UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

Electrical activity in the hippocampal formation of the rat : role of ascending monoamine-containing… Assaf, Souhile Y. 1978

Your browser doesn't seem to have a PDF viewer, please download the PDF to view this item.

Notice for Google Chrome users:
If you are having trouble viewing or searching the PDF with Google Chrome, please download it here instead.

Item Metadata

Download

Media
831-UBC_1978_A1 A88.pdf [ 24.55MB ]
Metadata
JSON: 831-1.0094477.json
JSON-LD: 831-1.0094477-ld.json
RDF/XML (Pretty): 831-1.0094477-rdf.xml
RDF/JSON: 831-1.0094477-rdf.json
Turtle: 831-1.0094477-turtle.txt
N-Triples: 831-1.0094477-rdf-ntriples.txt
Original Record: 831-1.0094477-source.json
Full Text
831-1.0094477-fulltext.txt
Citation
831-1.0094477.ris

Full Text

ELECTRICAL ACTIVITY IN THE HIPPOCAMPAL FORMATION OF THE RAT: ROLE OF ASCENDING MONOAMINE-CONTAINING SYSTEMS. by SOUHILE Y. ASSAF B.Sc, University of Western Ontario, 1974 M.Sc, University of Western Ontario, 1975 A THESIS SUBMITTED IN THE REQUIREMENTS DOCTOR OF PARTIAL FULFILLMENT OF FOR THE DEGREE OF • PHILOSOPHY in THE FACULTY OF GRADUATE STUDIES (Department of Physiology) We accept this thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA September, 1978 (c) Souhile Y. Assaf, 1978 In presenting th i s thes is in pa r t i a l fu l f i lment of the requirements for an advanced degree at the Univers i ty of B r i t i s h Columbia, I agree that the L ibrary sha l l make it f ree ly ava i lab le for reference and study. I further agree that permission for extensive copying of th is thesis for scholar ly purposes may be granted by the Head of my Department or by his representat ives. It is understood that copying or pub l i ca t ion of th is thesis fo r f inanc ia l gain sha l l not be allowed without my written permission. Department of The Univers i ty of B r i t i s h Columbia 2075 Wesbrook Place Vancouver, Canada V6T 1W5 Date ABSTRACT The e f f e c t s o f e l e c t r i c a l s t i m u l a t i o n o f t h e monoamine-c o n t a i n i n g n u c l e i , t h e median raphe (MR) and the l o c u s c o e r u l e u s ( L C ) , on e x t r a c e l l u l a r l y r e c o r d e d hippocampal e l e c t r i c a l a c t i v t y were s t u d i e d i n ur e t h a n e a n a e s t h e t i z e d r a t s . The f o l l o w i n g o b s e r v a t i o n s suggested t h a t s e r o t o n e r g i c and n o r a d r e n e r g i c systems o r i g i n a t i n g i n MR and LC, r e s p e c t i v e l y , modulate r h y t h m i c a l slow a c t i v i t y (RSA) r e c o r d e d i n the d e n t a t e g y r u s and/or t h e p o p u l a t i o n response of g r a n u l e c e l l s t o s t i m u l a t i o n of t h e p e r f o r a n t path (PP) i n p u t , 1. RSA (3-7 Hz,) r e c o r d e d i n the d e n t a t e g y r u s (DG) was r e l a t e d t o the b u r s t i n g d i s c h a r g e p a t t e r n of m e d i a l s e p t a l (MS) neurones. E l e c t r o l y t i c o r k a i n a t e l e s i o n s o f t h e MS a b o l i s h e d RSA t h e r e b y s u p p o r t i n g the c o n c l u s i o n t h a t r h y t h m i c a l hippocampal a c t i v i t y was i n i t i a t e d by MS neurones., 2. S t i m u l a t i o n o f MR r e s u l t e d i n d i s r u p t i o n o f t h e b u r s t i n g d i s c h a r g e o f MS neurones and d e s y n c h r o n i z a t i o n o f RSA. D e p l e t i o n o f f o r e b r a i n s e r o t o n i n f o l l o w i n g p r e t r e a t m e n t w i t h para-c h l o r o p h e n y l a l a n i n e (p-CPA) e l i m i n a t e d these r e s p o n s e s w h i l e g u i p a z i n e (1 mg/kg), a s e r o t o n i n a g o n i s t , mimicked t h e e f f e c t s o f MR s t i m u l a t i o n . These d a t a c o n f i r m e d t h a t a s e r o t o n i n -c o n t a i n i n g system mediates t h e d i s r u p t i o n of b u r s t i n g d i s c h a r g e i n MS and d e s y n c h r o n i z a t i o n o f hippocampal e l e c t r i c a l a c t i v i t y . 3. LC s t i m u l a t i o n evoked RSA. I n t r a c e r e b r a l i n j e c t i o n s o f 6-hydroxydopamine (6-OHDA) which d e p l e t e d hippocampal n o r a d r e n a l i n e (NA) d i d n o t e l i m i n a t e t h i s response. I n a d d i t i o n , i i i e l e c t r o l y t i c l e s i o n s of the LC d i d not e l i m i n a t e RSA. These d a t a suggested t h a t N A - c o n t a i n i n g a f f e r e n t s t o the hippocampal f o r m a t i o n a r e not e s s e n t i a l f o r t h e g e n e r a t i o n o f r h y t h m i c a l a c t i v i t y , 4. S t i m u l a t i o n o f t h e PP r e s u l t e d i n s y n a p t i c p o t e n t i a l s i n the d e n d r i t i c l a y e r of DG and p o p u l a t i o n s p i k e s r e c o r d e d i n t h e g r a n u l e c e l l body l a y e r . C o n d i t i o n i n g s t i m u l a t i o n o f ME o r LC i n c r e a s e d t h e a m p l i t u d e of t h e p o p u l a t i o n s p i k e evoked by a t e s t PP p u l s e w i t h o u t a l t e r i n g t h e a m p l i t u d e or r a t e o f r i s e o f the s y n a p t i c p o t e n t i a l . The e f f e c t s of MR and LC s t i m u l a t i o n were b l o c k e d by p-CPA and 6-0HDA, r e s p e c t i v e l y , s u g g e s t i n g t h a t these monoamine-containing n u c l e i s p e c i f i c a l l y i n f l u e n c e d t h e r e s p o n s i v e n e s s of d e n t a t e g r a n u l e c e l l s , 5. The o b s e r v a t i o n s t h a t changes i n the a m p l i t u d e of t h e p o p u l a t i o n s p i k e , such as t h o s e observed f o l l o w i n g s t i m u l a t i o n o f MR and LC, c o u l d o c c u r i n the absence of changes i n the s y n a p t i c p o t e n t i a l were c o n f i r m e d u s i n g an a d d i t i o n a l a f f e r e n t t o t h e d e n t a t e v i a the c o m m i s s u r a l p r o j e c t i o n . These data i n d i c a t e d t h a t e x t r i n s i c a f f e r e n t s t o DG may p o t e n t i a t e t h e d i s c h a r g e of a p o p u l a t i o n o f g r a n u l e c e l l s w i t h o u t enhancing s y n a p t i c p o t e n t i a l s . On the b a s i s o f t h e above d a t a , i t was c o n c l u d e d t h a t p h a r m a c o l o g i c a l l y d i s t i n c t n e u r o n a l systems o r i g i n a t i n g i n MR and LC modulate e l e c t r i c a l a c t i v i t y i n the hippocampal f o r m a t i o n . TABLE 01 -CONTENTS C e r t i f i c a t e Of Exam i n a t i o n ........................ i A b s t r a c t .......................................... i i Table o f c o n t e n t s . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , . i v Acknowledgements .............................. ... x i i Tab l e Of E g u i v a l e n t s And A b b r e v i a t i o n s ............. x i v L i s t Of T a b l e s .............................., x v i L i s t Of F i g u r e s ................................... x v i i 1 A 0 _ I n t r o d u c t i o n .................................. 1 1.1. Review Of The L i t e r a t u r e ...................... . 5 1.1.1 The Hippocampal Formation ................... 5 Major A f f e r e n t s ............................... 8 a. The P e r f o r a n t P a t h ...................... 8 b. C o m m i s s u r a l / A s s o c i a t i o n a l System ........ 11 c. S e p t a l - H i p p o c a m p a l P r o j e c t i o n ........... 12 d. Monoamine Pathways ...................... 18 Major Hippocampal E f f e r e n t s ................... 24 a. F i m b r i a - F o r n i x System 26 P l a s t i c i t y Of S y n a p t i c T r a n s m i s s i o n I n The Hippocampal F o r m a t i o n ...................... 27 1.1.2 The S e p t a l Area ............................ 30 Major A f f e r e n t s ............................... 32 a. Hippocampal I n p u t s . .... ................. 32 b. Hypothalamic I n p u t s ..................... 34 c. Amygdaloid I n p u t s .. .................. ... 35 d. Monoamine Pathways ....................... 35 1.1.3 R h y t h m i c a l A c t i v i t y I n The S e p t a l -Hippocampal A x i s ............................... 40 Spontaneous P a t t e r n s Of Hippocampal E l e c t r i c a l A c t i v i t y .................. ................. 41 Sources Of BSA ................................ 42 Ro l e Of A s c e n d i n g Systems I n The G e n e r a t i o n Of HS A ........................................ 44 Ro l e Of The S e p t a l Area I n The G e n e r a t i o n Of R SA ........... ....«.•.........*...........* . 47 Role Of P h a r m a c o l o g i c a l l y D i s t i n c t Systems I n The C o n t r o l Of S e p t a l - H i p p o c a m p a l A c t i v i t y . 49 a. A c e t y c h o l i n e ....................,........ 49 b. N o r a d r e n a l i n e ........................... 52 c. Dopamine 53 d. . S e r o t o n i n 54 e. Other T r a n s m i t t e r s ...................... 55 R e l a t i o n s h i p Of S e p t a l And Hippocampal A c t i v i t y To Behaviour ............................... 56 1.2 The P r e s e n t Study ............................ 61 0 Gen er a l Met ho ds .....,. ......*.....•..•..•.• • • .. . 6 3 2.1 S u r g i c a l P r e p a r a t i o n : ........................ 63 2.2 S t i m u l a t i n g And R e c o r d i n g P r o c e d u r e s ......... 64 S i n g l e U n i t A c t i v i t y And Evoked P o t e n t i a l s .,..,65 E l e c t r i c a l A c t i v i t y Of The Hippocampal F o r m a t i o n .................................. 67 2.3 Data A n a l y s i s : 69 S i n g l e U n i t A c t i v i t y .......................... 69 Evoked F i e l d s ••••.«.....*.................,.•. 71 Rh y t h m i c a l E l e c t r i c a l A c t i v i t y 74 2. 4 L e s i o n i n g T e c h n i q u e s ......................... 75 v i E l e c t r o l y t i c L e s i o n s . . . . . . . . . . . . . . . . . . . . . . . . . . 7 6 Acute And C h r o n i c T r a n s e c t i o n s ................ 7 6 N e u r o c h e m i c a l L e s i o n s . . . . . . . . . . . . . . . . . . . . . . . . . 7 8 2 . 5 Neurochemical Assays ......................... 8 0 2 . 6 H i s t o l o g i c a l A n a l y s i s ........................ 81 Chapter 3:„ Bole,Of The Septal^Area^In... The G e n e r a t j o g Of Hippocampal Electr^.gaj.^;A-cti,vity . . . . , . . . . . . . . . . . 8 2 3 . 1 I n t r o d u c t i o n 8 2 3 . 2 E x p e r i m e n t a l P r o c e d u r e s 8 3 3 . 3 R e s u l t s 8 4 Spontaneous E l e c t r i c a l A c t i v i t y Of The Hippocampus And The Dentate Gyrus .......... 8 4 S e p t a l U n i t A c t i v i t y . . . . . , - . . . . . . . . . . . . . . . . . . . . 9 0 Hippocampal Response P a t t e r n s A f t e r E l e c t r o l y t i c And K a i n i c a c i d L e s i o n s Of The S e p t a l Area . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 0 1 D i s c h a r g e P a t t e r n Of S e p t a l Neurones I n The I s o l a t e d F o r e b r a i n ......................... 1 0 9 3 . 4 D i s c u s s i o n . . . . . . , , * . . . . . . . . . . . , , . . . . . . . . . . . . . 11 5 D i s c h a r g e P a t t e r n Of S e p t a l Neurones . . . . . . . . . . 1 1 5 C o r r e l a t i o n s With M o r p h o l o g i c a l S t u d i e s ....... 1 1 6 R e g u l a t i o n Of Hippocampal A c t i v i t y ..,...,..,..117 3*5 SuHInicizry •••*••#*•••»•*•••#»•#•»••*•••*•••»*••# ^19 Chapter 4_JL Role__Qf_A_Ra£he-sero C o n t r o l .Of S e p t a l Hippocampal A c t i v i t y - . . . . . . . . . . . 1 2 3 4 . 1 I n t r o d u c t i o n • * * , , * 1 2 3 4 . 2 E x p e r i m e n t a l P r o c e d u r e s 1 2 4 4 . 3 B e s u l t s . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 2 6 v i i Response Of I-Neurones To Raphe S t i m u l a t i o n ...126 Response Of B-neurones To Raphe S t i m u l a t i o n ...132 E f f e c t Of Raphe S t i m u l a t i o n On Hippocampal E l e c t r i c a l A c t i v i t y 135 E f f e c t s Of p - c h l o r o p h e n y l a n i n e On S e p t a l -Hippocampal A c t i v i t y ....................... 140 E f f e c t s Of Q u i p a z i n e On Septal-Hippocampa1 A c t i v i t y . , 142 E f f e c t s Of 5 - h y d r o x y t r y p t o p h a n On Hippocampal E l e c t r i c a l A c t i v i t y ........................ 145 4.4 D i s c u s s i o n .................. ................. 148 E f f e c t Of MR S t i m u l a t i o n 0B I-neurones. .......148 E f f e c t Of MR S t i m u l a t i o n On B-neurones ........149 R e g u l a t i o n Of Hippocampal A c t i v i t y ............ 150 P o s s i b l e R o l e Of S e r o t o n i n .................... 151 4.5 Sum mary .......,.............................. 154 Chapter 5: —The •. - D o r s a l N o r a d r e n e r g i c System And Hippocampal E l e c t r i c a l A c t i v i t y .................. 157 5.1 I n t r o d u c t i o n ................................... 157 5.2 E x p e r i m e n t a l P r o c e d u r e s ......................158 5.3 R e s u l t s ........... ........................... 159 Spontaneous And Sensory Evoked P a t t e r n s Of E l e c t r i c a l A c t i v i t y . ......................,159 E f f e c t Of LC S t i m u l a t i o n On Hippocampal A c t i v i t y . 164 E f f e c t s Of 6-OHDA On RSA I n i t i a t e d By LC S t i m u l a t i o n .................... ........,..,169 E f f e c t s Of Amphetamine ,.....,,..,.,,..,,,..,,,172 5. 5 D i s c u s s i o n ...................................177 Does The D o r s a l NA Bundle Underly R SA ? .......177 A c t i o n Of Amphetamine ......................... 179 P o s s i b l e C o n t r i b u t i o n Of Urethane A n a e s t h e s i a .180 5.5 Summary .....181 Chapter 6: C h a r a c t e r i z a t i o n Of The P e r f o r a n t P a t h -d e n t a t e P r o j e c t i o n ............................... 182 6.1 I n t r o d u c t i o n 182 6.2 E x p e r i m e n t a l P r o c e d u r e s ......................187 6. 3 R e s u l t s .188 I d e n t i f i c a t i o n Of Dentate Responses. .......... 193 a. M o l e c u l a r L a y e r .......* ......193 b. C e l l Layer ..............................201 c. S i n g l e O n i t D i s c h a r g e ...................220 P o t e n t i a t i o n Of Dentate E x t r a c e l l u l a r Responses ........................ 230 a. P o p u l a t i o n EPSP .........................230 b. P o p u l a t i o n S p i k e 236 c. S i n g l e U n i t D i s c h a r g e ................... 241 6.4 D i s c u s s i o n ... 244 F i e l d A n a l y s i s Of The P e r f o r a n t P a t h I n p u t To The Dentate Gyrus ..........................244 Response Of S p o n t a n e o u s l y F i r i n g G - c e l l s . .....248 P o t e n t i a t i o n Of The E x t r a c e l l u l a r EPSP ........ 251 P o t e n t i a t i o n Of The P o p u l a t i o n Spike ..........252 QhSLR%SS.JI±^. Neurona 1 ,Transmission : I n ; The -Den t a t e Gyrus: R o l e Of The Commissural I n p u t ............. 256 7.1 I n t r o d u c t i o n •*.... ............... ,.-...*. , . . 256 ix 7.2 Experimental Procedures. ..................... 257 7. 3 Results 258 I d e n t i f i c a t i o n Of The Dentate Responses .......261 a. Molecular Layer ......................... 261 B. G-cell Layer ............................ 273 c. Single Unit Discharge ...................277 Origin Of The Commissural Pathway. ............ 286 Effects Of Commissural Stimulation On PP-evoked Responses .................................. 287 7.U Discussion .............................,.....298 Fi e l d Analysis Of The Commissural Input To The Dentate ....................................298 Effect Of Commissural Stimulation On PP Evoked Responses ............303 7. 5 Summary .. .... 305 Chap.ter 8 _i_ _ T he ai, Rap he- s er o ton i n S y stem • An d - N eurona .1 Transmission In The Dentate Gyrus ......,,......••306 8.1 Introduction .................................306 8.2 Experimental Procedures .....307 8.3 Results ...................................... 308 Effects Of MR Stimulation On G-cells ........,.308 Effects Of MB Stimulation On PP-evoked Responses .................................. 313 Relationship Between Effects Of MR Stimulation On Population Spike And Inh i b i t i o n Of Single Onits. .....................................320 Effects Of p-CPA On Respones Evoked By Stimulation Of MR 323 X 8. 1 D i s c u s s i o n *..325 E f f e c t s Of MB S t i m u l a t i o n On Spontaneous D i s c h a r g e Of G - c e l l s 327 E f f e c t s Of MB S t i m u l a t i o n On PP-evoked Responses 32 8 R e l a t i o n s h i p Between I n h i b i t i o n Of G - c e l l s And P o t e n t i a t i o n Of The P o p u l a t i o n S p i k e ..........329 R o l e F o r S e r o t o n i n : ...............329 8.5 Summary .......................... ............ 331 Chapter 9:. The_ L.ocus„_i-Coeruleus , And , , Neuronal T r a n s m i s s i o n . I n , The Dentate Gyrus ................332 9.1 I n t r o d u c t i o n ................................. 332 9.2 E x p e r i m e n t a l P r o c e d u r e s ...33 3 9.3 R e s u l t s ....333 E f f e c t s Of LC S t i m u l a t i o n On Dentate C e l l s ...,333 E f f e c t Of LC S t i m u l a t i o n On PP-evoked F i e l d Responses. .................................338 E f f e c t s Of 6-OHDA L e s i o n s Of The D o r s a l N o r a d r e n e r g i c Pathway ...................... 345 9.4 D i s c u s s i o n .................••................346 E f f e c t s Of LC S t i m u l a t i o n On G - c e l l D i s c h a r g e .346 E f f e c t Of LC S t i m u l a t i o n On PP-evoked Responses ....................................34 8 9.5 Summary ................................ .....,349 Chapter 10; G e n e r a l D i s c u s s i o n - . . . . . . . . . . . . . . . . . . . . . . 3 5 0 10,1 P a t t e r n s Of Hippocampal E l e c t r i c a l A c t i v i t y .................. .. ..... • . .. ....... .351 10.2 The S i g n i f i c a n c e Of Hippocampal P o p u l a t i o n x i Responses ...... .. .......354 10.3 Rhythmical A c t i v i t y And Neuronal Transmission In the Dentate Gyrus 358 References .......... • . 363 Vita ................................................400 x i i ACKNOWLEDGEMENTS The a u t h o r a p p r e c i a t e s t h e a s s i s t a n c e p r o v i d e d by t h e f o l l o w i n g d u r i n g p r e p a r a t i o n o f t h i s t h e s i s ; Dr. J . J . M i l l e r f o r the s u p e r v i s i o n , c r i t i c i s i m s and a s s i s t a n c e t h a t he p r o v i d e d t h r o u g h o u t t h e r e s e a r c h . Mrs. Helen B r a n d e j s who p r o v i d e d e x c e l l e n t t e c h n i c a l a s s i s t a n c e a t a l l s t a g e s o f t h e ex p e r i m e n t s and i n p a r t i c u l a r t he s p e c t r o f l u o r o m e t r i c a s s a y s . Mr. Tom R i c h a r d s o n f o r t h e f i l e s and d e v i c e s which made the computer a n a l y s i s p o s s i b l e . Mrs. Margaret S t u e r z l Smith f o r t h e r a d i o e n z y m a t i c a s s a y s f o r n o r a d r e n a l i n e . Ms. L i z . Barbour f o r a s s i s t a n c e i n t h e expe r i m e n t s r e p o r t e d i n Chapter 5. Messrs Kurt Henze and Ralph A s s i n a f o r r e p r o d u c t i o n o f the i l l u s t r a t i o n s and t h e i r generous t e c h n i c a l a s s i s t a n c e . Dr. Stephen Mason f o r p r o v i d i n g some o f t h e l e s i o n e d r a t s used i n C h a pter 9 and f o r r e a d i n g the t h e s i s . Drs. F. L i o y , H. McLennan And P, Vaughan formed my a d v i s o r y committee and read the t h e s i s , Drs. H,C. F i b i g e r , J.A, Pearson And A.G. P h i l l i p s f o r a nswering a l o t of g u e s t i o n s and p r o v i d i n g s u p p l i e s and f a c i l i t i e s whenever r e g u i r e d . Mr. Harry Kohne and Department of P h y s i o l o g y t e c h n i c a l s t a f f f o r b u i l d i n g c r u c i a l hardware. Messrs P h i l H i c k s , Don R u t h e r f o r d , Ron S k e l t o n , John W i l c o x and Dr. N e i l McNaughton f o r r e a d i n g e a r l y d r a f t s o f the t h e s i s and p r o v i d i n g v a l u a b l e d i s c u s s i o n s . Mrs. . Hary F o r s y t h And Mr. James Loo f o r t y p o g r a p h i c a l and s e c r e t a r i a l a s s i s t a n c e . The Research Was Funded By MRC Of Canada. DECLARATION The data p r e s e n t e d were o b t a i n e d between Sept, 1975 and Sept. 1978 when the a u t h o r was i n t r a i n i n g i n the Department of P h y s i o l o g y , U n i v e r s i t y o f B r i t i s h Columbia. Becords which b e s t i l l u s t r a t e c e r t a i n o b s e r v a t i o n s were s e l e c t e d . P o l y g r a p h r e c o r d s but not o s c i l l o s c o p e photographs or computer p l o t s were repro d u c e d by t h e I n s t r u c t i o n a l Media C e n t r e a t U.B.C. And were r e t o u c h e d a t t h e i r d i s c r e t i o n where n e c e s s a r y . The t e x t of the t h e s i s was composed u s i n g FMT documentation s o f t w a r e a v a i l a b l e on the U.B.C. computing system. x i v l i l k E OF EQUIVALENTS AND ABBREVIATIONS Eg.ivalents hippocampus = amnions horn = hippocampal f i e l d s CA1-4 d e n t a t e gyrus = f a s c i a d e n t a t a = d e n t a t e hippocampal f o r m a t i o n = hippocampus + de n t a t e gyrus p o t e n t i a t i o n = f a c i l i t a t i o n = enhancement o f t e s t r e s p o n s e s Ef§Suently Osed A b b r e v i a t i o n s AP a n t e r i o r - p o s t e r i o r s t e r e o t a x i c c o - o r d i n a t e B-neurone s e p t a l neurone which d i s c h a r g e s i n b u r s t s CNS c e n t r a l nervous system COMM hippocampal co m m i s s u r a l pathway DA dopamine DO d e n t a t e gyrus EC e n t o r h i n a l c o r t e x EPSP e x c i t a t o r y p o s t s y n a p t i c p o t e n t i a l G - c e l l d e n t a t e g r a n u l e c e l l HRP h o r s e r a d i s h p e r o x i d a s e I-neurone s e p t a l neurone d i s p l a y i n g i r r e g u l a r d i s c h a r g e L m e d i a l - l a t e r a l s t e r e o t a x i c c o - o r d i n a t e LC l o c u s c o e r u l e u s MF mossy f i b r e pathway MS m e d i a l s e p t a l n u c l e u s MR median raphe n u c l e u s NA n o r a d r e n a l i n e ( n o r e p i n e p h r i n e ) PP p e r f o r a n t path RSA r h y t h m i c a l slow a c t i v i t y X V p-CPA p-chlorophenylalanine 5- .HT 5-hydroxytryptan.ine (serotonin) 6- OHDA 6-hydroxydopamine x v i LIST OP TABLES Ta b l e I : Comparison Of S e p t a l Neurones I n f l u e n c e d By MR S t i m u l a t i o n In C o n t r o l And p-CPA-Treated Rats.,,,....................................... 139 Ta b l e I I : E f f e c t s Of C o n d i t i o n i n g S t i m u l a t i o n Of The Commissural Pathway On PP-evoked P o p u l a t i o n S p i k e s . . ........ .........................,,, 290 Ta b l e I I I : E f f e c t s Of p-CPA On Hippocampal 5-HT And Responses To S t i m u l a t i o n Of The Median Raphe. 326 x v i i Fig. 1- 1: Drawing Of The I n t r i n s i c Synaptic Organization Of The Hippocampal Formation..... 8 Fig. 2- 1: Schematic I l l u s t r a t i o n Of Ext r a c e l l u l a r Recording Technique. ................69 Fig. 2- 2: Measurements Of Amplitudes And Sate Of Rise Of Evoked F i e l d Potentials.................... 73 Fig. 3-1: Patterns Of E l e c t r i c a l A c t i v i t y Recorded From The Dentate Gyrus Of A Urethans Anaesthetized Rat. ................86 Fig, 3- 2: Location Of Recording Electrode In The Dentate Gyrus,••«..••..»«.<•••...........,,,.. 89 Fig. 3- 3: Differences In The Discharge Patterns Of Septal Neurones Recorded During Rhythmical Or Desynchronized Hippocampal A c t i v i t y . . . . . . . . 92 Fig. 3- 4: Localization Of B-neurones In The Septal A r e a . . . . . . . a , . . . ' , , , . . . * . . . . . . . » * . * , , ' « . , # * , , , ,.94 x v i i i F i g . 3- 5: C h a r a c t e r i s t i c s Of A S e p t a l B-neurone......... 97 F i g , 3- 6: B u r s t i n g D i s c h a r g e P a t t e r n Of S e p t a l Neurones F o l l o w i n g Hippocampal S t i m u l a t i o n . . . . 100 F i g , 3- 7: E f f e c t s Of E l e c t r o l y t i c And K a i n a t e L e s i o n s Of The S e p t a l Area On Hippocampal E l e c t r i c a l A c t i v i t y , ...103 F i g . 3- 8: E x t e n t Of E l e c t r o l y t i c And K a i n i c - i n d u c e d L e s i o n s I n The S e p t a l Area. 106 F i g , 3- 9: Neuronal D e g e n e r a t i o n F o l l o w i n g K a i n i c I n j e c t i o n s I n t o The S e p t a l Area. ....108 F i g . 3-11: D i s c h a r g e P a t t e r n s Of S e p t a l Neuronas I n The D e a f f e r e n t e d F o r e b r a i n . . . . . . . . . . . . . . . . . 114 F i g . 3-12: Schematic I l l u s t r a t i o n Of The Proposed R e l a t i o n s h i p Between The B r a i n s t e m , S e p t a l Area And The Hippocampal F o r m a t i o n . . . . . . . . . . . . 121 x i x F i g . 4 - 1 : Response Of S e p t a l I-neurones To Raphe S t i m u l a t i o n . .128 F i g . 4- 2: L o c a l i z a t i o n Of S t i m u l a t i n g E l e c t r o d e s I n The Region Of The Raphe And The Co r r e s p o n d i n g Response Of I-neurones.......... 131 F i g . 4- 3: C h a r a c t e r i s t i c s Of A B-neurone During S i n g l e P u l s e S t i m u l a t i o n Of MR........... 134 F i g . 4- 4: E f f e c t s Of R e p e t i t i v e MB S t i m u l a t i o n On The B u r s t i n g D i s c h a r g e P a t t e r n Of B-neurones I n C o n t r o l And p-CPA T r e a t e d R a t s . . . . . . . . . . . . . . . . 137 F i g . 4- 5: E f f e c t s Of R e p e t i t i v e MR S t i m u l a t i o n On Hippocampal E l e c t r i c a l A c t i v i t y I n C o n t r o l And p-CPA T r e a t e d r a t s . . . . . . . . . . . . . . . . . . . . . . . 141 F i g . 4 - 6 : E f f e c t s Of Q u i p a z i n e On Hippocampal E l e c t r i c a l A c t i v i t y , . . . . . . . . . . . . . , . . . . . , . . . . . , 1 4 4 F i g . 4- 7: E f f e c t s Of 5-HT On Hippocampal E l e c t r i c a l A c t i v i t y . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147 F i g . 4 - 8 : A Schematic I l l u s t r a t i o n Of The Proposed S y n a p t i c Arrangements Between The MR, Septum, And Hippocampal Formation.............156 X X F i g . 5- 1: R h y t h m i c a l E l e c t r i c a l a c t i v i t y Recorded I n The Dentate Gyrus Of C o n t r o l And T r e a t e d R a t s . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161 F i g . 5 - 2 : Freguency Of Spontaneously O c c u r r i n g And E l i c i t e d RSA 163 F i g . , 5- 3: E f f e c t s Of S t i m u l a t i o n I n The Region Of The Locus C o e r u l e u s On E l e c t r i c a l A c t i v i t y Of The Dentate Gyrus............................. 167 F i g . 5- 4: L o c a t i o n Of S t i m u l a t i n g E l e c t r o d e s I n The Region Of LC And I n t e n s i t y T h r e s h o l d F o r The G e n e r a t i o n Of RSA........................ 171 Fig.... 5- 5: E l e c t r i c a l A c t i v i t y Recorded I n The Dentate Gyrus Of A 6-OHDA T r e a t e d R a t . . . . . . . . . . . . . . . . 174 F i g . 5 - 6 : Freguency Of RSA As A F u n c t i o n Of LC S t i m u l u s I n t e n s i t y I n C o n t r o l And 6-OHDA L e s i o n e d R a t s . 176 F i g , , 5 - 7 : E f f e c t s Of Amphetamine On E l e c t r i c a l A c t i v i t y Of The Dentate Gyrus. 184 x x i F i g . 6- 1: Laminar O r g a n i z a t i o n Of A f f e r e n t s To The Dentate. 184 F i g . 6 - 2 : Depth P r o f i l e s Of The F i e l d P o t e n t i a l s Evoked I n The Dentate Gyrus By A Weak P e r f o r a n t P a t h V o l l e y 190 F i g . 6 - 3 : F i e l d P o t e n t i a l s Evoked I n The Dentate By PP S t i m u l a t i o n A t V a r i o u s I n t e n s i t i e s 19 2 F i g . . 6- 4: I d e n t i f i c a t i o n Of The F i e l d P o t e n t i a l s Recorded In The Dentate M o l e c u l a r L a y e r . . 19 5 F i g . , 6- 5: R e l a t i o n s h i p Between PP S t i m u l u s I n t e n s i t y And A m p l i t u d e Of Responses Recorded I n The D e n d r i t i c Region Of The Dentate, 197 F i g . 6- 6: I d e n t i f i c a t i o n Of The Evoked P o t e n t i a l Recorded At G - c e l l Body L a y e r . 200 F i g . 6- 7: Development Of The Dentate P o p u l a t i o n S p i k e . 20 3 x x i i F i g . 6- 8: E f f e c t s Of S t i m u l u s I n t e n s i t y On A m p l i t u d e And L a t e n c y Of The G - c e l l L a y e r r e s p o n s e 20 5 F i g . 6- 9: The C e l l L a y e r Response Recorded At V a r i o u s Depths I n The D e n t a t e . 20 8 F i g . 6-10: P r e s y n a p t i c And P o s t s y n a p t i c Responses F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t P a t h . 210 F i g , 6-11: A n t i d r o m i c And Orthodromic A c t i v a t i o n Of Dentate G r a n u l e C e l l s . 213 F i g . 6-12: I d e n t i f i c a t i o n Of The L a t e N e g a t i v e Component (N3) Recorded I n The Dentate F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t Path. 216 F i g . 6-13: D i s c h a r g e P a t t e r n Of Dentate Granule C e l l s . 219 F i g . 6-14: The Response Of An I d e n t i f i e d Dentate G r a n u l e C e l l To S t i m u l a t i o n Of The P e r f o r a n t P a t h . 222 F i g , 6-15: I d e n t i f i c a t i o n Of Dentate Neurones., 224 x x i i a Leaf x x i i i does not e x i s t . x x i v F i g . 6-16: Response Of H i l a r Neurones F o l l o w i n q S t i m u l a t i o n Of The P e r f o r a n t P a t h And The Mossy F i b r e s . 227 F i g . 6-17: P a i r e d P u l s e P o t e n t i a t i o n Of The E x t r a c e l l u l a r EPSP. 229 F i g . 6-18: S t a t i s t i c a l A n a l y s i s Of EPSP P o t e n t i a t i o n . 232 F i g . 6-19: P a i r e d - p u l s e P o t e n t i a t i o n Of The EPSP And The P o p u l a t i o n S p i k e Response., 238 F i g . 6-20: E f f e c t s Of S t i m u l u s I n t e n s i t y On P a i r e d -p u l s e P o t e n t i a t i o n Of The P o p u l a t i o n S p i k e . 240 F i g . 6-21: P o t e n t i a t i o n Of G - c e l l D i s c h a r g e By P a i r e d P u l s e S t i m u l a t i o n Of The P e r f o r a n t Path. 243 F i g . 6-22: I n t e r p r e t a t i o n Of The Responses Recorded I n The Dentate F o l l o w i n g PP S t i m u l a t i o n . 248 F i g . 7- 1: Laminar P r o f i l e s Of The F i e l d P o t e n t i a l s Evoked In The Dentate By S t i m u l a t i o n Of The P e r f o r a n t P a t h And Commissural Pathway. 260 X X V F i g . , 7- 2: Laminar P r o f i l e s Of P r i m a r y And Secondary F i e l d P o t e n t i a l s Recorded I n The Dentate F o l l o w i n g S t i m u l a t i o n Of The COM Pathway. 264 F i g . 7- 3: C h a r a c t e r i s t i c s Of E a r l y And L a t e F i e l d Responses Recorded I n The Dentate F o l l o w i n g S t i m u l a t i o n Of The Commissural Pathway. 266 F i g . 7- 4: F i e l d P o t e n t i a l s Recorded I n The G r a n u l e C e l l L a y e r F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t P a t h And Commissural Pathways. 273 F i g . 7- 5; Response Of Spontaneously F i r i n g D e n t a t e Neurones F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t Path And Commissural I n p u t . 279 F i g , , 7- 6: E f f e c t s Of Commissural S t i m u l a t i o n On Dentate F i e l d Responses Evoked By A T e s t P e r f o r a n t Path V o l l e y . 289 F i g . 8- 1: E f f e c t s Of MB S t i m u l a t i o n On Dentate G r a n u l e C e l l s . , 310 F i g . 8- 2: I n h i b i t i o n Of S pontaneously F i r i n g Dentate C e l l s By MR S t i m u l a t i o n . 312 x x v i F i g , , 8- 3: R e l a t i o n s h i p Between I n h i b i t i o n Of Dentate C e l l s And P o s i t i o n Of The S t i m u l a t i n g E l e c t r o d e In The Region Of MR. 315 F i g , 8 - 4 : E f f e c t s Of MR S t i m u l a t i o n On PP Evoked A c t i v a t i o n Of Dentate C e l l s . 317 F i g , 8- 5: E f f e c t s Of MR C o n d i t i o n i n g P u l s e s On The F i e l d Responses Evoked By The P e r f o r a n t P a t h . 319 F i g . 8- 6: F a c i l i t a t i o n Of The P o p u l a t i o n Spike Response F o l l o w i n g PP And MR C o n d i t i o n i n g P u l s e s . 322 F i g . 8- 7: R e l a t i o n s h i p Between I n h i b i t i o n Of G - c e l l s And A m p l i t u d e Of The P o p u l a t i o n S p i k e . 325 F i g , 9- 1: E f f e c t s Of LC S t i m u l a t i o n On Dentate Neurones. 335 F i g . 9- 2: E f f e c t s Of LC S t i m u l a t i o n On PP-evoked Dentate P o p u l a t i o n Responses. 340 F i g . 9- 3: R e l a t i o n s h i p Between The Number Of LC P u l s e s And A m p l i t u d e Of The P o p u l a t i o n S p i k e . 342 xx v i i F i g . 9 - 4 : H i s t o l o g i c a l L o c a l i z a t i o n Of S t i m u l a t i o n E l e c t r o d e I n LC. 342. 1 F i g . 9- 5; E f f e c t s Of 6-OHDA On The Response To LC S t i m u l a t i o n . 345 F i g , 10- 1: Proposed E x t r i n s i c I n f l u e n c e s On Neuronal T r a n s m i s s i o n I n The Dentate Gyrus. 360 1 JL 0 IlJIROrjJLJGTIQN The foundations of contemporary neuroscience are based on the assumption that the nervous system i s composed of i n d i v i d u a l nerve c e l l s that generate e l e c t r i c a l a c t i v i t y which, by chemical processes, i s transmitted to adjacent neurones at specialized regions c a l l e d 'synapses' (Langley, 1905; Sherrington,1906; Cajal, 1911; Dale, Peldberg and Vogt, 1936; Eccles, Katz and K u f f l e r , 1941) . /Although these concepts are now firmly established, new modes of synaptic organization including chemical and e l e c t r i c a l junctions between dendrites, somata and axons have extended this view of the nervous system, The mechanisms by which information i s transmitted from one region of the CNS to another and the i n t r i n s i c synaptic organization of the various nuclei are the concerns of systems neurophysiology. The methodology of the neurosciences has p a r a l l e l l e d these developments r e s u l t i n g i n a combination of electrophysiological, pharmacological and anatomical technigues. The use of microelectrodes for recording i n t r a c e l l u l a r and e x t r a c e l l u l a r e l e c t r i c a l a c t i v i t y of s i n g l e c e l l s combined with analysis of slow waves generated by populations of neurones has provided information about the synaptic organization of major neuronal systems. In order to understand the role of these systems i n the o v e r a l l 2 p a t t e r n o f b r a i n f u n c t i o n e l e c t r o p h y s i o l o g i c a l d a t a must be r e l a t e d t o b o t h a n a t o m i c a l and b i o c h e m i c a l d a t a . T h i s i n t e g r a t i v e approach i s p o s s i b l e i n t h e s t u d y of t h e hippocampal f o r m a t i o n which has a r e l a t i v e l y s i m p l e and w e l l d e f i n e d s y n a p t i c o r g a n i z a t i o n . I t s a f f e r e n t s t e r m i n a t e i n d i s c r e t e l a y e r s or s u b f i e l d s a l l o w i n g c o r r e l a t i o n o f n e u r o a n a t o m i c a l d a t a w i t h the c h a r a c t e r i s t i c f e a t u r e s o f i t s e l e c t r i c a l a c t i v i t y (Adey and Meyer, 1952; Green and A r d u i n i , 1954; G l o o r , Vera and S p e r t i , 1964; Andersen, E c c l e s and L o y n i n g , 1964; Lomo, 1971a). The spontaneous slow wave p a t t e r n s of t h e hippocampal f o r m a t i o n , p a r t i c u l a r l y r h y t h m i c a l slow a c t i v i t y (RSA; t h e t a r h y t h m ) , have been s t u d i e d most e x t e n s i v e l y and r e l a t e d t o t h e a c t i v i t y o f s i n g l e u n i t s i n t h e hippocampus and the s e p t a l a r e a (Jung and K o r n m u l l e r , 1938; Green and A r d u i n i , 1954; von E u l e r and Green, 1960; Kandel and Spencer, 1961; P e t s c h e , Stumpf and Gogolak, 1962)., I n a d d i t i o n t o the spontaneous p a t t e r n s o f hippocampal a c t i v i t y , c h a r a c t e r i s t i c e x t r a c e l l u l a r f i e l d p o t e n t i a l s are evoked by s t i m u l a t i o n o f t h e e n t o r h i n a l c o r t e x ( G l o o r e t a l , 1964; Lomo, 1971a). These f i e l d r e s p o n s e s are p o t e n t i a t e d by r e p e a t e d s t i m u l a t i o n and have t h u s been used t o s t u d y p r o c e s s e s a s s o c i a t e d w i t h n e u r o n a l t r a n s m i s s i o n and s t o r a g e o f i n f o r m a t i o n (Lomo, 197 1b; 3 B l i s s and L o i o , 1973; Douglas and Goddard, 1975; Steward, White, Cotman and Lynch, 1976). E a r l y s t u d i e s have proposed t h a t hippocampal e l e c t r i c a l a c t i v i t y i s c o n t r o l l e d by n e u r o n a l systems o r i g i n a t i n g i n d i f f u s e r e g i o n s o f t h e b r a i n s t e m (Green and A r d u i n i , 1954). While the p r e c i s e n e u r o a n a t o m i c a l and n e u r o c h e m i c a l o r i g i n s o f t h e s e systems a r e not known, the r e c e n t development of h i s t o c h e m i c a l t e c h n i g u e s f o r mapping p h a r m a c o l o g i c a l l y d i s t i n c t systems has r e v e a l e d t h a t the hippocampal f o r m a t i o n r e c e i v e s d i r e c t s e r o t o n i n - and n o r a d r e n a l i n e - c o n t a i n i n g p r o j e c t i o n s from the median raphe n u c l e u s and t h e l o c u s c o e r u l e u s , r e s p e c t i v e l y ( F a l c k , H i l l a r p , Thieme and Torp, 1962; Fuxe, 1965; U n g e r s t e d t , 1971; Hoore, 1975). S i n c e t h e s e monoamine systems a r e w i t h i n the d i f f u s e b r a i n s t e m r e g i o n s which modulate c o r t i c a l and hippocampal ' a r o u s a l 1 ( M o r u z z i and Magoun, 1949; Green and A r d u i n i , 1954; L i n d s l e y , 1960) , they may be i n v o l v e d i n d e t e r m i n i n g t h e c h a r a c t e r i s t i c f e a t u r e s of hippocampal e l e c t r i c a l a c t i v i t y . The p r e s e n t study examines t h e r o l e o f the a s c e n d i n g monoamine systems i n the c o n t r o l of slow e l e c t r i c a l a c t i v i t y and evoked f i e l d p o t e n t i a l s r e c o r d e d i n the hippocampal f o r m a t i o n of the r a t . The aim o f t h e e x p e r i m e n t s has been t o d e l i n e a t e t h e mechanisms by which t h e s e e x t r i n s i c i n p u t s i n f l u e n c e t h e spontaneous p a t t e r n s of r h y t h m i c a l hippocampal 4 a c t i v i t y and neuronal t r a n s m i s s i o n between the e n t o r h i n a l cortex and the dentate qyrus. 5 Ixli.- OF THE LITE8ATURE Ji1..1 The H i £ £ C c a m p a l Format i on The hippocampal f o r m a t i o n i s a horn shaped c o r t i c a l a r ea l y i n g a d j a c e n t t o t h e w a l l o f the l a t e r a l v e n t r i c l e s . T h i s r e g i o n i s s u b d i v i d e d i n t o the hippocampus p r o p e r , t h e d e n t a t e gyrus ( F a s c i a Dentata) and t h e s u b i c u l a r complex. The hippocampus proper can be f u r t h e r s u b d i v i d e d i n t o 4 f i e l d s <CA1, CA2, CA3 and CA4) i n which the major c e l l type i s the p y r a m i d a l c e l l ( L o r e n t e de No, 1934; B l a c k s t a d , 1956). The CA4 c e l l l a y e r e n t e r s t h e h i l u s o f t h e d e n t a t e gyrus which i s formed by a c u r v e d sheet of d e n t a t e g r a n u l e c e l l s . Both t h e hippocampal p y r a m i d a l c e l l s and d e n t a t e g r a n u l e c e l l s form w e l l d e f i n e d l a y e r s . As shown i n F i g . 1-1, t h e s e l a y e r s c o n t a i n a l m o s t e x c l u s i v e l y e i t h e r p y r a m i d a l c e l l b o d i e s {Stratum P y r a r a i d a l e ) , the d e n d r i t e s of p y r a m i d a l c e l l s {Stratum O r i e n s , Radiatum and Lacunosum), d e n t a t e g r a n u l e c e l l b o d i e s or the d e n d r i t e s o f g r a n u l e c e l l s ( Stratum M o l e c u l a r e ) . I n a d d i t i o n t o the l o n g axoned p y r a m i d a l and g r a n u l e c e l l s many c e l l t y p e s h a v i n g s h o r t axons can be found i n the hippocampal f o r m a t i o n . The l a t t e r neurones, which may be i n t r i n s i c to t h e hippocampal f o r m a t i o n , i n c l u d e G o l g i t y p e I I (basket) c e l l s which appear t o c o n t a c t numerous p y r a m i d a l and g r a n u l e c e l l s ( C a j a l , 1968). I n a d d i t i o n t o b a s k e t c e l l s t h e r e a r e s e v e r a l l a y e r s of 6 p o l y m o r p h i c c e l l s ( S t r a t a Polymorphe) or m o d i f i e d p y r a m i d a l c e l l s i n the d e n t a t e h i l u s ( L o r e n t e de No, 1934) . As shown i n F i g . 1 - 1 , t h e axons o f g r a n u l e c e l l s form the mossy f i b r e system which c o n t a c t s the d e n d r i t e s of CA3 c e l l s ( C a j a l , 1968; B a r b e r , Vaughan and Himer, 1974). The axons o f CA3 c e l l s p r o j e c t i n t o t h e f i m b r i a g i v i n g o f f S c h a e f f e r c o l l a t e r a l s which r e a c h t h e b a s a l d e n d r i t e s of CA1 and CA2 p y r a m i d a l c e l l s whose axons, i n t u r n , form t h e a l v e u s . T h i s s y n a p t i c o r g a n i z a t i o n s u g g e s t s t h a t any a f f e r e n t s which i n f l u e n c e d e n t a t e g r a n u l e c e l l s would s e g u e n t i a l l y a f f e c t CA3 and CA1 c e l l s a l o n g t h i s t r i - s y n a p t i c l o o p . The p r e c i s e l a m i n a r o r g a n i z a t i o n o f t h i s n e u r o n a l l o o p w i l l become more apparent f o l l o w i n g an e x a m i n a t i o n of t h e a f f e r e n t s t o t h e hippocampal f o r m a t i o n . H§-3°£ A f f e r e n t s A A 1'iie Pg£f 2£ant Path The major a f f e r e n t p r o j e c t i o n t o t h e hippocampal f o r m a t i o n i s from the e n t o r h i n a l c o r t e x by way of the p e r f o r a n t path (PP) f i b r e system (Lorente de No, 193 4) . The a n a t o m i c a l s t u d i e s of Hjorth-Simonsen (1973) demonstrate t h a t t h e l a t e r a l e n t o r h i n a l r e g i o n i s the s o u r c e o f PP f i b r e s which t e r m i n a t e i n the o u t e r p o r t i o n o f t h e d e n t a t e m o l e c u l a r l a y e r w h i l e t h e m e d i a l 7 1- I i Drawing o f The I n t r i n s i c Sy.nap.tiS O r g a n i z a t i o n Of The HipEOcamjaal Formatio|n Aj, t r a n s v e r s e s e c t i o n showing p y r a m i d a l c e l l l a y e r s CA1-CA4 and g r a n u l e c e l l s i n t h e d e n t a t e gyrus (DG). Axons o f g r a n u l e c e l l s form mossy f i b r e s (MF) which p r o j e c t t o CA3. P r o j e c t i o n s from CA3 are v i a t h e f i m b r i a t o t h e s e p t a l a r e a and by S c h a e f f e r c o l l a t e r a l s (Sch) onto d e n d r i t e s o f CA2 and CA1. Bjj, d e t a i l of t h e v a r i o u s l a y e r s from d o r s a l s u r f a c e of CA1 p y r a m i d a l c e l l t o t h e l o w e r blade o f the d e n t a t e g y r u s . The hippocampal f i s s u r e (HF) i s t h e d e m a r c a t i o n zone. T h i s f i g u r e i s m o d i f i e d from Green (1964) and C a j a l (1911). B . Alveus ^ S Oriens v s- Pyramidale I s . Radiatum A s- Lacunosum HF 1/ s Moleculare Q ~ " S. "Granufosum MF S. Polymorphe OO 9 e n t o r h i n a l r e g i o n g i v e s r i s e t o PP f i b r e s t h a t t e r m i n a t e i n t h e m i d d l e m o l e c u l a r l a y e r o f t h e d e n t a t e . , E l e c t r o n m i c r o s c o p i c i n v e s t i g a t i o n s suggest t h a t t h e mode of t e r m i n a t i o n i s p r i m a r i l y en £§ssaae onto d e n d r i t i c s p i n e s of d e n t a t e g r a n u l e c e l l s ( N a f s t a d , 1967). These a n a t o m i c a l d a t a have been c o n f i r m e d by e l e c t r o p h y s i o l o g i c a l a n a l y s i s o f f i e l d p o t e n t i a l s r e c o r d e d i n t h e d e n t a t e f o l l o w i n g s t i m u l a t i o n o f the PP or t h e e n t o r h i n a l c o r t e x i t s e l f ( G l o o r , Vera and S p e r t i , 1964; Lomo, 1971a; Steward, H h i t e , Cotman, and Lynch, 1976). The r e l a t i o n s h i p s between th e s e e x t r a c e l l u l a r p o t e n t i a l s and i n t r a c e l l u l a r r e c o r d i n g s of s i n g l e u n i t a c t i v i t y i n d i c a t e d a monosynaptic e x c i t a t o r y PP i n p u t onto the d e n d r i t e s of g r a n u l e c e l l s . The potency o f t h i s i n p u t i s r e f l e c t e d by the synchronous a c t i v a t i o n o f a p o p u l a t i o n of g r a n u l e c e l l s by s i n g l e p u l s e s t i m u l a t i o n o f t h e PP (Andersen, B l i s s and Skrede, 1971). Furthermore, t h e i n p u t from t h e PP t o t h e d e n t a t e i s o r g a n i z e d i n p a r a l l e l s t r i p s o r l a m e l l a e such t h a t s t i m u l a t i o n of a d i s c r e t e p a r t o f PP e x c i t e s g r a n u l e c e l l s o n l y i n a t h i n band o f t i s s u e r o u g h l y t r a n s v e r s e t o the l o n g i t u d i n a l a x i s of the hippocampus (Lomo,1971a). From r e c o r d i n g s o f s i n g l e u n i t a c t i v i t y i t appears t h a t an a f f e r e n t v o l l e y e x c i t e s a p o p u l a t i o n of g r a n u l e e e l s w i t h i n a l a m e l l a (Andersen e t a l . 10 1971;). Andersen, 1975)., Following the s i n g l e spike activation of granule c e l l s produced by PP stimulation, a long period of i n h i b i t i o n i s recorded (Lomo, 1971a). This i n h i b i t i o n i s s i m i l a r to that reported by Spencer and Kandel (1961) in CA1 c e l l s i n that nearly a l l c e l l s are i n h i b i t e d independent of whether they were previously activated by the stimulus. As w i l l be described i n a l a t e r section, t h i s i n h i b i t i o n i s postulated to r e s u l t from a recurrent i n h i b i t o r y mechanism (Gloor, 1963; Andersen et a l , 1964). In addition to the massive PP projection to the dentate a lesser number of PP f i b r e s are known to terminate in CA3 (Hjort h-Simonsen and Jeune, 1972; Hjorth-Simonsen, 1973) and possibly i n CA1 (Baisman, Cowan and Powell, 1965) subfields of the hippocampus., However, the sparse degeneration i n CA1 following lesions of the entorhinal cortex may be due to degeneration of f i b r e s en route to CA 3 (Hjorth-Simonsen, 1973). Electrophysiological evidence tends to support these anatomical data since monosynaptic potentials are recorded i n CA3 following stimulation of the PP (Gloor et a l , 1964), In contrast, the f i e l d s recorded i n CA1 have substantially smaller amplitudes and longer latencies than those recorded in the dentate or CA3 and may therefore be mediated through th t r i s y n a p t i c c i r c u i t r y described previously (Andersen, 1975; Winson and Abzug, 1978)., 11 5A. ••Commissural/Associational System The commissural (COMM) connections of the hippocampus have been mapped out using degeneration (Blackstad, 1956}, autoradiographic (Gottlieb and Cowan, 1973) and electrophysiological technigues (Andersen et a l , 1961; Deadwyler, White, Cotman and Lynch, 1975). Whereas a l l f i e l d s of the hippocampus and dentate gyrus receive COMM inputs, the o r i g i n of COMM fibr e s i s r e s t r i c t e d to CA3-CA4 of the con t r a l a t e r a l hippocampus (Gottlieb and Cowan, 1973; Hjorth-Simonsen and Laurberg, 1978). The large myelinated COMM f i b r e s leave the c o n t r a l a t e r a l hippocampus via the alveus proximal to the i n f e r i o r blade of the dentate and project through the ventral psalterium, re-entering the hippocampus near CA3. In the dentate, the COMM input i s r e s t r i c t e d to the inner molecular layer below the PP synapses, Remarkably, there i s no overlap between the entorhinal and COMM system. However, the same zone that i s innervated by the COMM input i s also the zone of termination of the associationa1 system originating in i p s i l a t e r a l CA3c (Simmer, 1970)., The si g n i f i c a n c e of the overlap between the termination of COMM and associational f i b r e s a r i s i n g from c o n t r a l a t e r a l and i p s i l a t e r a l CA3, respectively, i s not yet clear. There are few detailed electrophysiological studies of these projection systems and those available are discussed in 12 more d e t a i l i n Chapter 7, The COMM input to CA1 terminates on apical and basal dendrites of pyramidal c e l l s (Andersen, 1960; Gottlieb and Cowan, 1973). The input onto a p i c a l dendrites i s more massive as indicated by the dense degeneration following COfiM transections (Blackstad, 1958) and a larger negative wave evoked by stimulation of the COHM pathway (Cragg and Hamlyn, 1957). The s i m i l a r i t i e s between the synaptic wave evoked by stimulation of Schaeffer c o l l a t e r a l s and COMM stimulation suggests that both of these inputs terminate primarily on apical dendrites (Andersen, 1960; see Fig. 4 i n Andersen, 1975) Qx. Sepjtal-Hi£rjocam£al Projection Although the projections from the septal area to the hippocampus have been studied i n a number of species using various technigues, the precise organization of this system i s f a r from c l e a r , Powell (1963), on the basis of a Nauta-Gygax degeneration study i n the rat, concluded that f i b r e s from midline septal regions enter the body of the fornix reaching the hippocampus over the alveus to terminate throughout the hippocampus. Haisman (1966) using the same experimental preparation found that only the caudal aspect of the medial septal nucleus and the diagonal band send f i b r e s through the ventral portion of the 13 f i m b r i a which t e r m i n a t e i n a l l hippocampal r e g i o n s e x c e p t CA1. More r e c e n t l y , S i e g e l and T a s s o n i (1971) d i s a g r e e d w i t h Raisman (1966) s i n c e they r e p o r t e d t h a t l e s i o n s of the m e d i a l s e p t a l area r e s u l t i n d e g e n e r a t i o n o n l y i n the d o r s a l hippocampus i n c l u d i n g CA1. There i s no s i m p l e e x p l a n a t i o n f o r the t o p o g r a p h i c a l d i f f e r e n c e s found i n the above s t u d i e s . I n any case t h e i n t e r p r e t a t i o n s o f t h e p a t t e r n of d e g e n e r a t i o n f o l l o w i n g s e p t a l l e s i o n s have t o be t e n t a t i v e i n l i g h t of p o s s i b i l i t i e s t h a t l e s i o n s a l s o d i s r u p t a s c e n d i n g f i b r e s c o u r s i n g through t h e s e p t a l a r e a . Another t e c h n i g u e t h a t has been used t o map the s e p t a l - h i p p o c a m p a l p r o j e c t i o n s i s t h e o r t h o g r a d e t r a n s p o r t of t r i t i a t e d amino a c i d s t o the hippocampal f o r m a t i o n . The advantage o f t h i s t e c h n i g u e i s t h a t , w i t h proper p r e c a u t i o n s , i t m i n i m i z e s the c o n t r i b u t i o n o f f i b r e s of passage (Cowan, G o t t l i e b , H e n d r i c k s o n and P r i c e , 1972). F u r t h e r m o r e , a u t o r a d i o g r a p h y o f t e n r e v e a l s t e r m i n a l a r e a s which cannot be e a s i l y demonstrated u s i n g d e g e n e r a t i o n t e c h n i g u e s (Swanson and Cowan, 1977). F o l l o w i n g .3H-proline i n j e c t i o n s i n t o the m e d i a l s e p t a l - d i a g o n a l band a r e a , s u b s t a n t i a l l a b e l l i n g i s o bserved i n s e v e r a l r e g i o n s o f t h e hippocampus i n c l u d i n g CA2, CA3, CA4 o f the hippocampus and the h i l a r r e g i o n o f t h e d e n t a t e g y r u s . I n c o n t r a s t t o the d e g e n e r a t i o n e x p e r i m e n t s and those based on h i s t o c h e m i c a l l o c a l i z a t i o n o f a c e t y l c h o l i n e s t e r a s e 14 (reviewed below) Swanson and Cowan (1974) were unable t o f i n d e v i d e n c e f o r a s i g n i f i c a n t p r o j e c t i o n from the m e d i a l - s e p t a l d i a g o n a l band complex to CA 1 o r t h e i n n e r m o l e c u l a r l a y e r o f t h e d e n t a t e g y r u s . , More r e c e n t l y , Bose, H a t t o r i and F i b i g e r (1976) s u p p o r t e d t h e s e f i n d i n g s s i n c e they r e p o r t e d t h a t i n j e c t i o n s of 3H-l e u c i n e i n t o t h e m e d i a l s e p t a l n u c l e u s r e s u l t s i n s u b s t a n t i a l l a b e l l i n g of CA3 and CA4 zones of the hippocampus but o n l y s p a r s e l a b e l l i n g i n t h e m o l e c u l a r l a y e r of the d e n t a t e . However, i n f e c t i o n s of 3H-adenosine i n t o the m e d i a l s e p t a l n u c l e u s r e s u l t i n s u b s t a n t i a l t r a n s - s y n a p t i c l a b e l l i n g o f the i n n e r m o l e c u l a r l a y e r as w e l l as i n the h i l a r r e g i o n (Rose and S c h u b e r t , 1977)., I n a d d i t i o n t o the a n a t o m i c a l t e c h n i g u e s d e s c r i b e d above, h i s t o c h e m i c a l s t a i n i n g f o r a c e t y l c h o l i n e s t e r a s e (AchE) has been used t o map out the t e r m i n a t i o n o f the s e p t a l a f f e r e n t s . T h i s enzyme i s l o c a t e d i n d i s c r e t e l a y e r s i n t h e hippocampus and d e n t a t e g y r u s (Shute and L e w i s , 1967; Fonnum, 1970; Storm-Mathisen, 1970; M e l l g r e n and Srebo, 1973). The d e n t a t e r e g i o n s t a i n s more h e a v i l y f o r AchE than the hippocampus proper and the h e a v i e s t s t a i n i n g c o n s i s t s o f a band i n the i n n e r m o s t p o r t i o n of t h e m o l e c u l a r l a y e r a d j a c e n t t o t h e g r a n u l e c e l l b odies and t h e d e n t a t e h i l u s (Mosko et a l , 1973; Lynch and Cotman, 1975). F o l l o w i n g e l e c t r o l y t i c l e s i o n s o f the m e d i a l 15 septal-diagonal band area the above pattern of AchE staining i s abolished (Lewis et a l , 19 67; Srebro and Mellgren, 1974) and the o v e r a l l a c t i v i t y of AchE and cholineacetyltransferase (CAT) i s greatly reduced (McGeer, Hada, Tergo and Jung, 1969; Storm-Mathisen, 1970, 1972). In contrast, lesions of the l a t e r a l septum or l a t e r a l hypothalamus did not influence hippocampal AchE (McGeer et a l , 1969; Mellgren and Srebro, 1973) suggesting that a cholinergic input to the hippocampal formation originates i n or passes through the medial septal area. Under normal conditions the middle molecular region of the dentate which contains associational, commissural and entorhinal terminals does not s t a i n for AchE, but destruction of these systems r e s u l t s i n a marked i n t e n s i f i c a t i o n of AchE staining i n the deafferented zones (Lynch, Mathews, Mosko, Parks and Cotman, 1972; Lynch and Cotman, 1975). A subsequent lesion i n the medial septal area abolishes t h i s new pattern of AchE staining as well as the normal layered pattern of AchE i n the h i l u s (Lynch, 1972). On the basis of these data, i t has been suggested that septal afferents may undergo c o l l a t e r a l sprouting and take up positions in the outer molecular layer vacated by degenerating entorhinal afferents (See Lynch and Cotman, 1975 for review). Studies of the uptake of the precursor choline and 16 t h e subsequent r e l e a s e o f a c e t y l c h o l i n e (Ach) a l s o s u p p o r t t h e s u g g e s t i o n t h a t Ach i s p r o b a b l y r e l e a s e d i n t h e hippocampal f o r m a t i o n from t e r m i n a l s o r i g i n a t i n g i n the s e p t a l a r e a . Hippocampal synaptasomes have a h i g h a f f i n i t y uptake system f o r c h o l i n e which i s g r e a t l y reduced f o l l o w i n g m e d i a l s e p t a l l e s i o n s (Kuhar, Both and A g h a j a n i a n , 1972; Kuhar, 1975). T h i s d e c r e a s e i s o n l y i n t h e hippocampus and c o r t e x s u g g e s t i n g t h a t the uptake o f c h o l i n e may be a r e a s o n a b l y s p e c i f i c i n d i c a t o r o f Ach t u r n o v e r (Yamamura and Snyder, 1972). S t u d i e s which measure Ach r e l e a s e from the hippocampus, u t i l i z i n g a m o d i f i c a t i o n of the c o r t i c a l cup t e c h n i g u e , suggest t h a t t h e r e s t i n g r e l e a s e of Ach i s markedly i n c r e a s e d by s t i m u l a t i o n o f the m e d i a l septum (Smith, 1972, 1974; Tmdar, 1975, 1977). These e f f e c t s a r e s p e c i f i c t o m e d i a l s e p t a l s t i m u l a t i o n s i n c e s t i m u l a t i o n o f l a t e r a l septum o r c o n t r a l a t e r a l hippocampus does not i n f l u e n c e Ach r e l e a s e i n t h e hippocampus (Smith, 1972, 1974; Dudar, 1975). A d m i n i s t r a t i o n o f h e m i c h o l i n i u m - 3 (which b l o c k s c h o l i n e uptake) causes a g r a d u a l , dose-dependent d e p l e t i o n of Ach f o l l o w i n g m e d i a l s e p t a l s t i m u l a t i o n (Rommelspacker and Kuhar, 1974). , The a c t i o n of Ach on hippocampal neurones has been s t u d i e d u s i n g m i c r o - i o n t o p h o r e t i c t e c h n i g u e s (See S t r a u g h a n , 1975 f o r r e v i e w ) . Many hippocampal neurones a r e e x c i t e d by Ach (Herz and N a c i m i e n t o , 1965; 17 S a l m o i r a g h i and S t e f a n i s , 1965; B i s c o e and S t r a u g h a n , 1966). The r e c e p t o r s o f t h e s e c h o l i n o c e p t i v e neurones may be m u s c a r i n i c s i n c e t h e e x c i t a t i o n produced by Ach, but n o t t h a t produced by g l u t a m a t e , i s b l o c k e d by a t r o p i n e ( B i s c o e and S t r a u g h a n , 196 5; Wheal and M i l l e r , 1977). U n f o r t u n a t e l y , few o f t h e s e c h o l i n o c e p t i v e c e l l s were e l e c t r o p h y s i o l o g i c a l l y i d e n t i f i e d but i t appears t h a t b o t h p y r a m i d a l and g r a n u l e c e l l s a r e c h o l i n o c e p t i v e . More r e c e n t l y , Wheal and M i l l e r (1977) have r e p o r t e d t h a t e l e c t r i c a l s t i m u l a t i o n of t h e m e d i a l s e p t a l a r e a and t h e PP a c t i v a t e t h e same g r a n u l e c e l l s but o n l y the s e p t a l evoked a c t i v a t i o n i s b l o c k e d by a t r o p i n e . T h i s f u r t h e r s u p p o r t s the s u g g e s t i o n t h a t a c h o l i n e r g i c system o r i g i n a t i n g i n the m e d i a l s e p t a l r e g i o n e x c i t e s d e n t a t e g r a n u l e c e l l s . However, i t i s e x t r e m e l y d i f f i c u l t t o be sure t h a t s t i m u l a t i o n s e l e c t i v e l y e x c i t e s s e p t a l c e l l s and t h e i r axons but not f i b r e s o f passage c o u r s i n g through t h i s a r e a . Thus any c o n c l u s i o n s based on e l e c t r i c a l s t i m u l a t i o n of the s e p t a l a r e a can o n l y be t e n t a t i v e . I n c o n c l u s i o n , the n e u r o a n a t o m i c a l , h i s t o c h e m i c a l and p h a r m a c o l o g i c a l s t u d i e s r e v i e w e d above s t r o n g l y s u p p o r t the e x i s t e n c e of a s e p t a l - h i p p o c a m p a l p r o j e c t i o n which may use Ach as a t r a n s m i t t e r . , The p r e c i s e o r g a n i z a t i o n of s e p t a l t e r m i n a l s i n t h e d e n t a t e g y r u s i s s t i l l c o n t r o v e r s i a l s i n c e some s t u d i e s (Lynch e t a l , 1972; St o r m - M a t h i s e n , 1974) r e p o r t a s u b s t a n t i a l i n p u t t e r m i n a t i n g i n the s u p r a g r a n u l a r zone as w e l l as 18 i n t h e h i l u s whereas o t h e r s {Swanson and Cowan, 1975; Rose e t a l , 1976) o n l y r e p o r t s u b s t a n t i a l i n n e r v a t i o n o f the h i l u s . These d i f f e r e n c e s are r e l e v a n t f o r the i n t e r p r e t a t i o n of data based on i n t e n s i f i c a t i o n of AchE s t a i n i n g i n the r e - i n n e r v a t e d d e n t a t e (Lynch and Cotman, 1975) and t o d e t e r m i n e whether t h e s e p t a l i n p u t i s d i r e c t l y onto g r a n u l e c e l l s and/or neurones i n the h i l u s . £*., Monoamine Pathways The development o f t h e F a l c k - H i l l a r p h i s t o f l u o r e s c e n e e method f o r v i s u a l i z i n g monoamine c o n t a i n i n g c e l l b o d i e s and f i b r e s i n the CNS ( F a l c k , 1962; C a r l s o n , F a l c k and H i l l a r p , 1962; F a l c k , H i l l a r p , Thieme and Thorp, 1962) combined w i t h e x i s t i n g n e u r o a n a t o m i c a l and b i o c h e m i c a l t e c h n i g u e s have formed the b a s i s f o r s t u d y i n g the monoamine p r o j e c t i o n s t o the hippocampus., As o u t l i n e d below, t h e hippocampal f o r m a t i o n c o n t a i n s s u b s t a n t i a l amounts of t h e monoamines n o r a d r e n a l i n e (NA) and s e r o t o n i n (5-HT) but o n l y t r a c e amounts of dopamine ( f o r r e v i e w s see U n g e r s t e d t , 1971; Moore, 1975). The low c o n c e n t r a t i o n of dopamine i n t h e hippocampus s u g g e s t s t h a t i t does not p l a y an i m p o r t a n t r o l e i n s y n a p t i c t r a n s m i s s i o n i n t h i s a r e a a s i d e from b e i n g a p r e c u r s o r f o r n o r a d r e n a l i n e . The purpose of t h i s s e c t i o n i s t o d e s c r i b e t h e s o u r c e and p o s t u l a t e d p h y s i o l o g i c a l r o l e o f NA and 5-HT c o n t a i n i n g t e r m i n a l s i n the hippocampal f o r m a t i o n . 19 N o r a d r e n a l i n e I N A l l Fuxe (1964, 1965) f i r s t d e s c r i b e d N& c o n t a i n i n g c e l l b o d i e s l o c a t e d i n v a r i o u s n u c l e i i n the b r a i n s t e m . F l u o r e s c e n c e h i s t o c h e m i c a l mapping o f t h e i r p r o j e c t i o n s i n d i c a t e d t h a t t h e s e n u c l e i g i v e r i s e t o f i n e axons which descend and ascend f o r l o n g d i s t a n c e s r e a c h i n g widespread a r e a s i n t h e CNS (Anden, D a h l s t r o m , Fuxe, L a r s s o n , Olson and O n g e r s t e d t , 1966; Fuxe, H o k f e l t and U n g e r s t e d t , 1969; U n g e r s t e d t , 1971). Most r e l e v a n t t o t h e p r e s e n t t h e s i s i s the i d e n t i f i c a t i o n of a n u c l e u s i n the d o r s a l pons, t h e l o c u s c o e r u l e u s ( L C ) , c o n s i s t i n g of t i g h t l y packed c e l l s most o f which c o n t a i n NA (Anden et a l , 1966; U n g e r s t e d t , 1971). Axons from t h e LC p r o j e c t c a u d a i l y and r o s t r a l l y . The most prominent r o s t r a l p r o j e c t i o n i s through the d o r s a l NA bundle which p r o j e c t s i n t o t h e m e d i a l f o r e b r a i n bundle t o t e r m i n a t e throughout the f o r e b r a i n , i n c l u d i n g the s e p t a l r e g i o n , hippocampus and t h e e n t i r e c o r t e x (Fuxe, 1965; Anden e t a l , 1966; Fuxe, Hamberger and H o k f e l t , 1968; O n g e r s t e d t , 1971; l i n d v a l l and B j o r k l u n d , 1974, Jones and Moore, 1977; Moore, 1978). E l e c t r o l y t i c l e s i o n s i n the LC or d o r s a l NA b u n d l e r e s u l t i n a marked l o s s o f NA f l u o r e s c e n c e i n the above a r e a s (Anden e t a l , 1966; O n g e r s t e d t , 1971). B i o c h e m i c a l e x p e r i m e n t s p r o v i d e a d d i t i o n a l s u p p o r t f o r an NA c o n t a i n i n g p r o j e c t i o n t o t h e hippocampal f o r m a t i o n s i n c e d e s t r u c t i o n of the d o r s a l bundle s e v e r e l y reduces NA c o n t e n t of the hippocampus (Moore, 1973; K o b a y a s h i , 20 P a l k o v i t s , K o p i n and J a c o b o w i t z , 1974) whereas e l e c t r i c a l s t i m u l a t i o n o f t h i s pathway ca u s e s a r e l e a s e o f NA i n the hippocampus ( A r b u t h n o t t , Crow, Fuxe, Olson and U n g e r s t e d t , 1970). The NA c o n t a i n i n g f i b r e s r e a c h t h e hippocampus v i a the f o r n i x or by l o o p i n g around the c o r p u s c a l l o s u m and e n t e r i n g through t h e cingulum ( U n g e r s t e d t , 1971; L i n d v a l l and B j o r k l u n d , 1974; Moore, 1 975) . However, u s i n g an immunofluorescence t e c h n i g u e t o v i s u a l i z e d o p a m i n e - b e t a - h y d r o x y l a s e (DBH), Swanson and Hartman (1975) f a i l e d t o f i n d D B H - c o n t a i n i n g f i b r e s i n t h e f o r n i x - f i m b r i a s u g g e s t i n g t h a t t h e c a u d a l r o u t e may be more i m p o r t a n t . , The d i f f i c u l t i e s i n d i s c r i m i n a t i n g between NA t e r m i n a l s and f i b r e s based on NA or DBH f l u o r e s c e n c e make a p r e c i s e d e s c r i p t i o n of t h e i r d i s t r i b u t i o n t e n t a t i v e . However, combined w i t h a u t o r a d i o g r a p h i c t e c h n i g u e s (Jones and Moore, 1977) , a l a y e r e d NA d i s t r i b u t i o n i n t h e hippocampus may be described.„The de n s e s t t e r m i n a l r e g i o n i s t h e d e n t a t e h i l u s and a d j a c e n t mossy f i b r e - C A 3 zone f o l l o w e d by moderately dense p r o j e c t i o n to t h e s t r a t u m lacunosum of CA1 (Swanson and Hartman, 1975; Jones and Moore, 1977). The m o l e c u l a r l a y e r o f t h e d e n t a t e and t h e area of p y r a m i d a l c e l l b o d i e s were o n l y s p a r s e l y l a b e l l e d . More r e c e n t l y , a h i s t o f l u o r e s c e n c e t e c h n i g u e f o r v i s u a l i z i n g a d r e n e r g i c r e c e p t o r s was used t o study t h e d i s t r i b u t i o n 21 o f p o s s i b l e NA r e c e p t o r s i t e s (Melamed, l a h a v and A t l a s , 1977). In c o n t r a s t t o NA hist©fluorescence and a u t o r a d i o g r a p h i c s t u d i e s d e s c r i b e d above, t h i s s t u d y r e p o r t s a h i g h d e n s i t y of b e t a - a d r e n e r g i c r e c e p t o r s i t e s ( i . e . s i t e s which b i n d a f l u o r e s c e n t analogue of p r o p r a n o l o l ) i n t h e d e n t a t e g r a n u l e c e l l l a y e r . , The e f f e c t s of m i c r o - i o n t o p h o r e t i c a l l y a p p l i e d NA on neurones i n t h e hippocampal f o r m a t i o n i n d i c a t e t h a t i t produces a l o n g l a t e n c y i n h i b i t i o n o f p y r a m i d a l c e l l s (Herz and N a c i m i e n t o , 1965; S a l m o i r a g f a i and S t e f a n i s , 1965; B i s c o e and S t r a u g h a n , 1966; See St r a u g h a n , 1975 f o r review) . S e g a l and Bloom (1974 a,b) attempted t o r e l a t e t h e a c t i o n o f i o n t o p h o r e t i c a l l v a p p l i e d NA and t h e e f f e c t s of e l e c t r i c a l s t i m u l a t i o n o f LC on hippocampal p y r a m i d a l c e l l s , , These a u t h o r s r e p o r t e d t h a t LC s t i m u l a t i o n o r d i r e c t a p p l i c a t i o n of NA i n h i b i t p y r a m i d a l c e l l s . Moreover, t h e s e e f f e c t s were b l o c k e d by b e t a - a d r e n e r g i c a n t a g o n i s t s and were absent i n 6 -hydroxydopamine-treated r a t s ( S e g a l and Bloom, 1974b). Although t h e s e data a r e s u g g e s t i v e , t h e r e i s s t i l l no f i r m p h y s i o l o g i c a l e v i d e n c e i m p l i c a t i n g NA-c o n t a i n i n g t e r m i n a l s i n s y n a p t i c f u n c t i o n of the hippocampal f o r m a t i o n . 22 S e r o t o n i n 1 5 - H T J L I The 5-HT i n n e r v a t i o n o f the hippocampal f o r m a t i o n o r i g i n a t e s from neurones w i t h i n t h e m i d b r a i n raphe n u c l e i (Dahlstrom and Fuxe, 1965; Anden e t a l , 1966), B i o c h e m i c a l s t u d i e s by Lorens and G u l d b e r g (1974) i n d i c a t e t h a t t h e median raphe ( n u c l e u s c e n t r a l i s s u p e r i o r ) and not the d o r s a l raphe i s the s o u r c e of hippocampal 5-HT, H i s t o f l u o r e s c e n c e (Fuxe e t a l , 1970; B j o r k l u n d e t a l , 1973) and a u t o r a d i o g r a p h i c (Moore and H a l a r i s , 1975) s t u d i e s i n d i c a t e t h a t axons a r i s i n g from t h e median raphe ascend i n t h e m e d i a l f o r e b r a i n bundle t o r e a c h the hippocampal f o r m a t i o n v i a the f o r n i x and the c i n g u l u m , H i t h i n t h e hippocampal f o r m a t i o n tha d i s t r i b u t i o n of 5-HT i s almost i m p o s s i b l e to s t u d y u s i n q h i s t o c h e m i c a l methods due t o the low f l u o r e s c e n c e and s m a l l s i z e o f the t e r m i n a l s . Even t h e g l y o x y l i c a c i d method ( L i n d v a l l and B j o r k l u n d , 1974) which improves t h e l o c a l i z a t i o n o f c a t e c h o l a m i n e s does not a i d i n t h e d e t a i l e d a n a l y s i s o f 5-HT systems. However, d a t a based on a u t o r a d i o g r a p h i c p r o cedures suggest t h a t the d i s t r i b u t i o n o f axons a r i s i n g from t h e median raphe n u c l e u s e s s e n t i a l l y o v e r l a p s the NA i n n e r v a t i o n d e s c r i b e d i n t h e p r e v i o u s s e c t i o n (Conrad, L e o n a r d and P f a f f , 1974; Moore and H a l a r i s , 1975), The densest t e r m i n a t i o n i n t h e hippocampal f o r m a t i o n i s r e s t r i c t e d t o a band a p p r o x i m a t e l y 65 um wide, a l o n g the v e n t r a l 23 border of d e n t a t e g r a n u l e c e l l s (Moore, 1975). I t cannot be d etermined whether t h e s e t e r m i n a l s a r e on c e l l b o d i e s of g r a n u l e c e l l s o r s u b j a c e n t h i l a r neurones. There i s s p a r s e t o moderate i n n e r v a t i o n of t h e d e n t a t e m o l e c u l a r l a y e r (See Moore, 1975 F i g . , * ! ) . . I t must be kept i n mind t h a t the above t e r m i n a l p a t t e r n does not n e c e s s a r i l y r e f l e c t 5-HT i n n e r v a t i o n s i n c e some raphe neurones a r e not s e r o t o n e r g i c . More r e c e n t l y H a l a r i s et a l (1976) have shown t h a t i n j e c t i o n s of r a d i o l a b e l e d 5-HTP i n t o the MB r e s u l t i n a s i m i l a r p a t t e r n o f t e r m i n a t i o n t o the amino a c i d . F u r thermore, " s e l e c t i v e " d e s t r u c t i o n o f 5-HT t e r m i n a l s w i t h 5 , 6 - d i h y d r o x y t r y p t a m i n e r e s u l t s i n g r e a t l y reduced a x o n a l t r a n s p o r t from t h e median raphe t o the hippocampal f o r m a t i o n . Both of these o b s e r v a t i o n s s u p p o r t t h e c o n c l u s i o n t h a t t h e p a t t e r n o f i n n e r v a t i o n d e s c r i b e d above r e f l e c t s the d i s t r i b u t i o n o f 5-HT c o n t a i n i n g t e r m i n a l s i n t h e hippocampal f o r m a t i o n . , S e r o t o n i n (5-HT) has been i o n t o p h o r e t i c a l l y a p p l i e d t o p y r a m i d a l c e l l s i n a n a e s t h e t i z e d c a t s ( S t e f a n i s , 1964; B i s c o e and S t r a u g h a n , 1965) and r a t s ( S e g a l and Bloom, 1974a). I n most cases 5-HT caused a s h o r t l a t e n c y (2-15 sec) d e p r e s s i o n of n e u r o n a l d i s c h a r g e which, u n l i k e t h e d e p r e s s e n t a c t i o n o f NA,did not p e r s i s t a f t e r the drug a p p l i c a t i o n had been stopped. In f a c t , NA was a weak d e p r e s s a n t of n e u r o n a l f i r i n g r e l a t i v e t o 5-HT when comparable e j e c t i o n 24 c u r r e n t s were used { B i s c o e and Straughan, 1966). N e i t h e r the mechanism by which 5-HT d e p r e s s e s hippocampal neurones nor t h e p h y s i o l o g i c a l s i g n i f i c a n c e of t h i s a c t i o n are known. For t h e s e r e a s o n s any p o s t u l a t e d r o l e f o r 5-HT or NA i n s y n a p t i c t r a n s m i s s i o n i n t h e hippocampus must be t e n t a t i v e ; many c e l l s may have r e c e p t o r s f o r s u b s t a n c e s not n o r m a l l y r e l e a s e d onto them as t r a n s m i t t e r s . Major Hippocampal E f f e r e n t s A l t h o u g h t h e e f f e r e n t c o n n e c t i o n s of the hippocampal f o r m a t i o n have been s t u d i e d f o r more than a c e n t u r y , our u n d e r s t a n d i n g o f t h e i r o r g a n i z a t i o n i s c o m p a r a t i v e l y vague. Only r e c e n t l y have d e t a i l e d d e s c r i p t i o n s o f the hippocampal e f f e r e n t s o t h e r than t h e f o r n i x been at t e m p t e d . The new d a t a have prompted a r e - e v a l u a t i o n o f t h e w i d e l y h e l d view t h a t t h e f o r n i x i s t h e major, i f not t h e o n l y , e f f e r e n t from t h e hippocampal f o r m a t i o n (Maynert, 1872; Von Gudden, 1881; C a j a l , 1911; Nauta, 1956; G u i l l e r y , 1957) and have emphasized t h e i m p o r t a n c e of c a u d a l l y - d i r e c t e d e f f e r e n t s t o c o r t i c a l and s u b c o r t i c a l r e g i o n s ( C h r o n i s t e r and White, 1975; H j o r t h - S i m o n s e n , 1971, 1973; Swanson and Cowan, 1977). , Another new development has been i n r e g a r d t o what r e g i o n s o f the hippocampal f o r m a t i o n g i v e r i s e to e x t r a h i p p o c a m p a l p r o j e c t i o n s . At t h e s i m p l e s t l e v e l . 25 t h e d e n t a t e g y r u s does not have any e x t r a h i p p o c a m p a l p r o j e c t i o n s s i n c e i t s o u t f l o w c o n s i s t s a l m o s t e n t i r e l y of t h e mossy f i b r e p r o j e c t i o n t o CA3-CA4 p y r a m i d a l c e l l s . L i k e w i s e t h e CA4 p y r a m i d a l c e l l s have no e x t r a h i p p o c a m p a l p r o j e c t i o n but do g i v e r i s e t o a s s o c i a t i o n a l and c o m m i s s u r a l p r o j e c t i o n s t o t h e i p s i l a t e r a l and c o n t r a l a t e r a l d e n t a t e , r e s p e c t i v e l y (Zimmer, 1970; G o t t l i e b and Cowan, 1973). F i e l d CA3 g i v e s r i s e t o e x t e n s i v e i n t r a h i p p o c a m p a l a s s o c i a t i o n a l / c o m m i s s u r a l p r o j e c t i o n s a s w e l l as an e x t r a h i p p o c a m p a l p r o j e c t i o n t o the s e p t a l a r e a ( G o t t l i e b and Cowan, 1973; Saisman e t a l , 1966). Perhaps the most s u b s t a n t i a l e x t r a h i p p o c a m p a l p r o j e c t i o n a r i s e s from CAi and p r o j e c t s through the pre c o m m i s s u r a l f o r n i x b i l a t e r a l l y t o the l a t e r a l s e p t a l a r e a (Swanson and Cowan, 1977) and th r o u g h the a l v e u s t o the a d j a c e n t s u b i c u l u m ( H j o r t h - S i m o n s e n , 1973; Swanson and Cowan, 1977) . The s u b i c u l u m , i n t u r n , g i v e s r i s e t o t h e massive p o s t c o m m i s s u r a l f o r n i x (Baisman et a l , 1966; Swanson and Cowan, 1977). On t h e b a s i s of t h e above d a t a , t h e hippocampus proper c o n t r i b u t e s o n l y t o t h e pr e c o m m i s s u r a l f o r n i x whereas t h e s u b i c u l u m i s t h e s o u r c e of t h e e n t i r e p o s t -c o m m i s s u r a l p r o j e c t i o n t o the a n t e r i o r t h a l a m i c and mammillary n u c l e i . I n a d d i t i o n to t h e s e d i r e c t e x t r a h i p p o c a m p a l p r o j e c t i o n s d e s c r i b e d i n more d e t a i l below, i n d i r e c t c o n n e c t i o n s o f t h e hippocampal f o r m a t i o n through t h a l a m i c n u c l e i may i n f l u e n c e t h e 26 f r o n t a l and p a r i e t a l l o b e s t h e r e b y c o m p l e t i n g the so-c a l l e d Papez (1937) c i r c u i t . l i l f e r i a ^ F o r n i x System Although E l l i o t Smith (1897) c o n c l u d e d t h a t the arrangement o f t h e f o r n i x conforms to t h e same fundamental p l a n i n a l l v e r t e b r a t e s , many subseguent s t u d i e s have shown s i g n i f i c a n t d i f f e r e n c e s between s p e c i e s and i n d i f f e r e n t s t r a i n s of the same s p e c i e s ( E d i n g e r and W a l l e n b e r g , 1902; Simpson, 1952; Cowan and P o w e l l , 1955; V a l e n s t e i n and Nauta, 1959). I n t h e r a t , the d i r e c t hippocampal p r o j e c t i o n s c o n t a i n e d i n the pr e c o m m i s s u r a l f o r n i x p r o j e c t t o the s e p t a l n u c l e i and t h e p r e o p t i c r e g i o n and t h r o u g h t h e columns o f t h e f o r n i x t o the mammillary body and the m e d i a l hypothalamic r e g i o n ( C a j a l , 1911; Nauta, 1956; Swanson and Cowan, 1 9 7 1 ) . O n l y a few f o r n i c a l f i b r e s r e a c h t h e m i d b r a i n i n t h e r a t . I n c o n t r a s t , t h e c a t f o r n i x has a massive p r o j e c t i o n t o t h e r o s t r a l raid-brain and few f i b r e s t o t h e m e d i a l h y p o t h a l a m i c r e g i o n (Nauta, 1959). I n p r i m a t e s the d o r s a l f o r n i x p r o j e c t i o n s a r e l e s s pronounced than they a r e i n r o d e n t s and do not appear t o t e r m i n a t e c a u d a l t o t h e mammillary b o d i e s ( P a l e t t i and C r e s w e l l , 1976). S e v e r a l p r o p o s a l s have been made r e g a r d i n g the t o p o g r a p h i c a l o r g a n i z a t i o n o f t h e hippocampal e f f e r e n t s v i a t h e f i m b r i a - f o r n i x . Raisman e t a l (1966) suggested 27 t h a t CA1 p r o j e c t s p r i n c i p a l l y t o the m e d i a l s e p t a l r e g i o n w h i l e CA3 p r o j e c t s t o the l a t e r a l septum. In c o n t r a s t S i e g e l e t a l (1974) a l s o u s i n g r a t s , suggest t h a t t h e m e d i a l and l a t e r a l s e p t a l r e g i o n s a re i n n e r v a t e d by t h e d o r s a l and v e n t r a l hippocampus, r e s p e c t i v e l y . The p o s t c o m m i s s u r a l f o r n i x o r i g i n a t i n g i n t h e s u b i c u l u m may a l s o be t o p o g r a p h i c a l l y o r g a n i z e d i n t h e d o r s o v e n t r a l a x i s . Swanson and Cowan (1977) suggest t h a t the d o r s a l s u b i c u l u m p r o j e c t s p r e d o m i n a n t l y upon the mammillary complex and the v e n t r a l s u b i c u l u m p r o j e c t s t o t h e r o s t r a l hypothalamus and t h e b a s a l f o r e b r a i n i n c l u d i n g the n u c l e u s accumbens and t h e bed n u c l e u s o f the s t r i a t e r m i n a l i s . P l a s t j c i t y •• Of Sjrnarrtic T r a n s m i s s i o n I n The Hippocampal F o r m a t i o n S i n c e t h e major a f f e r e n t s t o the hippocampal f o r m a t i o n occupy d i s c r e t e d e n d r i t i c l a y e r s which can be i d e n t i f i e d u s i n g b oth n e u r o a n a t o m i c a l and e l e c t r o p h y s i o l o g i c a l t e c h n i g u e s , i t i s an i d e a l system f o r e v a l u a t i n g s t r u c t u r a l and f u n c t i o n a l m o d i f i c a t i o n s of s y n a p t i c t r a n s m i s s i o n . S t r u c t u r a l p l a s t i c i t y has been e v a l u a t e d by d e t e r m i n i n g t h e a n a t o m i c a l r e a d j u s t m e n t s which o c c u r f o l l o w i n g e l i m i n a t i o n of a major a f f e r e n t t o the same r e g i o n . F u n c t i o n a l p l a s t i c i t y has been s t u d i e d p r i m a r i l y by d e t e r m i n i n q t h e changes i n e f f i c a c y o f s y n a p t i c t r a n s m i s s i o n r e s u l t i n g from p r e v i o u s a c t i v a t i o n o f t h e same pathway. 28 Both of these approaches may contribute to an understanding of the mechanisms associated with processing and storage of information, however, only functional p l a s t i c i t y has direct relevance to the present work and w i l l be reviewed below. Several reviews on terminal p r o l i f e r a t i o n and synaptogenesis i n the hippocampal formation following deafferentation are available (Lynch and Cotman, 1975; ficSilliams and Lynch, 1978)., A feature of a l l excitatory afferents i n the hippocampal formation studied so far i s the increase i n synaptic transmission during r e p e t i t i v e stimulation. The increased e f f i c a c y i s seen as an enhancement i n both the amplitude and rate of r i s e of e x t r a c e l l u l a r f i e l d responses and an increase in the number of c e l l s which are activated by the test pulse (Lomo, 1971b; B l i s s and Lomo, 1973), The simplest example of t h i s phenomenon was provided by Lomo (1971b) using double pulse stimulation of the perforant path-dentate projection in the rabbit. The monosynaptic test EPSP and population spike (the synchronous discharge of a number of c e l l s ) responses were both potentiated by a preceding (20-300 mesc) conditioning volley. The test EPSP was potentiated as much as 10 0% while the population spike could be potentiated up to 300%. These observations have been recently confirmed i n the rat (Steward et a l , 1976; Assaf and M i l l e r , 1978) and in the i n v i t r o hippocampal s l i c e (Dudek et a l , 1975). 29 I f some e x c i t a t o r y synapses i n t h e hippocampal f o r m a t i o n are s t i m u l a t e d w i t h p a r t i c u l a r f r e q u e n c i e s { u s u a l l y 4-15/sec) t h e r e i s a g r a d u a l i n c r e a s e i n the a m p l i t u d e o f evoked p o t e n t i a l s u n t i l a maximum response i s o b t a i n e d a f t e r 10-30 s t i m u l i ( B l i s s and Lomo, 1973; Andersen, 1975). With h i g h e r s t i m u l a t i o n r a t e s , the d u r a t i o n o f t h e enhanced p o t e n t i a l s i s s h o r t e r , g i v i n g way t o a p e r i o d o f d e p r e s s i o n u s u a l l y l a s t i n g l e s s than 5 minutes. T h i s s o - c a l l e d " freguency f a c i l i t a t i o n " i s more o b v i o u s i n the mossy f i b r e - C A 3 pathway t h a n i n t h e p e r f o r a n t p a t h - d e n t a t e p r o j e c t i o n (Andersen, 1975). T e t a n i c s t i m u l a t i o n g i v e s r i s e t o an i n c r e a s e d r e s p o n s i v e n e s s of the s t i m u l a t e d pathway t o s i n g l e v o l l e y s d e l i v e r e d s e v e r a l hours l a t e r ( B l i s s and Lome, 1973), S i m i l a r p o s t - t e t a n i c p o t e n t i a t i o n has r e c e n t l y been demonstrated i n i n v i t r o hippocampal s l i c e (Lynch e t a l , 1977; T y l e r and A l g e r , 1975; Andersen e t a l , 1977; Dunwiddie and Lynch, 1978). The observed p o t e n t i a t i o n i s not dependent on changes i n t h e p r e s y n a p t i c f i b r e v o l l e y and i s not observed i n a d j a c e n t s y n a p t i c i n p u t s which were not t e t a n i z e d (Lynch e t a l , 1977; Andersen et a l , 1977). I n f a c t , Dunwiddie and Lynch (1978) r e c e n t l y r e p o r t e d a d e p r e s s i o n o f t h e n o n - t e t a n i z e d pathway which o c c u r s s i m u l t a n e o u s l y w i t h p o t e n t i a t i o n o f t h e s t i m u l a t e d pathway. 30 Mechanisms which mediate the above forms cf functional p l a s t i c i t y are not known. Andersen (1975) suggests that they may r e f l e c t a changed a b i l i t y of the presynaptic terminal to deliver increased amounts of transmitter. More recently, Dunwiddie, Madison and Lynch (1 978) observed that synaptic transmission i s necessary for producing long-term potentiation. These and other p o s s i b i l i t i e s are discussed i n more d e t a i l at the end of Chapter 8. lxl±2 The Septal Area The septal area comprises a group of nuclei located between the anterior horns of the l a t e r a l v e n t r i c l e s . Their extent i s bounded ventrally by the decussations of the anterior commissure; dorsallv by the corpus callosum; r o s t r a l l y by the f r o n t a l cortex and caudally by the descending columns of the fornix (Hines, 1922; Young, 1936; Andy and Stephan, 1965). On the basis of i t s cytoarchitecture the septal area of the rat may be subdivided into medial, l a t e r a l , posterior and ventral d i v i s i o n s (Swanson and Cowan, 1974). The medial septal region consists of the medial septal nucleus dors a l l y and the nucleus of the diagonal band of Broca v e n t r a l l y . The distinguishing feature of the medial septal region i s the mass of large neurones which i n Golgi preparations have long spine-free dendrites (Tombol and Petsche, 1969; Swanson and Cowan, 1974). These large bipolar neurones are interposed by 31 many s m a l l neurones. The l a t e r a l s e p t a l r e g i o n c o n s i s t s p r i m a r i l y of medium s i z e d c e l l s which can be grouped i n t o d o r s a l , i n t e r m e d i a t e and v e n t r a l s u b d i v i s i o n s (Swanson and Cowan, 1974). The p o s t e r i o r s e p t a l r e g i o n c o n s i s t s of t h e s e p t o - f i m b r i a l n u c l e u s which i s embedded w i t h i n the p r e c o m m i s s u r a l f o r n i x and the t r i a n g u l a r s e p t a l n u c l e u s which c o n t a i n s t i g h t l y packed c e l l s w i t h i n the v e n t r a l hippocampal commissure. The v e n t r a l s e p t a l r e g i o n i s p o o r l y d e f i n e d w i t h t h e e x c e p t i o n of t h e bed n u c l e u s o f t h e s t r i a t e r m i n a l i s which r e c e i v e s a f f e r e n t s from t h e amygdala (Andy and Stephan, 1965, 1974; Tombol and P e t s c h e , 1969; Swanson and Cowan, 1974)., Although the above d e s c r i p t i o n and t e r m i n o l o g y a r e based p r i m a r i l y on i n v e s t i g a t i o n s i n t h e r a t , t h e y may be a p p l i c a b l e t o o t h e r mammalian s p e c i e s s i n c e t h e s e p t a l r e g i o n r e t a i n s t h e above f e a t u r e s i n p r i m a t e e v o l u t i o n . T h i s u n i f o r m i t y i s m a i n t a i n e d d e s p i t e the p r o g r e s s i v e enlargement and r e l a t i v e development o f the prim a t e septum (Andy and Stephan, 1965, 1974). 32 Major A f f e r e n t s The s e p t a l r e g i o n r e c e i v e s i t s major a f f e r e n t s from the hippocampal f o r m a t i o n v i a t h e f o r n i x system, from the c o r t i c o - m e d i a l amygdala v i a t h e s t r i a t e r m i n a l i s and from h y p o t h a l a m i c and m i d b r a i n r e g i o n s by way of t h e m e d i a l f o r e b r a i n bundle. More r e c e n t l y , a s c e n d i n g monoamine systems o r i g i n a t i n g i n t h e pons and m i d b r a i n have been shown t o e x t e n s i v e l y i n n e r v a t e t h e s e p t a l r e g i o n . On the b a s i s o f t h e s e c o n n e c t i o n s , the s e p t a l r e g i o n o c c u p i e s a p i v o t a l p o s i t i o n between m i d b r a i n and f o r e b r a i n a r e a s , Jbs. Hj£ppcampal Xg.£Uts The major i n p u t from t h e hippocampal f o r m a t i o n v i a t h e p r e c o m m i s s u r a l f o r n i x t e r m i n a t e s i n t h e l a t e r a l s e p t a l r e g i o n ( P o w e l l et a l , 1957; Raisman e t a l , 1966; Swanson and Cowan, 1977) , Raisman e t a l (1966) observed t h a t l e s i o n s i n f i e l d s CA1 and C A3 r e s u l t i n d e g e n e r a t i o n d i s t r i b u t e d t o t h e d o r s a l and l a t e r a l p a r t s of t h e l a t e r a l s e p t a l n u c l e u s . More r e c e n t l y , Swanson and Cowan (1977) s u g g e s t e d t h a t t h e above p a t t e r n o f d e g e n e r a t i o n does not i n d i c a t e t h a t t h e s o l e s o u r c e of t h e p r e c o m m i s s u r a l f o r n i x i s t h e hippocampus proper s i n c e f i b r e s o r i g i n a t i n g from the s u b i c u l u m are a l s o d e s t r o y e d by t h e s e l e s i o n s . On t h e b a s i s of a u t o r a d i o g r a p h i c s t u d i e s , Swanson and Cowan (1977) c o n c l u d e d t h a t t h e major s o u r c e o f t h e s e p r e c o m m i s s u r a l 33 f i b r e s may be the i p s i l a t e r a l subicular region. The afferents to the l a t e r a l septal reqion are topographically organized such that dorsal hippocampal regions project exclusively to the dorsal aspect of the l a t e r a l septal nucleus and ventral hippocampal f i e l d s project to the ventral portions of the l a t e r a l septal area (Raisman et a l , 1966; Siegel and Tassoni, 1971; Swanson and Cowan, 1977). The Electrophysiological studies of McLennan and M i l l e r (1974b,1976) support this topographical organization since short latency f i e l d responses were recorded in the l a t e r a l septal area following stimulation of the fimbria. These data, taken together with the previously described septal-hippocamapal projection (section 1.1.1), establish a r e c i p r o c a l l i n k between the septal area and the hippocampal formation. The precommissural fornix terminates i n the l a t e r a l septal nucleus and t h i s , i n turn, projects to the medial septal area which sends efferents to the hippocampal formation. 34 3U Y %9%h 3 1 a mi c I npat s The medial forebrain bundle (MFB) i s the major input from the hypothalamus and the midbrain (Milhouse, 1969). Since the MFB consists of f i b r e s o r i g i n a t i n g in both of these regions, i t is d i f f i c u l t to assess the contribution of hypothalamic nuclei to t h i s system, however, G u i l l e r y (1957) suggested that a hypothalamo-septal group of fi n e f i b r e s originates in l a t e r a l hypothalamic areas and projects to the l a t e r a l septal region. On the other hand, large f i b r e s appear to originate i n the midbrain and project primarily to the medial septal region (Guillery, 1957; Nauta and Kuypers, 1958; Morest, 1961; Wolf and Sutin, 1966). This anatomical d i f f e r e n t a t i o n i s partly supported by the observations that e l e c t r i c a l stimulation i n the l a t e r a l hypothalamus res u l t s i n desynchronization of hippocampal e l e c t r i c a l a c t i v i t y and disruption of the bursting discharge of septal neurones; i n contrast, stimulating more medially produces the opposite e f f e c t s (Anchel and Lindsley, 1972; Wilson et a l , 1S75). However, any conclusions based on the above anatomical or functional data should be re-evaluated in l i q h t of the p o s s i b i l i t y that electrodes in these s i t e s may stimulate ascending monoamine-containing f i b r e s which pass through t h i s area to terminate i n the septal area (see below). 3 5 C._ A i y g d a l o i d I n p u t s The major i n p u t from t h e amygdala i s v i a t h e s t r i a t e r m i n a l i s which o r i g i n a t e s i n the c o r t i c o m e d i a l amygdaloid n u c l e u s and i n n e r v a t e s t h e bed n u c l e u s of t h e s t r i a t e r m i n a l i s i n the v e n t r a l s e p t a l r e g i o n {Fox, 1 9 4 0 ; G l o o r , 1 9 5 5 ; Nauta, 1 9 6 6 ; Cowan e t a l , 1 9 6 5 ; Leonard and S c o t t , 1 9 7 1 ) . A second i n p u t o r i g i n a t e s i n t h e b a s o l a t e r a l amygdaloid n u c l e i and p r o j e c t s v i a t h e v e n t r a l a m y g d a l o f u g a l pathway t o the d i a g o n a l band r e g i o n {Fox, 1 9 4 0 ; Nauta, 1 9 5 6 ; Leonard and S c o t t , 1 9 7 1 ) . I t i s no t e w o r t h y t h a t both t h e s t r i a t e r m i n a l i s and t h e amy g d a l o f u g a l pathways a l s o p r o j e c t v e n t r a l l y p ast the s e p t a l r e g i o n t o t e r m i n a t e i n the p r e o p t i c a r e a , hypothalamus and i n t o t h e m e d i a l f o r e b r a i n bundle. On the b a s i s of e l e c t r o p h y s i o l o g i c a l e v i d e n c e Renaud ( p e r s o n a l communication) s u g g e s t s t h a t t h e same neurone i n the amygdala can p r o j e c t t o s e p t a l and h y p o t h a l a m i c s i t e s . Ms. Monoamine Pathways N o r a d r e n a l i n e j N A ) T ; On the b a s i s of F a l c k - H i l l a r p and g l y o x y l i c a c i d h i s t o c h e m i c a l s t u d i e s , the r e g i o n a l d i s t r i b u t i o n o f NA t e r m i n a l s w i t h i n t h e s e p t a l a r e a has been found t o be dense i n l a t e r a l septum and sp a r s e i n the media l s e p t a l a r e a (Moore et a l , 1 9 7 1 ; L i n d v a l l and B j o r k l u n d , 1 9 7 4 ) . 36 Recently, Moore (1978) has noted that the bed nucleus of s t r i a terminalis receives a very dense NA innervation. Moreover, the glyoxylic acid preparations reveal numerous f i n e terminals near c e l l bodies and dendrites which may make en JBstssage synapses i n the septal area and then continue to more r o s t r a l forebrain and c o r t i c a l areas (Ongerstedt, 1971; Moore et a l , 1971; L i n d v a l l and Bjorklund, 1974; Moore, 1975) As i n the case of other forebrain regions, the septal area has been shown to receive NA-containing afferents from the locus coeruleus (LC) (Ungerstedt, 1971; Segal and Landis, 1974; Jones and Moore, 1971). However, Moore (1978) has recently shown that lesions of LC r e s u l t in only a 48% decrease i n septal NA content, suggesting that a substantial proportion of septal NA originates i n other regions. Since transections caudal to LC also result i n substantial (47%) depletions of of septal NA, the other sources of septal NA may be in more caudal brainstem s i t e s . Dopamine I D f t j i At one time, the catecholaminergic innervation of the septal area was considered to consist almost exclusively of NA nerve terminals (Moore et a l , 1971). More recently, histofluorescence studies have demonstrated a high density of DA terminals in the medial aspect of the l a t e r a l septal region (Lindvall, 37 1975; Moore, 1975, 1978). Furthermore, L i n d v a l l (1975) has shown that transection of the medial forebrain bundle at the l e v e l of the r o s t r a l substantia nigra, or b i l a t e r a l e l e c t r o l y t i c lesions of the A10 mesencephalic c e l l group res u l t i n a complete loss of septal DA terminals. Small i n j e c t i o n s of HBP into the l a t e r a l septal area r e s u l t i n extensive l a b e l l i n g of neurones i n the ventrolateral extent of the A10 area at the l e v e l of the interpeduncular nucleus (Assaf and M i l l e r , 1977; Moore, 1978). Taken together these anatomical and biochemical studies suggest a direct dopaminergic f i b r e system which originates i n the A10 region of the ventromedial tegmentum and innervates the l a t e r a l septal area . In contrast, there are r e l a t i v e l y few f i b r e s to the medial septal region. The functional role of DA in the septal region i s not yet clear. Assaf and M i l l e r (1977) have shown that e l e c t r i c a l stimulation i n the region of A10 results i n a short latency excitation of septal neurones which i s dependent on an intact dopamine system. The d i s t r i b u t i o n of the activated neurones i s similar to that of the dopamine innervation based on the histochemical experiments mentioned above. 38 S e r o t o n i n jtS-HT).:. On t h e b a s i s o f a u t o r a d i o g r a p h i c and b i o c h e m i c a l d a t a , s e r o t o n i n - c o n t a i n i n g c e l l b odies l o c a l i z e d t o t h e m i d b r a i n raphe n u c l e i have been shown t o e x t e n s i v e l y i n n e r v a t e t h e s e p t a l r e g i o n (Moore and H e l l e r , 1967, Moore, 1974; Conrad e t a l , 1974; Lorens and G u l d b e r g , 1974). L e s i o n s i n t h e m i d b r a i n raphe or m e d i a l f o r e b r a i n bundle s i g n i f i c a n t l y reduce 5-HT c o n t e n t of the s e p t a l a r e a (Moore and H e l l e r , 1967; Kuhar e t a l , 1972; L o r e n s and G u l d b e r g , 1974; Jacobs e t a l , 1974). The p r o x i m i t y o f t h e d o r s a l and median raphe n u c l e i make i t d i f f i c u l t t o l o c a l i z e the e x a c t o r i g i n of s e p t a l 5-HT u s i n g l e s i o n i n g t e c h n i g u e s . However, the r e s u l t s of a u t o r a d i o g r a p h i c (Conrad e t a l , 1974; H a l a r i s e t a l , 1S76) and HHP t r a c i n g t e c h n i g u e s ( J . J . M i l l e r , u n p u b l i s h e d o b s e r v a t i o n s ) i n d i c a t e t h a t t h e median raphe c o n t r i b u t e s s u b s t a n t i a l l y more f i b r e s t o t he s e p t a l a r e a than does t h e d o r s a l raphe. U n l i k e t h e c a t e c h o l a m i n e s , 5-HT does not f l u o r e s c e w e l l i n F a l c k - H i l l a r p p r e p a r a t i o n s and t h u s mapping i t s d i s t r i b u t i o n w i t h i n t h e s e p t a l a rea i s d i f f i c u l t . To overcome t h i s d i f f i c u l t y the d i s t r i b u t i o n o f raphe p r o j e c t i o n s t o t h e septum was s t u d i e d u s i n g a u t o r a d i o g r a p h i c t r a c i n g o f amino a c i d s and t h e 5-HT p r e c u r s o r 5-HTP (Conrad et a l , 1974; H a l a r i s e t a l , 1976). These s t u d i e s i n d i c a t e t h a t raphe axons run f o r w a r d i n t o the m e d i a l f o r e b r a i n bundle and r e a c h 39 the s e p t a l a r e a v i a t h e d i a g o n a l band. There i s dense i n n e r v a t i o n t h r o u g h o u t the m e d i a l s e p t a l n u c l e u s e x c e p t f o r a dense band i n i t s most l a t e r a l a s p e c t s (Moore, 1975). H a l a r i s e t a l (1976) c a u t i o n t h a t some of t h e t h e f i b r e s i n t h e m e d i a l s e p t a l n u c l e u s a r e f i b r e s of passage which c o n t i n u e i n t o the f o r n i x . Some o f t h e s e f i b r e s p r o b a b l y p r o j e c t t o the hippocampus s i n c e l e s i o n s i n t h e m e d i a l s e p t a l n u c l e u s s i g n i f i c a n t l y reduce HBP t r a n s p o r t from t h e hippocampus t o t h e median raphe (Assaf and M i l l e r , 1978). P r e v i o u s s t u d i e s have i m p l i c a t e d 5-HT i n the c o n t r o l of s e p t a l u n i t d i s c h a r g e . S t e f a n i s (1964) r e p o r t e d t h a t t h e i o n t o p h o r e t i c a p p l i c a t i o n of 5-HT d e p r e s s e s the spontaneous d i s c h a r g e of s e p t a l neurones. More r e c e n t l y , S e g a l (1976) r e p o r t e d t h a t raphe s t i m u l a t i o n r e s u l t s i n a l o n g l a t e n c y (>25 msec) i n h i b i t i o n o f s e p t a l neurones. 40 1I.IJL3 8h£thmiGal Activity. In The Se£tal-Hi£jBocamjal Axis The slow e l e c t r i c a l a c t i v i t y of the neocortex and hippocampus have been studied i n r e l a t i o n to the l e v e l of 'arousal', the sleep-waking cycle, attention and movement {Moruzzi and Magoun, 1949; Green and Arduini, 1954; Lindsley, 1960; Vanderwolf, 1972). These studies indicate that low voltage fast a c t i v i t y i n the neocortex and rhythmical slow a c t i v i t y recorded in the hippocampal formation may be considered as 'activation patterns'. The su b c o r t i c a l mechanisms underlying these patterns are not cl e a r , although there i s extensive evidence that ascending inputs from the brainstem determine the patterns of neocortical and hippocampal a c t i v i t y . , Furthermore, i t has been suggested that the rhythm for slow e l e c t r i c a l a c t i v i t y of the hippocampus i s not due to the direct action of brainstem afferents on the hippocampus but may be generated by the rhythmical bursting discharge of septal neurones which, i n turn, relay i t to the hippocampus (Petsche, Stumpf and Gogolak, 1962). The aim of the present section i s to review the studies which have led to the above concepts, , 41 Spontaneous P a t t e r n s Of Hj.£ioGaffl£al E l e c t r i c a l A c t i v i t y In the t e r m i n o l o g y o f c l i n i c a l e l e c t r o p h y s i o l o g y slow e l e c t r i c a l waves a r e d e s c r i b e d on t h e b a s i s of t h e f r e g u e n c y and p a t t e r n of t h e i r o c c u r r e n c e , a m p l i t u d e , and shape. The e l e c t r i c a l a c t i v i t y o f t h e r a t hippocampus c o n s i s t s of a m i x t u r e o f l a r g e (up t o 2mV) and s m a l l waves h a v i n g f r e g u e n c i e s r a n g i n g between 2-50 Hz. However, the o c c u r r e n c e o f t h e s e d i f f e r e n t t y p e s o f waves i s not random and p a t t e r n s which a r e dominated by c e r t a i n f r e g u e n c i e s and a m p l i t u d e s have been r e c o r d e d . The most c o n s p i c u o u s i s a r h y t h m i c a l slow a c t i v i t y (fiSA) c o n s i s t i n g of a t r a i n of r o u g h l y s i n u s o i d a l waves w i t h a f r e g u e n c y v a r y i n g between 3-12 Hz. . The f i r s t o b s e r v a t i o n of RSA was made by Jung and K o r n m u l l e r (1938) i n the r a b b i t ' s hippocampus and i n a c c o r d a n c e w i t h t h e t e r m i n o l o g y of c l i n i c a l e l e c t r o p h y s i o l o g y i s d e s i g n a t e d a * t h e t a rhythm' s i n c e i t s f r e g u e n c y ranged between 4-7 hz. A l t h o u g h i n some s p e c i e s the f r e g u e n c y of RSA i s beyond t h i s r a n g e , t h e term * t h e t a * i s s t i l l used t o denote the r e g u l a r hippocampal a c t i v i t y i ndependent o f i t s f r e g u e n c y . In c o n t r a s t t o RSA, a d e s y n c h r o n i z e d a c t i v i t y o f a m i x t u r e o f l a r g e i r r e g u l a r waves and f a s t low v o l t a g e a c t i v i t y i s a l s o r e c o r d e d from the hippocampus (Green and A r d u i n i , 1954; Vanderwolf, 1971). Both t y p e s of a c t i v i t y may be r e c o r d e d s p o n t a n e o u s l y , b u t RSA i s more 42 p r o m i n e n t l y d u r i n g s t a t e s o f a r o u s a l (Green and A r d u i n i , 1954, See s e c t i o n s 1.3.3). Sources Of HSA Although p a t t e r n s o f e l e c t r i c a l a c t i v i t y s i m i l a r t o those r e c o r d e d i n t h e hippocampus have been observed i n o t h e r r e g i o n s of the b r a i n , n o t a b l y the thalamus (Green and A r d u i n i , 1954), t h e hippocampus has been e s t a b l i s h e d as a s o u r c e of * t h e t a * s i n c e 1) a c l e a r n u l l zone and phase r e v e r s a l s have been demonstrated i n the hippocampus (Green et a l , 1960) and 2) hippocampal a c t i v i t y has been r e l a t e d t o the d i s c h a r g e o f l o c a l neurones which were o b s e r v e d t o f i r e i n b u r s t s i n phase w i t h t h e r e g u l a r slow waves (Kandel and S p e n c e r , 1961). Based on t h e above c r i t e r i a which e s t a b l i s h the s o u r c e of the e l e c t r i c a l a c t i v i t y , i t was i n i t i a l l y t h o u g h t t h e p y r a m i d a l c e i l l a y e r was the s o l e g e n e r a t o r of RSA (Green, M a x w e l l , S c h i n d l e r and Stumpf, 1960). More r e c e n t l y , two g e n e r a t o r s of RSA have been d e s c r i b e d (Winson, 1974;1975; B l a n d e t a l , 1976). One g e n e r a t o r i s l o c a t e d i n s t r a t u m o r i e n s o f CA1 and the o t h e r more v e n t r a l i n the m o l e c u l a r l a y e r o f d e n t a t e g y r u s . I f r e c o r d i n g s a r e t a k e n s i m u l a t n e o u s l y from 43 these 'independent' generators, the RSA waves are 180 degrees out of phase. The physiological s i g n i f i c a n c e and the independence of the two d i f f e r e n t generators have not yet been established. The events underlying RSA are not known. Von Euler and Green (1960) suggested that RSA i s a r e f l e c t i o n of a series of declining spikes generated within the same neurone and compared i t to 'inactivation process' described by Granit and P h i l l i p s (1956). Furthermore, they suggested that phasic afferent inputs from the septal area cause synchronously occurring 'inactivation processes' i n a population of neurones and thereby RSA. On the other hand Kandel and Spencer (1961) and Andersen et a l (1964) have suggested that i n t r i n s i c mechanisms including delayed a f t e r - p o l a r i z a t i o n of pyramidal neurones and recurrent basket i n h i b i t i o n (Gloor, 1963) favour the development of 'theta' in the presence of a steady and sustaining excitatory inputs. Although there i s s t i l l controversy over whether RSA i s an i n t r i n s i c property of the hippocampus or the result of rhythmic afferent impulses, there i s general agreement that i t i s related to the discharge pattern of septal neurones. 44 B o l e Of A s p e n d i n g Systems I n The G e n e r a t i o n Of RSA The f i r s t s u g g e s t i o n t h a t t h e p r o d u c t i o n of RSA was r e l a t e d t o a s c e n d i n g i n f l u e n c e s from t h e m i d b r a i n was made by Green and A r d u i n i (1954) who observed t h a t p e r i p h e r a l s e n s o r y s t i m u l a t i o n e l i c i t e d h ippocampal RSA and n e o c o r t i c a l d e s y n c h r o n i z a t i o n . I n a d d i t i o n , e l e c t r i c a l s t i m u l a t i o n of the r e t i c u l a r f o r m a t i o n produced RSA and n e o c o r t i c a l d e s y n c h r o n i z a t i o n s i m u l t a n e o u s l y . S i n c e H o r u z z i and Hagoun (1949) had e a r l i e r used n e o c o r t i c a l d e s y n c h r o n i z a t i o n as an i n d i c a t o r of * a r o u s a l ' . Green and A r d u i n i (1954) proposed t h a t RSA i s the 'hippocampal a r o u s a l p a t t e r n * . The i n i t i a l o b s e r v a t i o n of Green and A r d u i n i (1954) t h a t e l e c t r i c a l s t i m u l a t i o n o f the r e t i c u l a r f o r m a t i o n e l i c i t s RSA has been c o n f i r m e d by many s t u d i e s ( T o r i i , 1961; Yokata and F u j i m o r i , 1964; Kawamura, Nakamura and T o k i z a n e , 196 1; P e t s c h e e t a l , 1962; Anchel and L i n d s l e y , 1972; Hacadar e t a l , 1974). However, t h e term ' r e t i c u l a r f o r m a t i o n ' i s a f u n c t i o n a l c l a s s i f i c a t i o n o f c o l l e c t i o n s o f n u c l e i h a v i n g a c a u d a l - r o s t r a l e x t e n t from the medulla t o t h e m i d b r a i n . These n u c l e i c o n t a i n c e l l s which have l o n g d e s c e n d i n g and a s c e n d i n g axons p r o j e c t i n g t o many r e g i o n s i n c l u d i n g t h e f o r e b r a i n and c o r t e x ( B r o d a l and R o s s i , 1955; S c h e i b e l and S c h e i b e l , 1958; Nauta and Kuypers, 1958; Magni and W i l l i s , 1963). I n an attempt t o map more p r e c i s e l y t h e o r i g i n s o f t h e s e a s c e n d i n g systems 45 which e l i c i t RSA Eacadar et a l {1974) demonstrated that e l e c t r i c a l stimulation of anatomically d i s t i n c t mesencephalic and pontine regions e l i c i t e d c h a r a c t e r i s t i c hippocampal patterns. They reported that s i t e s i n r e t i c u l a r i s pontis o r a l i s , locus coeruleus, r e t i c u l a r i s g i g a n t o c e l l u l a r i s and the midbrain tegmentum e l i c i t RSA whereas stimulation of raphe nuclei and nucleus r e t i c u l a r i s pontis caudalis produced hippocampal desynchronization. T o r i i (1961) had e a r l i e r proposed that some d i s t i n c t ascending systems may produce hippocampal desynchronization whereas adjacent systems e l i c i t RSA., This proposal was based on the observation that e l e c t r i c a l stimulation i n the medic-ventral midbrain tegmentum or l a t e r a l hypothalmus produces an increase in the fast (15-30 Hz) a c t i v i t y of the rabbit hippocampus whereas stimulation i n the dorso-lateral tegmentum and medial hypothalamic region e l i c i t s RSA. , Although the above studies demonstrate that ascending inputs from the brainstem are capable of e l i c i t i n g c h a r a c t e r i s t i c hippocampal patterns they do not prove that the brainstem i s necessary for the production of RSA. Stimulation of s i t e s such as the posterior hypothalamus are also e f f e c t i v e i n producing hippocampal RSA ( T o r i i , 1961; Yakota and Fujimori, 1964; Bland and Vanderwolf, 1972). This may be due, in part, to stimulation of ascending f i b r e s coursing through the hypothalamus. However, a r o s t r a l midbrain 46 t r a n s e c t i o n a t t h e m e s o - d i e n c e p h a l i c -jun c t i o n ( h i g h c e r v e a u i s o l e ) does n e t b l o c k RSA g e n e r a t e d by s t i m u l a t i o n o f t h e p o s t e r i o r hypothalamus (Kawamura and Domino, 1968), F u r t h e r m o r e , RSA i s "recorded i n c a t s which have a c h r o n i c h i g h c e r v e a u i s o l e (Olmstead and V i l l a b l a n c a , 1977). A l t h o u g h these s t u d i e s s uggest t h a t s i t e s o t h e r than the b r a i n s t e m may p l a y a r o l e i n the g e n e r a t i o n of RSA, t h e s e a r e a s a r e not e s s e n t i a l s i n c e massive h y p o t h a l a m i c l e s i o n s do not a b o l i s h RSA (Shishaw and V a n d e r w o l f , 1973). R a t h e r , t h e s e s t u d i e s i n d i c a t e t h a t a s c e n d i n g systems which produce RSA a r e v e r y d i f f u s e and n e i t h e r t h e b r a i n s t e m nor t h e p o s t e r i o r h y p o t h a l a m i c r e g i o n s a r e a b s o l u t e l y n e c e s s a r y f o r t h e p r o d u c t i o n o f RSA. T h e r e f o r e s m a l l and l a r g e l e s i o n s of v a r i o u s r e g i o n s from the medulla t o the d i e n c e p h a l o n do not e l i m i n a t e RSA a l t o g e t h e r . The o nly e x c e p t i o n s a r e l e s i o n s i n v o l v i n g the m e d i a l s e p t a l - d i a g o n a l band a r e a . I n c h r o n i c and a c u t e p r e p a r a t i o n s e l e c t r o l y t i c l e s i o n s o f t h e m e d i a l septum a b o l i s h a l l RSA produced fay s e n s o r y s t i m u l a t i o n o r e l e c t r i c a l s t i m u l a t i o n of the v a r i o u s s i t e s d i s c u s s e d above (Green and A r d u i n i , 1954; Brucke, P e t s c h e , P i l l a t and Deisenhammer, 1959; Stumpf, 1965 f o r r e v i e w ) . Furthermore, RSA does not r e c o v e r f o l l o w i n g l a r g e s e p t a l l e s i o n s (Brugge, 1 965) and even t h e RSA n o r m a l l y a s s o c i a t e d w i t h w a l k i n g o r a c t i v e s l e e p i s a b o l i s h e d (Brugge, 1965; Kolb and Whishaw, 1977). These d a t a suggest t h a t t h e s e p t a l a r e a p l a y s a 47 c r i t i c a l r o l e i n the g e n e r a t i o n of RSA. 12.1s. Qf. The S e p t a l Area I n The G e n e r a t i o n Of RSA F o l l o w i n g t h e i n i t i a l l e s i o n e x p e r i m e n t s d e s c r i b e d above, a r e l a t i o n s h i p between t h e d i s c h a r g e p a t t e r n of s e p t a l neurones and hippocampal e l e c t r i c a l a c t i v i t y was d e s c r i b e d by P e t s c h e et a l (1962), Many c e l l s i n the m e d i a l s e p t a l - d i a g o n a l band r e g i o n f i r e i n r h y t h m i c b u r s t s which a r e synchronous w i t h RSA. The same neurones f i r e i n an i r r e g u l a r o r random manner d u r i n g hippocampal d e s y n c h r o n i z a t i o n (Petsche e t a l , 1962; Stumpf, P e t s c h e and Gogolak, 1962; Petsche e t a l , 1965; Gogolak, P e t s c h e , S t e r c and Stumpf, 1967) . R e t i c u l a r or p e r i p h e r a l s e n s o r y s t i m u l a t i o n i n i t i a t e s the b u r s t i n g d i s c h a r g e p a t t e r n o f m e d i a l s e p t a l neurones and RSA ( P e t s c h e e t a l , 1965; W i l s o n e t a l , 1976) . The mechanisms u n d e r l y i n g t h e r e l a t i o n s h i p between t h e b u r s t i n g d i s c h a r g e p a t t e r n o f s e p t a l neurones and hippocampal a c t i v i t y a r e s t i l l not clear.,McLennan and M i l l e r (1974,1976) demonstrated t h a t s e p t a l u n i t a c t i v i t y i s under hippocampal c o n t r o l . E l e c t r i c a l s t i m u l a t i o n o f the f i m b r i a , t h e major e f f e r e n t p r o j e c t i o n from t h e hippocampal f o r m a t i o n t o t h e septum, r e s u l t e d i n an a c t i v a t i o n - i n h i b i t i o n seguence o f l a t e r a l s e p t a l neurones and a s y n c h r o n i z a t i o n of the b u r s t i n g d i s c h a r g e p a t t e r n d i s p l a y e d by m e d i a l s e p t a l neurones..On t h e b a s i s o f t h e s e d a t a they suqgested 48 t h a t RSA i s not s i m p l y t h e consequence of a s e p t a l "pacemaker 1 but t h a t t h e hippocampal o u t p u t i n i t i a t e t h e b u r s t i n g d i s c h a r g e p a t t e r n of m e d i a l s e p t a l neurones v i a feedback l o o p w i t h i n t h e s e p t a l r e g i o n . These s t u d i e s have prompted a r e - e v a l u a t i o n of the r o l e o f t h e s e p t a l a r e a i n t h e g e n e r a t i o n of BSA , The o n l y e v i d e n c e t h a t the s e p t a l area may i n f a c t i n i t i a t e BSA i s based on t h e o b s e r v a t i o n s t h a t 1) e l e c t r o l y t i c l e s i o n s of the s e p t a l a r e a b l o c k BSA (reviewed above) and 2) some s e p t a l c e l l s c o n t i n u e t o b u r s t when RSA i s not p r e s e n t i n t h e v i c i n i t y o f a r e c o r d i n g e l e c t r o d e i n th e hippocampus (Petsche e t a l , 1962; Ranck, 1975). In th e model proposed by McLennan and M i l l e r (1976) s e p t a l l e s i o n s c o u l d be a b o l i s h i n g hippocampal RSA by e i t h e r d i s r u p t i n g m i d b r a i n a f f e r e n t s which may pass through the s e p t a l a r e a t o r e a c h t h e hippocampus o r op e n i n g the feedback l o o p through t h e s e p t a l a r e a . The second o b s e r v a t i o n t h a t some s e p t a l c e l l s c o n t i n u e t o b u r s t when RSA i s not p r e s e n t i s ha r d e r t o account f o r u s i n g McLennan and M i l l e r ' s (1976) model, however, i t must be remembered t h a t i n the e x p e r i m e n t s of P e t s c h e e t a l (1962) hippocampal RSA was e l i m i n a t e d by i n d u c i n g s e i z u r e s i n the hippocampus. Perhaps a s t e a d y i n p u t from hippocampus t o t h e l a t e r a l septum i s s u f f i c i e n t to cause the b u r s t i n g d i s c h a r g e o f m e d i a l s e p t a l neurones. R e g a r d l e s s of the mechanisms i n v o l v e d , i t i s apparent t h a t t h e s e p t a l a r e a i s o f c r i t i c a l i m p o r t a n c e i n t h e g e n e r a t i o n o f RSA. 49 Role Of Pharmacologically D i s t i n c t Systems In The Control Of Septal-Hippocampal A c t i v i t y Many investigations have attempted to correlate e f f e c t i v e brainstem s i t e s which e l i c i t hippocampal RSA with p a r t i c u l a r anatomical substrates ( T o r i i , 1961, 1966; Anchel and Lindsley, 1972; Macadar et a l , 1974). The r e s u l t s of these studies indicate rather diffuse systems o r i g i n a t i n g i n brainstem regions without implicating s p e c i f i c nuclei. Another approach has been to delineate these ascending systems on the basis of t h e i r neurotransmitters. By far the most studied putative transmitter i s acetycholine (Ach), but the, more recent i d e n t i f i c a t i o n of ascending monoamine systems has prompted an investigation of t h e i r role i n hippocampal and septal a c t i v i t y . h± Acetycholine Bremer and Chatonnet (1949) f i r s t observed that an i n t r a c o r o t i d i n j e c t i o n of Ach produces neocortical desynchronization i n rabbits and cats. A s i m i l a r effect was obtained by increasing the a v a i l a b i l i t y of Ach at synaptic s i t e s by i n a c t i v a t i o n of acetycholinesterase (AchE) using physostigmine (eserine) or diisopropylflurophosphate (DFP) (Bremer and Chatonnet, 1949; Rinaldi and Himwich, 1955b; Monnier and Romanouski, 1962). A possible anatomical system mediating the e f f e c t s of Ach on neocortical 'arousal' 50 was d e s c r i b e d u s i n g h i s t o c h e m i c a l s t a i n i n g f o r AchE ( K r n j e v i c and S i l v e r , 1965; Lewis and Shute, 1967; Shute and L e w i s , 1967). f u r t h e r e v i d e n c e t h a t Ach may be i n v o l v e d i n n e o c o r t i c a l a c t i v i t y was p r o v i d e d by s t u d i e s showing an i n c r e a s e d c o r t i c a l r e l e a s e o f Ach s i m u l t a n e o u s w i t h n e o c o r t i c a l d e s y n c h r o n i z a t i o n produced by e l e c t r i c a l s t i m u l a t i o n of the m i d b r a i n r e t i c u l a r f o r m a t i o n { C e l e s i a and Jasper,1966; P h i l l i s , 1968). A c h o l i n e r g i c system has a l s o been i m p l i c a t e d i n t h e p r o d u c t i o n of hippocampal RSA. , I n t r a c a r o t i d i n j e c t i o n s of Ach or DFP produce l o n g t r a i n s o f RSA i n r a b b i t s ( R i n a l d i and Himwich, 1955a) and e s e r i n e when a d m i n i s t e r e d v i a a v a r i e t y o f r o u t e s e l i c i t s h ippocampal RSA i n eve r y s p e c i e s t e s t e d ( B r u c k e , S a i l e r and Stumpf, 1950; B r a d l e y and N i c h o l s o n , 1962; Monnier and Romanouski, 1962; Stumpf e t a l , 1962; V a n d e r w o l f , 1975) . E s e r i n e a l s o i n i t i a t e s t he b u r s t i n g d i s c h a r g e p a t t e r n o f s e p t a l neurones (Stumpf e t a l , 1962). On the o t h e r hand, c h o l i n e r g i c m u s c a r i n i c a n t a g o n i s t s ( a t r o p i n e and s c o p o l a m i n e b l o c k hippocampal RSA ( B r a d l e y and N i c h o l s o n , 1962) and t h e b u r s t i n g p a t t e r n of m e d i a l s e p t a l u n i t s (Stumpf e t a l , 1962). However, Vanderwolf (1975) has r e c e n t l y r e p o r t e d t h a t movement r e l a t e d RSA (3-12 Hz) i s not b l o c k e d by a t r o p i n e whereas i m m o b i l i t y r e l a t e d (4-7 Hz) RSA i s b l o c k e d . On t h e b a s i s of t h e s e data Vanderwolf (1975) s u g g e s t s t h a t t h e r e are a t l e a s t two p h a r m a c o l o g i c a l l y d i s t i n c t forms 51 of BSA which have d i f f e r e n t f r e q u e n c i e s and r e l a t i o n s t o b e h a v i o u r . The e f f e c t s o f Ach and e s e r i n e on t h e g e n e r a t i o n o f BSA a r e dependent on an i n t a c t s e p t a l a r e a . F o l l o w i n g s e p t a l l e s i o n s e s e r i n e i s i n e f f e c t i v e i n p r o d u c i n g RSA, however, i t c o n t i n u e s to enhance the f a s t a c t i v i t y o f t h e hippocampus (Stumpf, 1965; T o r i i and H i k l e r , 1966), The e v i d e n c e f o r a c h o l i n e r g i c s e p t o - h i p p o c a m p a l p r o j e c t i o n (Reviewed i n S e c t i o n I I ) s u g g e s t s t h a t e s e r i n e may be enhancing t h e a c t i v i t y of t h i s system. E l e c t r i c a l s t i m u l a t i o n of m i d b r a i n r e t i c u l a r f o r m a t i o n o r the m e d i a l s e p t a l r e g i o n enhance Ach r e l e a s e i n t h e d o r s a l hippocampus (Smith, 1972; Dudar, 1975,1977;. F u r t h e r m o r e , e l e c t r o l y t i c l e s i o n s of t h e m e d i a l s e p t a l a r e a b l o c k t h e r e l e a s e of Ach evoked by b r a i n s t e m s t i m u l a t i o n (Dudar, 1977). A l t h o u g h t h i s s u g g e s t s t h a t the septo-hippocampal c h o l i n e r g i c system may mediate t h e e f f e c t s o f e s e r i n e on RSA they do not e l i m i n a t e the s t r o n g p o s s i b i l i t i e s t h a t 1) e s e r i n e enhances t h e a c t i v i t y o f an e x t r i n s i c c h o l i n e r g i c i n p u t t o t h e s e p t a l a r e a o r 2) e s e r i n e enhances c h o l i n e r g i c t r a n s m i s s i o n i n s y n a p t i c mechanisms i n t r i n s i c to the s e p t a l a r e a . The above s t u d i e s t aken t o g e t h e r suggest t h a t Ach i s i n v o l v e d i n t h e g e n e r a t i o n of RSA, but the a n a t o m i c a l s u b s t r a t e s f o r these a c t i o n s a r e not known. 52 Bj. N o r a d r e n a l i ne The f i r s t s u g g e s t i o n t h a t a d r e n e r g i c systems may p l a c e a r o l e i n n e o c o r t i c a l or hippocampal ' a r o u s a l * i s based on t h e o b s e r v a t i o n s t h a t i n t r a v e n o u s a d r e n a l i n e or NA produce n e o c o r t i c a l d e s y n c h r o n i z a t i o n ( R o t h b a l l e r , 1956). However, i n j e c t i o n s o f a d r e n a l i n e or NA d i r e c t l y i n t o the c e r e b r a l c i r c u l a t i o n v i a t h e c a r o t i d a r e t e r y r a t produce a s i m i l a r d e s y n c h r o n i z a t i o n ( M a n t e g a z z i n i , Paeck and S a n t i b o n e z , 1959) s u g g e s t i n g t h a t s y s t e m i c e f f e c t s may mediate the observed d e s y n c h r o n i z a t i o n . The a d d i t i o n a l o b s e r v a t i o n s t h a t v a s o p r e s s i n (Capon, 1960) and c o m p r e s s i o n of t h e d e s c e n d i n g t h o r a c i c a o r t a (Baust e t a l , 1963) i n c r e a s e b l o o d p r e s s u r e and produce n e o c o r t i c a l d e s y n c h r o n i z a t i o n have s t r e n g t h e n e d the p r o p o s a l t h a t a d r e n a l i n e and NA a c t i n d i r e c t l y t o produce c o r t i c a l ' a r o u s a l ' . More r e c e n t l y , a r o l e f o r t h e d o r s a l NA bundle o r i g i n a t i n g i n the l o c u s c o e r u l e u s (LC) i n n e o c o r t i c a l and hippocampal a r o u s a l has been p o s t u l a t e d . J o n e s e t a l (1973) e l e c t r o l y t i c a l l y l e s i o n e d LC, c a u s i n g d e p l e t i o n o f f o r e b r a i n NA and observed a d r a m a t i c d e c r ease i n waking t i m e and c o r t i c a l a c t i v a t i o n . L i k e w i s e 6-hydroxydopamine i n j e c t i o n s i n t o the LC and c a t e c h o l a m i n e d e p l e t i n g drugs decrease c o r t i c a l d e s y n c h r o n i z a t i o n and waking time ( L i d b r i n k , 1974; J o u v e t , 1974) . 53 I t has a l s o been suggested t h a t NA c o n t a i n i n g neurones o r i g i n a t i n g i n t h e LC a r e i n v o l v e d i n the p r o d u c t i o n of RSA. T h i s s u g g e s t i o n was based on t h e o b s e r v a t i o n s t h a t e l e c t r i c a l s t i m u l a t i o n of LC (Macadar e t a l , 1974) and s y s t e m i c i n j e c t i o n s o f amphetamine produce RSA ( B r a d l e y and N i c h o l s o n , 1962.; Longo, 1962; V a n d e r w o l f , 1975; A s s a f , Barbour and M i l l e r , u n p u b l i s h e d r e s u l t s ) . The e f f e c t o f amphetamine may be r e l a t e d t o i t s a b i l i t y t o i n c r e a s e r e l e a s e o f NA and dopamine or i t s d i r e c t a c t i o n on dopamine r e c e p t o r s (Moore, 1977). £*. Dop_afflin e A l t h o u g h , a t t h i s t i m e , t h e r e i s no d i r e c t e v i d e n c e , o t h e r than the e f f e c t s o f amphetamine and apomorphine (Brucke e t a l , 1957) r e l a t i n g dopamine to RSA, i t has been suggested to p l a y a r o l e i n t h e p r o d u c t i o n of c o r t i c a l " a r o u s a l 1 ( H o r n y k i e w i c z , 1966). An i n c r e a s e i n b r a i n dopamine l e v e l s f o l l o w i n g s y s t e m i c i n j e c t i o n s of L-Dopa r e s u l t s i n n e o c o r t i a l d e s y n c h r o n i z a t i o n ( M o n t e g a z z i n i and G l a s s e r , 1960; Hawer and Domaw, 1970; Thut and Rech, 1972). T h i s e f f e c t appears to be r e l a t e d t o an i n c r e a s e i n DA and not t o NA s i n c e i t i s not b l o c k e d w i t h d i s u l f i r a m which p r e v e n t s the c o n v e r s i o n o f DA to NA (Hawer and Domer, 1970). An i n d i r e c t a c t i o n mediated by the m e s o l i m b i c DA system p r o j e c t i n g t o the s e p t a l area ( L i n d v a l l , 1975; 54 Assaf and M i l l e r , 1977) i s more l i k e l y . P i . S e r o t o n i n S e r o t o n i n (5-HT) has been i m p l i c a t e d i n c o r t i c a l ' a r o u s a l * based on the i n i t i a l o b s e r v a t i o n t h a t c e r e b r a l l e v e l s o f 5-HT changes d u r i n g the d i u r n a l c y c l e ( J o u v e t , 1974). The a d d i t i o n a l o b s e r v a t i o n t h a t e l e c t r o l y t i c d e s t r u c t i o n of t h e m i d b r a i n raphe n u c l e i r e d u c e s f o r e b r a i n 5-HT and s i g n i f i c a n t l y d e c r e a s e s slow-wave s l e e p p r o v i d e d f u r t h e r s u p p o r t f o r t h e above r e l a t i o n s h i p ( J o u v e t , 1974). The r o l e o f 5-HT i n t h e c o n t r o l o f hippocampal e l e c t r i c a l a c t i v i t y i s a l s o c o n t r o v e r s i a l . Domer and Longo (1962) f i r s t r e p o r t e d t h a t tha i n t r a v e n o u s i n f u s i o n o f t h e 5-HT p r e c u r s o r 5 - h y d r o x y t r y p t o p h a n r e s u l t e d i n hippocampal d e s y n c h r o n i z a t i o n . More r e c e n t l y , Macadar et a l (1974) r e p o r t e d t h a t e l e c t r i c a l s t i m u l a t i o n i n t h e r e g i o n o f t h e m i d b r a i n raphe i n c a t s r e s u l t s i n d e s y n c h r o n i z a t i o n o f hippocampal a c t i v i t y . I n c o n t r a s t , s t i m u l a t i o n o f t h e same s i t e i n the r a b b i t produces c o r t i c a l a c t i v a t i o n and hippocampal BSA ( P o l e and Monnier, 1970), However, t h e l a t t e r s t u d y d i d not r e l a t e t h e e f f e c t s of s t i m u l a t i o n i n the r e g i o n o f the raphe t o 5-HT l e v e l s , i n f a c t , l a r g e doses of m e t h y l s e r g i d e (10-20 mg/kg,i, p.) and p-CPA (500 mg/kg i. p . ) do not b l o c k t h e a b i l i t y o f s t i m u l a t i o n i n t h e r e g i o n of the raphe t o e l i c i t BSA ( B o b i n s o n , 1978). 55 T h e r e f o r e , some of t h e s e observed s y n c h r o n i z i n g e f f e c t s may be due t o n o n - s e r o t o n e r g i c systems p a s s i n g a d j a c e n t t o t h e raphe n u c l e i . Other T r a n s m i t t e r s G l u t a m i c a c i d may be i n v o l v e d i n c o r t i c a l a c t i v a t i o n s i n c e t h e r e l e a s e o f g l u t a m i c a c i d from the n e o c o r t e x i s reduced f o l l o w i n g l e s i o n s of the r e t i c u l a r f o r m a t i o n ( J a s p e r , Khan and E l l i o t , 1965) and i n c r e a s e d f o l l o w i n g e l e c t r i c a l s t i m u l a t i o n of s i t e s i n the r e t i c u l a r f o r m a t i o n ( J a s p e r and Koyama, 1969). Although g l u t a m i c a c i d has been p o s t u l a t e d as t h e t r a n s m i t t e r of t h e p e r f o r a n t path i n p u t t o t h e hippocampus (Nadler e t a l , 1 9 7 6 ) , i t has not been i m p l i c a t e d i n the g e n e r a t i o n of RSA. E l e c t r i c a l s t i m u l a t i o n or t r a n s e c t i o n of t h e p e r f o r a n t p a th d i d not d i s r u p t the g e n e r a t i o n o f RSA (Adey, M e r i l l e s and Sunderland,1956; Andersen, B l a n d , Myhrer and S c h w a r t z k r o i n , 1977). The i n h i b i t o r y amino a c i d gamma-aminobutyric a c i d (GABA) may a l s o be i n v o l v e d i n t h e g e n e r a t i o n of RSA. McLennan and M i l l e r (1974a) have shown t h a t the b u r s t i n g d i s c h a r g e p a t t e r n o f m e d i a l s e p t a l neurones was d i s r u p t e d by e l e c t r o p h o r e t i c a p p l i c a t i o n of the GABA a n t a g o n i s t b i c u c u l l i n e . I n a d d i t i o n , t h e s y s t e m i c i n j e c t i o n of b i c u c u l l i n e d i s r u p t s the b u r s t i n g d i s c h a r g e of s e p t a l neurones ( S e g a l , 1976) and hippocampal RSA (Assaf and M i l l e r , u n p u b l i s h e d o b s e r v a t i o n s ) . The 56 p o s s i b i l i t y t h a t GABA mediates t h e r e c u r r e n t i n h i b i t i o n of hippocampal neurones (Andersen e t a l , 1964b) s u q g e s t s t h a t i t may be i n v o l v e d i n t h e g e n e r a t i o n o f RSA. R e l a t i o n s h i p Of Ser>tal An d Hippocampal A c t i v i t y To-Behaviour Hippocampal RSA has been r e l a t e d t o changes i n o v e r t movement (Van d e r w o l f , 1969) and t o i n f e r r e d p r o c e s s e s i n c l u d i n g i n f o r m a t i o n p r o c e s s i n g (Routtenberg and K r a m i s , 1968), a t t e n t i o n (Bennet e t a l , 1973), reward and punishment (Gray, 1970), m o t i v a t i o n ( K o n o r s k i et a l , 1968; Grastyan e t a l , 1966) and l e a r n i n g ( E l a z a r and Adey, 1967; Winson, 1978 ). Vanderwolf (1969) f i r s t o bserved t h a t c e r t a i n o v e r t motor b e h a v i o u r s of r a t s were c o n s i s t e n t l y accompanied by RSA, w h i l e o t h e r b e h a v i o u r s were not (Vanderwolf e t a l , 1975 f o r r e v i e w ) . . I n the f r e e l y moving r a t v o l u n t a r y b e h a v i o u r s such as w a l k i n g , swimming and m a n i p u l a t i o n of o b j e c t s are accompanied by 7-12 Hz RSA, w h i l e i m m o b i l i t y o r * s t e r e o t y p e d * b e h a v i o u r s such as chewing, u r i n a t i o n and p e l v i c t h r u s t i n g a r e accompanied by i r r e g u l a r hippocampal a c t i v i t y (tfishaw and V a n d e r w o l f , 1973) . One major e x c e p t i o n t o the r e l a t i o n s h i p between o v e r t motor b e h a v i o u r and RSA i s a c t i v e (REM) s l e e p s i n c e the hippocampal e l e c t r i c a l a c t i v i t y c o n s i s t s o f RSA (Vanderwolf, 1971; H a r p e r , 1971) . 57 The above h y p o t h e s i s r e l a t i n g hippocampal e l e c t r i c a l a c t i v i t y t o movement i s s t i l l c o n t r o v e r s i a l . Bennet (1969) and o t h e r s (Feder and Ranck, 1975) have observed t h a t movements such as w a l k i n g and l e v e r p r e s s i n g are not always accompanied by RSA. However, Whishaw and Vanderwolf (1973) have suggested t h a t the f a i l u r e t o o b s e r v e a c l o s e r e l a t i o n s h i p between RSA and w a l k i n g or l e v e r p r e s s i n g i s due t o an i n t e r a c t i o n between t h e e l e c t r o d e placement and type o f response such t h a t f a s t a c t i v i t y g e n e r a t e d i n n e i g h b o u r i n g s i t e s c a n c e l out RSA i n the immediate v i c i n i t y of the e l e c t r o d e . Another c h a l l e n g e t o t h e movement h y p o t h e s i s a r i s e s from Klemm's (1970,1971,1976) o b s e r v a t i o n s t h a t RSA can be r e c o r d e d d u r i n g i m m o b i l i t y . More r e c e n t l y , Vanderwolf et a l (1975), p r i m a r i l y on t h e b a s i s of p h a r m a c o l o g i c a l data ( r e v i e w e d b e l o w ) , have proposed t h a t t h e r e e x i s t s two d i f f e r e n t a s c e n d i n g systems to t h e hippocampus, one p r o d u c i n g low f r e g u e n c y RSA d u r i n g i m m o b i l i t y (Type I) and t h e o t h e r h i g h e r f r e g u e n c y RSA which i s c o r r e l a t e d w i t h movement (Type I I ) . Another approach t h a t has been taken t o r e l a t e t h e s e p t a l - h i p p o c a m p a l a x i s t o b e h a v i o u r i s t h a t of r e c o r d i n g the a c t i v i t y o f s i n g l e hippocampal and s e p t a l neurones and a t t e m p t i n g t o r e l a t e d i s c h a r g e p a t t e r n s t o o v e r t and i n f e r r e d p r o c e s s e s . Ranck (19 7 3 ) , r e c o r d i n g s i n g l e u n i t s i n t h e d o r s a l hippocampal f o r m a t i o n of u n r e s t r a i n e d r a t s , d i v i d e d t h e r e c o r d e d neurones i n t o two groups on the b a s i s of t h e i r f i r i n g r e p e r t o i r e . One 58 group, which he c a l l e d 'complex s p i k e c e l l s ' , f i r e a b u r s t c o n t a i n i n g d i f f e r e n t s i z e s p i k e s and t h e o t h e r t y p e was c a l l e d ' t h e t a c e l l ' s i n c e i t f i r e d s i m p l e a c t i o n p o t e n t i a l s which were r e l a t e d t o ongoing RSA. Ranck (1973,1975) o b s e r v e d t h a t ' t h e t a ' c e l l s i n c r e a s e t h e i r f i r i n g r a t e d u r i n g hippocampal RSA and p o i n t e d out t h a t V a n derwolf's (1971) d e s c r i p t i o n of ' v o l u n t a r y movement' r e l a t e d RSA i s t h e b e s t d e s c r i p t i o n f o r the r e l a t i o n s h i p of ' t h e t a * c e l l s t o b e h a v i o u r . I n c o n t r a s t 'complex s p i k e ' c e l l s had no s i m p l e r e l a t i o n t o RSA, but t h e r e was a c o r r e l a t i o n between t h e f i r i n g of these c e l l s and t h e b e h a v i o u r of the r a t . The most common r e l a t i o n s h i p between t h e f i r i n g of a 'complex s p i k e ' c e l l and b e h a v i o u r was an i n c r e a s e i n d i s c h a r g e r a t e d u r i n g approach and s u c c e s s f u l a p p e t i t i v e b e h a v i o u r . S i m i l a r r e c o r d i n g i n the s e p t a l a r e a a l s o r e v e a l a r e l a t i o n s h i p between t h e d i s c h a r g e p a t t e r n o f s e p t a l neurones and ongoing b e h a v i o u r . I n the m e d i a l s e p t a l n u c l e u s two major t y p e s o f c e l l s were i d e n t i f i e d ' t h e t a c e l l s * which i n c r e a s e t h e i r f i r i n g r a t e o n l y d u r i n g hippocampal RSA and ' t i g h t group c e l l s * whose f i r i n g o c c u r r e d d u r i n g s p e c i f i c consumatory b e h a v i o u r s (Ranck (1974).,The f i r i n g p a t t e r n o f * t h e t a c e l l s ' r e c o r d e d by Ranck (1974) s u g g e s t s t h a t t h e y may be the same as B-neurones r e c o r d e d by P e t s c h e , Stumpf and Gogolak (1962) i n the r a b b i t and by A s s a f and M i l l e r (1978) i n t h e r a t . O t her s t u d i e s r e l a t i n g the d i s c h a r g e of s e p t a l neurones t o b e h a v i o u r a l or p h y s i o l o g i c a l s t a t e s suggest 59 an increase i n discharge rate during dehydration and sensory stimulation(Hayat anf Feldman, 1974), At t h i s time, the relationships between the a c t i v i t y of the septal-hippocampal axis are not understood s u f f i c i e n t l y to allow an appreciation of the role of the septum and hippocampus i n behaviour. However, several theories which try to account for seme of the above observations have been proposed. Vinogradova (1975) postulated that the hippocampus receives generalized signals through r e t i c u l o s e p t a l input which do not contain q u a l i t a t i v e information about s t i m u l i reaching the hippocampus. The 'theta* bursts of septal neurones rhythmically modulate the dendriti c a c t i v i t y of hippocampal neurones creating the conditions when comparison or matching of inputs w i l l be possible only for signals which come in s t r i c t l y determined time guanta. Nadel and 0*Keefe (1974) suggested that the type of information which the hippocampus processes i s s p a t i a l . According to t h i s theory, the hippocampus functions as a cognitive map which i s used by the animal to move to s p e c i f i c locations i n i t s environment and to solve problems concerning s p a t i a l r e l a t i o n s between objects i n the environment. This theory accounts, in part, for d e f i c i t s i n s p a t i a l learning following hippocampal and septal lesions {Hinson, 1978) and the r e l a t i o n s h i p between the discharge pattern of hippocampal neurones 60 and unique places i n the animal's s p a t i a l environment (Nadel and O'Keefe, 1974). 61 IA.2 The P r e s e n t Study On the b a s i s o f t h e e v i d e n c e r e v i e w e d above d i f f e r e n t p a t t e r n s of hippocampal e l e c t r i c a l a c t i v i t y a r e i n i t i a t e d by a n a t o m i c a l l y d i f f u s e r e g i o n s i n t h e b r a i n s t e m . The medial s e p t a l a r e a i s c r i t i c a l l y i n t e r p o s e d between t h e b r a i n s t e m and t h e hippocampal f o r m a t i o n and may mediate RSA. Meuroanatomical and n e u r o c h e m i c a l e v i d e n c e f o r monoamine p r o j e c t i o n s t o the s e p t a l a r e a and the hippocampus suggest t h a t t h e p u t a t i v e t r a n s m i t t e r s s e r o t o n i n (5-HT) and N o r a d r e n a l i n e (NA) may be i n v o l v e d i n the g e n e r a t i o n of d i s t i n c t p a t t e r n s o f s e p t a l - h i p p o c a m p a l a c t i v i t y . The f i r s t p a r t of t h i s t h e s i s examines 1) the r o l e o f the s e p t a l a r e a i n the g e n e r a t i o n of hippocampal e l e c t r i c a l a c t i v i t y i n t h e urethane a n a e s t h e t i z e d r a t ( c h a p t e r 3) 2) the r o l e of t h e 5-HT p r o j e c t i o n from the median raphe n u c l e u s i n t h e c o n t r o l o f s e p t a l - h i p p o c a m p a l a c t i v i t y ( c h a p t e r 4) and 3) whether t h e d o r s a l NA bundle from the l o c u s c o e r u l e u s produces r h y t h m i c a l hippocampal a c t i v i t y ( c h a p t e r 5 ) . A second f e a t u r e of the hippocampal f o r m a t i o n t h a t w i l l be e x p l o r e d i n t h i s t h e s i s i s t h e p o t e n t i a t i o n of n e u r o n a l t r a n s m i s s i o n between the e n t o r h i n a l c o r t e x and t h e d e n t a t e g y r u s . P r e v i o u s s t u d i e s have i n d i c a t e d t h a t c o n d i t i o n i n g s t i m u l a t i o n of the p e r f o r a n t path (PP) , t h e e n t o r h i n a l p r o j e c t i o n t o t h e d e n t a t e , r e s u l t s i n a marked p o t e n t i a t i o n of t h e f i e l d r e s p o n s e s evoked by a 62 subsequent t e s t v o l l e y . However i t i s not known how the a c t i v i t y of o t h e r a f f e r e n t s t o the d e n t a t e such as monoamine systems a f f e c t PP-evoked f i e l d r e s p o n s e s . The second p a r t of t h i s t h e s i s was t h e r e f o r e d e s i g n e d t o : 1) c h a r a c t e r i z e t h e f i e l d p o t e n t i a l s r e c o r d e d i n t h e d e n t a t e gyrus f o l l o w i n g PP s t i m u l a t i o n and t o s t u d y the f e a t u r e s of homosynaptic p o t e n t i a t i o n of t h i s i n p u t ( c h a p t e r 6) and 2) d e t e r m i n e i f e x t r i n s i c a f f e r e n t s , p a r t i c u l a r l y the monoamine p r o j e c t i o n s d e s c r i b e d above, a l t e r the f i e l d r e s p o n s e s evoked i n t h e d e n t a t e by a t e s t PP v o l l e y ( c h a p t e r s 7-9). I n c h a p t e r 7 a l a m i n a r p r o f i l e o f the c o m m i s s u r a l i n p u t t o the d e n t a t e w i l l be p r e s e n t e d i n o r d e r t o v e r i f y p r e v i o u s s u g g e s t i o n s t h a t t h i s i n p u t t e r m i n a t e s a d j a c e n t t o PP synapses i n the m o l e c u l a r l a y e r o f t h e d e n t a t e g y r u s . The e f f e c t s of c o n d i t i o n i n g s t i m u l a t i o n of t h i s i n p u t on f i e l d r e s p o n s e s evoked by t h e PP w i l l be compared t o r e s u l t s o b t a i n e d f o l l o w i n g c o n d i t i o n i n g s t i m u l a t i o n of t h e monoamine systems which do not p r o j e c t t o the area of the PP synapses. T h i s approach may be u s e f u l i n d e t e r m i n i n g whether h e t e r o s y n a p t i c p o t e n t i a t i o n of n e u r o n a l t r a n s m i s s i o n i s a f e a t u r e of t h e hippocampal f o r m a t i o n . 63 ZsSL GENERAL METHODS 2., 1 S u r g i ca 1 P r e p a r a t i o n : Male W i s t a r r a t s (Roodlyn Farms, Guel p h , O n t a r i o ) w e i g h i n g between 250-400 g were used f o r a c u t e r e c o r d i n g e x p e r i m e n t s . The a n i m a l s were a n a e s t h e t i z e d w i t h i n t e r p e r i t o n e a l i n j e c t i o n s o f e t h y l carbamate (urethane 1.0-1.5g/kg) and su p p l e m e n t a l doses t o produce a s t a t e o f l i g h t a n a e s t h e s i a were g i v e n as ne c e s s a r y though a p r e v i o u s l y i m p l a n t e d j u g u l a r c a n n u l a . Body temperature was monitored by a r e c t a l t h e r m i s t o r probe and m a i n t a i n e d between 36.5-37 C u s i n g a h e a t i n g pad r e g u l a t e d by a temperature c o n t r o l u n i t (EKEG E l e c t r o n i c s ) . The a n i m a l s were p l a c e d i n a Kopf s t e r e o t a x i c frame (Model 1204) w i t h the head r i g i d l y f i x e d by t h e i n c i s o r bar at 4.5-5.0 mm below z e r o t h e r e b y p o s i t i o n i n g the s k u l l i n the h o r i z o n t a l p l a n e . A m i d l i n e i n c i s i o n a p p r o x i m a t e l y 15 mm i n l e n g t h was made thro u g h the s c a l p and s k i n f l a p s were r e t r a c t e d and h e l d w i t h mosquito f o r c e p s . The s u r f a c e o f t h e s k u l l was then s c r a p e d and d r i e d r e v e a l i n g bregma, lambda and t h e s a g g i t a l s u t u r e . One o r two p i e c e s of bone (7x7 mm) were removed and t h e dura i n c i s e d , so t h a t the u n d e r l y i n g c o r t i c a l t i s s u e was exposed o v e r a r e a s r o u g h l y c o r r e s p o n d i n g t o b o u n d a r i e s 2.0 mm a n t e r i o r t o bregma, 4.0 mm p o s t e r i o r t o bregma and 4.0 mm on e i t h e r 64 s i d e of the s a g g i t a l s u t u r e f o r f o r e b r a i n placements and t o bo u n d a r i e s 2.0 mm a n t e r i o r t o lambda, 3.0 mm p o s t e r i o r t o lambda and 3.0 mm on e i t h e r s i d e o f t h e s a g g i t a l s u t u r e f o r placements i n the b r a i n s t e m . Extreme c a r e was t a k e n not to damage the s u p e r i o r s a g g i t a l s i n u s when removing the bone or i n c i s i n g t h e dura. The exposed c o r t e x was co v e r e d w i t h warm s a l i n e t o keep t h e a r e a moist t h r o u g h o u t the e x p e r i m e n t s . When p o o l i n g o f s a l i n e was not p o s s i b l e warm m i n e r a l o i l was used. 2j>_2 S t i m u l a t i n g And E e c o r d i n g P r o c e d u r e s S t i m u l a t i n g e l e c t r o d e s were s t e r e o t a x i c a l l y l o wered i n t o v a r i o u s r e g i o n s , which a r e d e t a i l e d i n subsequent method s e c t i o n s , u s i n g a Kopf e l e c t r o d e h o l d e r . F o r m i c r o e l e c t r o d e placements a Kopf h y d r a u l i c m i c r o d r i v e (Model 607W) was used t o lower t h e e l e c t r o d e i n 1 uM s t e p s . The a n t e r i o r - p o s t e r i o r (AP) s t e r e o t a x i c z e r o f o r a l l f o r e b r a i n e l e c t r o d e p lacements was measured from t h e j u n c t i o n o f t h e s a g g i t a l s u t u r e and bregma w h i l e b r a i n s t e m placements were measured from s t e r e o t a x i c z e r o at t h e t i p o f t h e ear bar. A l l l a t e r a l placements were t a k e n from the m i d l i n e p o s i t i o n o f the s a g i t t a l s i n u s and v e r t i c a l c o o r d i n a t e s were a l l measured from the s u r f a c e of the c o r t e x . The p o s i t i o n s f o r a l l t h e s e placements were m o d i f i e d from t h e Konig and K l i p p e l (1963) a t l a s of the r a t b r a i n . 65 C o n c e n t r i c b i p o l a r e l e c t r o d e s (SNE 100, Rhodes i n s t r u m e n t s ) h a v i n g a t i p s e p a r a t i o n of 0.5 mm and BC r e s i s t a n c e i n normal s a l i n e o f 75-100 K ohms were used f o r e l e c t r i c a l s t i m u l a t i o n . S i n g l e and r e p e t i t i v e monophasic r e c t a n g u l a r p u l s e s were d e l i v e r e d by an i s o l a t e d s t i m u l a t o r ( D i g i t i m e r t y p e 2533) and u n l e s s s p e c i f i e d o t h e r w i s e p u l s e parameters were 0.05-0.1 msec i n d u r a t i o n and 1-20 V i n t e n s i t y (20-400 uA) . The s t i m u l a t o r s were c o n t r o l l e d by a f o u r c h a n n e l programmer ( D i g i t i m e r Model D4030).,The c u r r e n t s p r e a d from th e s e e l e c t r o d e s was e s t i m a t e d t o range between 0.15-0.5 mm from the t i p when s t i m u l a t i n g w i t h i n t e n s i t i e s o f 10-100uA (Bagshaw and Evans, 1976). S i n g l e U n i t A c t i v i t y And Evoked P o t e n t i a l s E x t r a c e l l u l a r u n i t a c t i v i t y and evoked f i e l d p o t e n t i a l s were rec o r d e d u s i n g g l a s s m i c r o p i p e t t e s c o n t a i n i n g 4M NaCl h a v i n g t i p diameters of 1-2 uM and DC r e s i s t a n c e between 2-5 M ohms. These m i c r o e l e c t r o d e s were prepar e d from C o r n i n g pyrex c a p i l l a r y t u b i n g ( o u t s i d e d i a m e t e r 1. 5 mm) which were heated and p u l l e d i n a C a n b e r r a - t y p e m i c r o e l e c t r o d e p u l l e r and the r e s u l t a n t t i p s were broken back t o the d e s i r e d w i d t h under m i c r o s c o p i c o b s e r v a t i o n . The e l e c t r o d e s were f i l l e d under a d i s s e c t i n g microscope by i n j e c t i n g the e l e c t r o l y t e i n t o the s h a f t of the e l e c t r o d e u s i n g a l o n g 31 gauge hypodermic n e e d l e . The s o l u t i o n was pushed f u r t h e r down the s h a f t u s i n g a pyrex probe u n t i l 66 c a p i l l a r y action drew i t to the t i p of the electrode. The microelectrode was connected to the positive pole of a custom made voltage follower using an insulated s i l v e r wire. The reference or negative pole was grounded to the stereotaxic frame. The recorded e l e c t r i c a l a c t i v i t y was led through a Tektronix 5A22N amplifier and displayed on a Tektronix D4 4 oscilloscope and Tektronix D13 dual beam storage oscilloscope. E x t r a c e l l u l a r unit a c t i v i t y was f i l t e r e d using a 100Hz-10kHz bandpass, while evoked potential recordings were f i l t e r e d using a 10Hz-3KHz bandpass. In some cases both unit a c t i v i t y and evoked potentials were recorded simultaneously on the two beams of the oscilloscope. Amplifier outputs were connected to a custom made audiomonitor. The amplified signals were also led into a voltage discriminator (HP.I Instruments, Model 120) which compared the recorded signals with a predetermined triggering potential so that action potentials having a minimum amplitude were represented by an output sig n a l . The signals from the discriminator were fed into a Schmidt trigger and the res u l t i n q pulses were subseguently led into a PDP 11/10 (D i g i t a l Corp.) computer programmed to analyze the records as described in the next section. The analyzed data were plotted on a visu a l display terminal or an i n t e r a c t i v e d i g i t a l p l o t t e r (Tektronix model 4662). 67 E l e c t r i c a l A c t i v i t y Of Tlje HiEPO£§fflEi§i EQJEfflatiGn The e l e c t r i c a l a c t i v i t y o f t h e hippocampus and d e n t a t e gyrus was r e c o r d e d u s i n g b i p o l a r s t a i n l e s s s t e e l e l e c t r o d e s { t i p separation=0,5 mm). As shown i n F i g . 2-1, the s i g n a l s r e c o r d e d from t h e s e e l e c t r o d e s were f i l t e r e d (2-30 H z ) , a m p l i f i e d and d i s p l a y e d on an o s c i l l o s c o p e and r e c o r d e d on a pol y g r a p h ( G i l s o n model MSP). One beam o f a d u a l beam s t o r a g e o s c i l l o s c o p e was used t o d i s p l a y t h e r e c o r d e d e l e c t r i c a l a c t i v i t y and the second beam was used t o d i s p l a y t h e s i m u l t a n e o u s n e u r o n a l d i s c h a r g e s i n t h e s e p t a l a r e a . A T e k t r o n i x P o l a r o i d camera (model C-5) was used t o photograph the r e s u l t i n g o s c i l l o s c o p e sweeps. 2 ..3 Data A n a l y s i s : S i B g l e U n i t A c t i v i t y S e v e r a l methods were used t o a n a l y z e t h e spontaneous d i s c h a r g e p a t t e r n of a neurone and d e t e r m i n e whether i t was i n f l u e n c e d by e l e c t r i c a l s t i m u l a t i o n of a p a r t i c u l a r r e g i o n . The spontaneous d i s c h a r g e r a t e was a n a l y z e d u s i n g a f i r s t o r d e r i n t e r v a l h i s t o g r a m (IHT) . A peak on an IHT shows a p r e f e r r e d i n t e r v a l between f i r i n g s i n a s p i k e t r a i n . S i m i l a r i l y the nth o r d e r i n t e r v a l h i s t o g r a m i s the d i s t r i b u t i o n o f t h e i n t e r v a l s between a g i v e n s p i k e and t h e nth+1 s p i k e . The sum of the s e IHT's o f o r d e r 1 t o n i s known as t h e a u t o c o r r e l a t i o n f u n c t i o n whereby the peaks i n d i c a t e the p r e f e r r e d p e r i o d s of t h e s p i k e 68 2- l i Schematic I l l u s t r a t i o n Of Extracellular Recording Technique., Slow e l e c t r i c a l a c t i v i t y i s recorded from the dentate gyrus using bipolar s t a i n l e s s s t e e l electrodes. Single unit a c t i v i t y i s recorded simultaneously from the septal area using glass micropipettes f i l l e d with 4M NaCl. The signals are amplified and displayed on an oscilloscope for photography or plotted on an x-y graphics terminal. Filter D.I-3KHZ Win Disci dow rimin. v PDP-11 Computer Dual-Beam Oscilloscope r\ A W A r \i y y \j v V "I I I 1' I vr I i "> THPI r A ,f i i t J 5 0 0 / J V 200msec Plotter 70 t r a i n . R h y t h m i c a l n e u r o n a l d i s c h a r g e p a t t e r n s r e s u l t i n an a u t o c o r r e l o g r a m d i s p l a y i n g a damped o s c i l l a t i o n whereas t h a t c o r r e s p o n d i n g t o a randomly f i r i n g neurone i s f l a t ( P e r k e l , G e r s t e i n and Moore, 1967), An a d d i t i o n a l s t a t i s t i c a l measure o f t h e spontaneous d i s c h a r g e which was used i s t h e j o i n t i n t e r v a l d e n s i t y ( R o d i e c k , U r i n g and G e r s t e i n , 1962) i n which t h e a b s c i s s a v a l u e o f each p o i n t i s t h e d u r a t i o n of a g i v e n i n t e r v a l and the c o r r e s p o n d i n g o r d i n a t e v a l u e i s t h e d u r a t i o n o f t h e subsequent i n t e r v a l . T h i s a n a l y s i s g i v e s a c l e a r i n d i c a t i o n of the b u r s t i n g d i s c h a r g e p a t t e r n o f a neurone. In s t i m u l a t i o n e x p e r i m e n t s , a po s t s t i m u l u s t i m e (PST) h i s t o g r a m was used t o a s s e s s t h e changes i n s p i k e a c t i v i t y r e s u l t i n g from s t i m u l a t i o n of a p a r t i c u l a r r e g i o n a g a i n s t a background of spontaneous f i r i n g neurones. The d i s t r i b u t i o n o f s p i k e s i n t i m e r e l a t i v e to t h e s t i m u l u s was summed over many re p e a t e d s t i m u l u s p r e s e n t a t i o n s or epochs. A peak on a PST h i s t o g r a m shows a p r e f e r r e d t i m e o f d i s c h a r g e r e l a t i v e t o the s t i m u l u s ( G e r s t e i n and K i a n g , 1960) whereas a f l a t PST i n d i c a t e s a l a c k o f r e s p o n s e t o t h e s t i m u l u s . P e s t s t i m u l u s data were a l s o d i s p l a y e d i n a r a s t e r e d form whereby each s p i k e appears as a d o t and t h e d e n s i t y of d o t s b e f o r e and a f t e r t h e s t i m u l u s i n d i c a t e s the p r o b a b i l i t y t h a t the background d i s c h a r g e r a t e o f t h e neurone was a l t e r e d by p r e s e n t a t i o n of t h e s t i m u l u s . 71 Evoked F i e l d s Evoked f i e l d potentials »ere either photographed from the storage oscilloscope or plotted when computer averaging was used. The averaging was performed by converting the analog amplifier output to d i g i t a l voltage values which were sampled in 35 microsecond bins by the PDP 11/10 computer. Twenty to t h i r t y consecutive stimulus presentations (ISI=1-5 sec) were used to obtain a t y p i c a l average response. Latencies were measured from the beginning of the stimulus a r t i f a c t s to the peak of the evoked wave (peak latency) and/or to the onset of the evoked wave (onset latency). The evoked responses were always displayed with negative p o l a r i t y upwards as in F i g . 2-2. The amplitude of each f i e l d response was measured from baseline to peak. In addition the rate of r i s e of evoked f i e l d s was measured as the net voltage change from the onset of the f i e l d to a predetermined i n t e r v a l (usually 1.0-1.5 msec) a f t e r onset. The rate of r i s e was measured whenever amplitude measurements were l i k e l y to be contaminated by spike discharges occurring near the peak of the evoked f i e l d as i n the case of EPSP's (Fig, 2-2). Special care was also taken to measure the amplitude of compound spikes, such as population spikes described i n chapter 6 , which were sometimes superimposed on evoked waves. As shown i n Fig. 2-2, the amplitude of the population spike was measured as the voltage change from maximum positive deflection to the peak negative deflection of the 72 O G J L ; 2- 2: Measurements Of Amglitudes And Rate Of Rise Of Evoked F i e l d Potentials, hz f i e l d potentials such as the e x t r a c e l l u l a r EPSP were analysed with respect to peak amplitude and rate of r i s e measured 1.0-1.5 msec after onset. B: compound spikes which were superimposed on evoked waves were analysed with respect to th e i r amplitude (maximum negativity to maximum pos i t i v i t y ) , 73 74 spike. This method of measuring population spike amplitudes was found to be as r e l i a b l e as that previously used by Steward et a l (1976) who expressed the spike amplitude as the average of the height of the ascending and descending limbs of the population spike. In addition, the present method i s more suitable for automated data analysis. a l t e r a t i o n s i n evoked responses were expressed as a percentage of the control amplitude or rate of r i s e . Bhythmical E l e c t r i c a l A c t i v i t y The slow wave a c t i v i t y of the hippocampus and dentate gyrus was analyzed with respect to i t s freguency, amplitude and shape. The e l e c t r i c a l a c t i v i t y was c l a s s i f i e d as rhythmical slow a c t i v i t y (SSa) i f i t consisted of roughly sinusoidal waves with a frequency varying from 3-12 Hz and minimum amplitudes of 0.3 mV. E l e c t r i c a l a c t i v i t y that did not meet a l l three c r i t e r i a was c l a s s i f i e d as i r r e g u l a r . No attempt was made to subclassify i r r e g u l a r a c t i v i t y , although i t s average amplitude was often noted (Fig. 3-1). When e l e c t r i c a l stimulation or drug administration was used to a l t e r the spontaneously occurring e l e c t r i c a l a c t i v i t y an event marker on a separate channel of the polygraph was used to time the onset and termination of the stimulus t r a i n or i n j e c t i o n period. The freguency of BSA was computed by manually counting the number of sinusoidal •theta* waves occurring during 1 second 75 s a m p l i n g p e r i o d s . T y p i c a l l y , r a t e s were c a l c u l a t e d b e g i n n i n g one minute b e f o r e the s t i m u l u s t r a i n u n t i l r e c o v e r y t o background a c t i v i t y . S t a t i s t i c a l A n a l y s i s : the s t a t i s t i c a l t e s t s which were used f o r comparisons o f d a t a were a l l d esigned t o d i s p r o v e the n u l l h y p o t h e s i s . F o r comparisons between two independent p o p u l a t i o n s such as the f r e g u e n c y of RSA b e f o r e and d u r i n g s t i m u l a t i o n o r p o t e n t i a t i o n of evoked r e s p o n s e s a c o n v e n t i o n a l S t u d e n t s t - t e s t was used. When comparisons were made between two dependent measures , such as whether or not a p o p u l a t i o n o f c e l l s i s i n h i b i t e d , the b i n o m i a l d i s t r i b u t i o n was used. 2 i i i L e s i o n i n g Technigues A l l l e s i o n s were performed when the a n i m a l s were a n a e s t h e t i z e d w i t h sodium p e n t o b a r b i t a l (50 mg/kg i . p , ) . The a n i m a l s were p o s i t i o n e d i n t h e s t e r e o t a x i c frame and t h e s k u l l exposed a s d e s c r i b e d i n t h e p r e v i o u s s e c t i o n s . S m a l l h o l e s were d r i l l e d i n the c a l v a r i u m (1.0-1.5 M i n diameter) and a sharp needle was used t o puncture t h e u n d e r l y i n g dura and the l e s i o n i n g e l e c t r o d e or c a n n u l a was then s t e r e o t a x i c a l l y l owered t o the d e s i r e d depth, A l e s i o n was t h e n made u s i n g one o f t h e methods d e s c r i b e d below. F o l l o w i n g the l e s i o n s t h e i n c i s i o n was sewn up and t h e a n i m a l s a l l o w e d t o r e c o v e r i n a te m p e r a t u r e c o n t r o l l e d cage. 76 E l e c t r o l y t i c L e s i o n s The l e s i o n i n g e l e c t r o d e s were made by s c r a p i n g t h e i n s u l a t i o n o f f t h e i n n e r p o l e o f an SNE-100 e l e c t r o d e under a d i s s e c t i n g microscope so t h a t 0.5-0. 75 mm o f t h e t i p was exposed. The p o s i t i v e p o l e of a Grass l e s i o n i n g d e v i c e (Model LM5A) was co n n e c t e d t o the i n n e r t i p of t h e e l e c t r o d e and the n e g a t i v e p o l e was connected to t h e t a i l o f t h e a n i m a l . A good c o n n e c t i o n between the n e g a t i v e p o l e and t h e a n i m a l was s e c u r e d by wrapping a m o i s t c o t t o n swab around t h e a n i m a l * s t a i l . The l e s i o n i n g c u r r e n t (1.0-1.5 mA) was p r e a d j u s t e d b e f o r e anodal DC c u r r e n t was passed f o r 10-15 seconds. Acute And C h r o n i c T r a n s e c t i o n s Acute t r a n s e c t i o n s were made u s i n g a m o d i f i e d s u r g i c a l b l a d e (1.0 mm X10.0 mm) which was mounted on a Kopf s t e r e o t a x i c h o l d e r . The dura o v e r l y i n g t h e i n t e n d e d c o u r s e o f t h e t r a n s e c t i o n was g e n t l y removed b e f o r e the b l a d e was s t e r e o t a x i c a l l y l o w e r e d and moved t o complete t h e t r a n s e c t i o n . To t r a n s e c t the hippocampal commissure i n the v e n t r a l p s a l t e r i u m t h e Jcnife was lowered t o a p o s i t i o n 4.5 mm p o s t e r i o r t o bregma; 0.5 mm from the m i d l i n e and 4.0 mm below t h e s u r f a c e o f t h e c o r t e x ; t h e b l a d e was the n moved p a r a l l e l t o t h e s a g g i t a l s i n u s t o a p o s i t i o n 2.0 mm a n t e r i o r t o bregma. To t r a n s e c t b r a i n s t e m a f f e r e n t s t o the f o r e b r a i n t h e s a g g i t a l s i n u s was t i e d o f f and 77 removed before the knife was lowered to a position 2.5-3.0 mm behind bregma; 4.5 mm from the midline and 2.5 mm below the surface of the cortex. As the knife was moved across the coronal plane i t was gradually lowered further reaching a depth of 7.0-8.0 mm at the midline. At t h i s point i t was gradually retracted as i t approached the c o n t r a l a t e r a l side. The procedure was necessary to account f o r the shape of the base of the s k u l l and avoid damaging the basi l a r a r t e r i e s . Chronic transection of the hippocampal commissure was performed by d r i l l i n g two small holes in the s k u l l , one positioned 1.0 mm from the midline and 2.0 mm anterior to bregma and the other 1.0 mm from the midline and 5.0 mm posterior to bregma, A s t e r i l e curved s u r g i c a l needle (6-0) with attached thread was lowered through one hole and c a r e f u l l y brought up through the other. The two ends of the thread were then gently l i f t e d thereby transecting the overlying f i b r e s . The curve of the needle assured a minimum depth of 4.5 mm at the l e v e l of the ventral psalterium. 78 Neurochemical L e s i o n s Chemicals were i n j e c t e d i n t r a c e r e b r a l l y u s i n g a Hamilton 30 gauge m i c r o s y r i n g e a t t a c h e d t o a Kcpf s t e r e o t a x i c h o l d e r . The m i c r o s y r i n g e was f i l l e d t a k i n g s p e c i a l c a r e t o e l i m i n a t e a i r b u b b l e s . The m i c r o s y r i n g e was s t e r e o t a x i c a l l y l owered v e r y s l o w l y t o m i n i m i z e t i s s u e damage. A p r e d e t e r m i n e d volume was i n j e c t e d u s i n g a micrometer a t the r a t e of 1.0 u l / 1 0 minutes. At t h e end o f the i n j e c t i o n p e r i o d t h e c a n n u l a was l e f t i n p l a c e f o r a t l e a s t 10 minutes t o a l l o w u n d i s t u r b e d d i f f u s i o n of the s o l u t i o n away from the t i p b e f o r e i t was withdrawn,. The d o r s a l n o r a d r e n e r g i c bundle (AP=+2.6; L=+1.1; V=-4.5 below c o r t e x ) was s e l e c t i v e l y d e s t r o y e d u s i n g b i l a t e r a l i n j e c t i o n s o f 6-hydroxydopamine hydrobromide (6-OHDA, R e g i s C h e m i c a l ; 6 ug d i s s o l v e d 2.0 u l of 0.15M NaCl w i t h 0.2mg/ml a s c o r b i c a c i d ) . P r e v i o u s s t u d i e s have suggested t h a t 6-OHDA i s s p e c i f i c a l l y t a k e n up by c a t e c h o l a m i n e - c o n t a i n i n g axons r e s u l t i n g i n t h e i r subsequent d e s t r u c t i o n . ( T r a n z e r and Thoenen, 1967; Sachs and J o n s s o n , 1975) the degree of n o n s p e c i f i c t i s s u e damage at t h e i n j e c t i o n s i t e r e s u l t i n g from t h e n e e d l e p e n e t r a t i o n and t h e i n j e c t i o n d i d not exceed 0.25 mm i n any p e n e t r a t i o n . C e n t r a l s e r o t o n i n (5-HT) was s e l e c t i v e l y d e p l e t e d u s i n g i n t r a g a s t r i c i n j e c t i o n s o f p - c h l o r o p h e n y l a l a m i n e (p-CPA, 400 mg/kg; P f i z e r Inc.) suspended i n 2-3 ml of 79 s a l i n e . The s u s p e n s i o n was p r e p a r e d by a d d i n g a few drops o f p o l y s o r b a t e (Tween 80) to wet t h e s o l i d . . The p-CPA was th e n c o m p l e t e l y d i s s o l v e d a t pH 11 by a d d i n g 5 M NaOH. The s u s p e n s i o n was then d i l u t e d u s i n g 5M HC1 and s a l i n e t o f i n a l pH o f 7.0. Given i n t h i s manner p-CPA has been found to b l o c k 5-HT b i o s y n t h e s i s p o s s i b l y by i n h i b i t i o n o f e n z y m a t i c t r y p t o p h a n h y d r o x y l a t i o n (Koe and Weissman, 1966) . The r e s u l t i n g d e p l e t i o n o f 5-HT i s maximal 2-4 days f o l l o w i n g t h e i n j e c t i o n . C e l l b odies i n t h e s e p t a l a rea were d e s t r o y e d u s i n g d i r e c t i n j e c t i o n s of k a i n i c a c i d (1.0-3.0 ug i n 1.0 u l of s a l i n e ) . K a i n i c a c i d i s an analogue of the e x c i t a n t amino a c i d g l u t a m a t e which has been c l a i m e d to s p e c i f i c a l l y d e s t r o y c e l l b o d i e s i n t h e a r e a of the i n j e c t i o n w i t h o u t d e s t r o y i n g axons t e r m i n a t i n g i n or p a s s i n g t h r o u g h t h e i n j e c t i o n s i t e (Olney, Ho and Rhee, 1974; McGeer and McGeer, 1977). 80 2 A 5 Neu r o c h e m i c a l Assays The e f f e c t i v e n e s s o f 6-OHDA and p-CPA i n d e p l e t i n g c e n t r a l n o r a d r e n a l i n e (NA) and s e r o t o n i n (5-HTj , r e s p e c t i v e l y , was e v a l u a t e d by a s s a y i n g t h e c o n c e n t r a t i o n s o f t h e s e s u b s t a n c e s i n t h e caudate n u c l e u s , the hippocampal f o r m a t i o n and t h e s e p t a l a r e a . At the t e r m i n a t i o n o f t h e acu t e e l e c t r o p h y s i o l o g i c a l e x p e r i m e n t s t h e anim a l s were d e c a p i t a t e d and t h e above b r a i n a r e a s promptly removed. The c o n c e n t r a t i o n o f NA was measured u s i n g a r a d i o e n z y m a t i c assay a c c o r d i n g t o the methods of C o y l e and Henry (1973). The c o n c e n t r a t i o n o f NA i n 6-OHDA t r e a t e d r a t s was compared t o l e v e l s i n an i m a l s t h a t t h a t were a l s o a n a e s t h e t i z e d and used f o r e l e c t r o p h y s i o l o g i c a l e x p e r i m e n t s as w e l l as t o v a l u e s o b t a i n e d from unoperated c o n t r o l s . S e r o t o n i n c o n c e n t r a t i o n was measured u s i n g a s p e c t r o f l u o r o m e t r i c assay a c c o r d i n g t o methods p r e v i o u s l y d e s c r i b e d by R i c h a r d s o n and J a c o b o w i t z (1973). The a n i m a l s were k i l l e d i m m e d i a t e l y a f t e r the e l e c t r o p h y s i o l o g i c a l e x p e r i m e n t s and t h e b r a i n a r e a s were d i s s e c t e d out and f r o z e n a t -40 degrees C u n t i l the f o l l o w i n g morning. They were them homogenized and assayed on a Turner model 430 spectroflourometer» 81 ZtJk Si§i2l53i£§2 Analysis The l o c a t i o n o f s t i m u l a t i n g s i t e s was marked by p a s s i n g low DC anodal c u r r e n t (10-15 uA f o r 10 seconds) t h r o u g h the s t a i n l e s s s t e e l e l e c t r o d e r e s u l t i n g i n s m a l l i r o n d e p o s i t s from t h e e l e c t r o d e t i p . The d e p o s i t s were s u b s e q u e n t l y v i s u a l i z e d as b l u e / g r e e n s p o t s i n h i s t o l o g i c a l s e c t i o n s when the p r e p a r a t i o n was p e r f u s e d w i t h a p o t a s s i u m f e r r o c y a n i d e / f o r m a l i n m i x t u r e (1g/100 m l ) . The c y a n i d e r e a c t s w i t h Fe++ r e s u l t i n g i n the P r u s s i a n b l u e r e a c t i o n p r o duct which i s e a s i l y o b served under t h e l i g h t m icroscope. M i c r o e l e c t r o d e p o s i t i o n s were determined by e i t h e r e j e c t i n g pontamine sky b l u e dye (20% w/v i n 4a NaCl) e l e c t r o p h o r e t i c a l l y (Thomas and W i l s o n , 1965) o r by b r e a k i n g o f f t h e e l e c t r o d e t i p so t h a t t h e e l e c t r o d e t r a c k c o u l d be v i s u a l i z e d a f t e r t h e f i x a t i o n ( f o r example See F i g . 3-2) . The animals were k i l l e d by an overdose o f t h e a n a e s t h e t i c agent and p e r f u s e d i n t r a c a r d i a l l y w i t h 100-200 ml of 0.9% NaCl f o l l o w e d by 100 ml of 10% b u f f e r e d f o r m a l i n , , H i s t o l o g i c a l v e r i f i c a t i o n of the e l e c t r o d e s i t e s was made by c u t t i n g 50 uM f r o z e n s e c t i o n s and s t a i n i n g them w i t h c r e s y l v i o l e t or s a f f r a n i n . 82 CHAPTER • 3s - ROLE OF THE SEPT AL • ,ARE& -1H THE GENERATION- OF • HIgPQCAMPAL ELECTRICAL ACTIVITY I n t r o d u c t i o n As r e v i e w e d i n c h a p t e r 1, the e l e c t r i c a l a c t i v i t y o f t h e hippocampal f o r m a t i o n c o n s i s t s o f a m i x t u r e of slow and f a s t waves having v a r y i n g a m p l i t u d e s which a r e r e l a t e d t o ongoing p h y s i o l o g i c a l s t a t e s . The most c o n s p i c u o u s a c t i v i t y i s a synchronous or r e g u l a r slow a c t i v i t y (RSA) which i s r e c o r d e d d u r i n g s e n s o r y s t i m u l a t i o n o r d i r e c t a c t i v a t i o n of the a s c e n d i n g m i d b r a i n r e t i c u l a r f o r m a t i o n . I n r a b b i t s , r h y t h m i c a l hippocampal a c t i v i t y has been r e l a t e d t o the d i s c h a r g e p a t t e r n o f neurones i n t h e m e d i a l s e p t a l {MS) r e g i o n . P e t s c h e e t a l (1962) observed t h a t d u r i n g RSA a p o p u l a t i o n o f MS neurones f i r e i n b u r s t s which occur d u r i n g a c e r t a i n phase o f RSA waves, whereas, d u r i n g d e s y n c h r o n i z a t i o n the same neurones f i r e i n an i r r e g u l a r o r random manner. The a d d i t i o n a l o b s e r v a t i o n t h a t e l e c t r o l y t i c l e s i o n s of the s e p t a l a r e a a b o l i s h RSA (Brucke et a l , 1959) has l e d t o t h e p r o p o s a l t h a t t h e septum 'paces' e l e c t r i c a l a c t i v i t y o f t h e hippocampal f o r m a t i o n ( P e t s c h e et a l , 1962; Stumpf, 1965). A t r u e pacemaker i n the s e p t a l area would s u p p o r t the h y p o t h e s i s t h a t RSA i s generated by p h a s i c a f f e r e n t i n p u t s which produce p e r i o d i c d e p o l a r i z a t i o n o f 83 hippocampal neurones (Ton E u l e r and Green, 1960). R e c e n t l y , McLennan and M i l l e r (1976) have p r o v i d e d e l e c t r o p h y s i o l o g i c a l e v i d e n c e t h a t , i n t h e r a t , the b u r s t i n g d i s c h a r g e p a t t e r n o f MS neurones may be a conseguence r a t h e r than the cause of hippocampal RSA. T h i s prompted a r e - e v a l u a t i o n o f t h e e v i d e n c e t h a t the MS i n i t i a t e s RSA. The purpose o f t h e p r e s e n t c h a p t e r i s t o 1) c h a r a c t e r i z e the d i s c h a r g e p a t t e r n s of s e p t a l neurones i n the u rethane a n a e s t h e t i z e d r a t and r e l a t e t h e s e p a t t e r n s t o ongoing e l e c t r i c a l a c t i v i t y o f the hippocampus and 2) d e s c r i b e mechanisms which c o u l d u n d e r l i e the observed r e l a t i o n s h i p s . 3JL2 E x p e r i m e n t a l P r o c e dures Slow e l e c t r i c a l a c t i v i t y was r e c o r d e d from the CA1 r e g i o n o f the d o r s a l hippocampus or the upper b l a d e of t h e d e n t a t e g y r u s i n 71 r a t s u s i n g t e c h n i g u e s d e t a i l e d i n c h a p t e r 2. The s i m u l t a n e o u s d i s c h a r g e of s i n g l e neurones i n t h e s e p t a l a r e a (0.2 mm a n t e r i o r t o 0.3 mm p o s t e r i o r to bregma; 0.1-0.5 mm from the m i d l i n e ; 3 . 5 -6.0 mm below the s u r f a c e o f t h e c o r t e x ) was r e c o r d e d e x t r a c e l l u l a r l y . S e v e r a l r e c o r d i n g e l e c t r o d e p e n e t r a t i o n s were made i n t o t h e s e p t a l r e g i o n and i n d i v i d u a l neurones were c l a s s i f i e d on t h e b a s i s o f t h e s t e r e o t a x i c p o s i t i o n o f the r e c o r d i n g e l e c t r o d e , the r esponse t o s t i m u l a t i o n of the f i m b r i a (McLennan and 84 K i l l e r , 1974a) and t h e d i s c h a r g e p a t t e r n of neurones a n a l y z e d a s o u t l i n e d i n c h a p t e r 2. Hippocampal e l e c t r i c a l a c t i v i t y was a l s o r e c o r d e d i n a n i m a l s which p r e v i o u s l y (3-20 days) had e i t h e r an e l e c t r o l y t i c l e s i o n (1.5 mfl ano d a l c u r r e n t f o r 10-15 seconds) or an i n t r a s e p t a l i n j e c t i o n (1-3 ug i n 1 u l of s a l i n e ) of t h e n e u r o t o x i n k a i n i c a c i d which, u n l i k e e l e c t r o l y t i c MS l e s i o n s , d e s t r o y s n e u r o n a l p e r i k a r y a w i t h o u t i n t e r f e r i n g w i t h axons o f passage (See appendix a ) . In 4 a d d i t i o n a l r a t s an a c u t e t r a n s e c t i o n was made a t t h e l e v e l of the hypothalamus t o e v a l u a t e t h e c o n t r i b u t i o n of b r a i n s t e m a f f e r e n t s t o t h e d i s c h a r g e p a t t e r n o f s e p t a l neurones, 3.3.3 R e s u l t s Spontaneous E l e c t r i c a l a c t i v i t y Of The Hipfiocamjaus JLfid The Dentate Gyjrus E l e c t r i c a l a c t i v i t y was r e c o r d e d from the CA1, p y r a m i d a l c e l l l a y e r and the upper b l a d e of the d e n t a t e g r a n u l e c e l l r e g i o n . The r e c o r d s o b t a i n e d from b o t h of t h e s e s i t e s c o n s i s t e d of e i t h e r 1) an i r r e g u l a r or d e s y n c h r o n i z e d a c t i v i t y c o n s i s t i n g of slow and f a s t waves h a v i n g v a r y i n g a m p l i t u d e s ( F i g 3-1B or 2) a r h y t h m i c p a t t e r n of slow waves having c o n s t a n t a m p l i t u d e s and f r e q u e n c i e s which ranged from 3-7 Hz ( F i g 3-1a). The l a t t e r p a t t e r n , r e f e r r e d t o as RSA or 85 : l l Patterns Of E l e c t r i c a l A c t i v i t y Recorded From The Dentate Gyrus O f A Urethane-Anaesthetized Rat. I x P i spontaneously occurring RSA shown at fast and slow sweeps., B x l l desynchronized e l e c t r i c a l a c t i v i t y recorded from the same position 3 minutes a f t e r the previous records, Ci somatosensory stimulation (indicated by the s o l i d line) s h i f t s the e l e c t r i c a l a c t i v i t y from desynchronization to RSA. I i . records obtained following the death of the animal indicating minimal contribution of e l e c t r i c a l noise. 86 87 •t h e t a ' , could a l s o be e l i c i t e d by somatosensory s t i m u l a t i o n such as p i n c h i n g the animal's t a i l ( Fig 3-1C). The p a t t e r n o f e l e c t r i c a l a c t i v i t y recorded from CA1 was always s i m i l a r to t h a t recorded from the dentate with r e s p e c t to i t s synchrony and freguency. Moreover, RSA was never recorded i n CA1, i n the absence of ESA from the dentate. For t h i s reason only r e c o r d s obtained from h i s t o l o g i c a l l y v e r i f i e d s i t e s i n the dentate ( F i g . 3-2) w i l l be presented i n the present t h e s i s . , The frequency o f RSA observed i n urethane a n a e s t h e t i z e d r a t s (3-7 Hz) i s s i g n i f i c a n t l y lower than the freguency of RSA i n f r e e l y moving r a t s (3-12 Hz) p r e v i o u s l y r e p o r t e d by Vanderwolf (1 975). S i m i l a r e f f e c t s of urethane on RSA freguency have been observed i n r a b b i t s ( Stumpf, 1965). The spontaneous occurrence of RSA was r e l a t e d to the depth of a n a e s t h e s i a , but sensory-evoked RSA c o u l d be demonstrated at a wide range of a n a e s t h e t i c doses. In order to e l i m i n a t e the p o s s i b i l i t y of any extraneous movement a r t i f a c t s which might be a s s o c i a t e d with the p a t t e r n s of e l e c t r i c a l a c t i v i t y , 5 animals p r e v i o u s l y a n a e s t h e t i z e d with urethane were then immobilized with f l a x e d i l (20 mg/kg, i.v . ) and a r t i f i c i a l l y v e n t i l a t e d . The spontaneous and sensory-evoked p a t t e r n s of e l e c t r i c a l a c t i v i t y recorded under these c o n d i t i o n s d i d not d i f f e r from those obtained i n non-immobilized r a t s . Furthermore, the c o n t r i b u t i o n of e l e c t r i c a l n o i s e generated by the 88 EIGj. 3- • 2^. Loeation Of gecording Electrode Xs. The Dentate Gyrus, y Ai A schematic drawing of the hippocampal formation showing pyramidal c e l l layer of the hippocampus proper (triangles) and the granule c e l l s of the dentate gyrus ( c i r c l e s ) , B_i Coronal section of the rat brain showing electrode positions (arrow) from which ESA was recorded in the upper blade of the dentate gyrus. Abbreviations: CA1-3 Pyramidal c e l l layer CC Corpus Callosum DG Dentate Gyrus Ffl Fimbria HF Hippocampal Fissure H Dentate Hilus 89 90 equipment and room f i x t u r e s was checked r o u t i n e l y by r e c o r d i n g from t h e a n i m a l i m m e d i a t e l y a f t e r death. As shown i n F i g 3- 1f, such a r t i f a c t s d i d not c o n t r i b u t e s u b s t a n t i a l l y t o t h e r e c o r d e d a c t i v i t y . Sep_tal O n i t A c t i v i t y _ A c t i o n p o t e n t i a l s w i t h s i g n a l t o n o i s e r a t i o s g r e a t e r than 4:1 (160:40 uV) were r e c o r d e d from 431 neurones i n t h e m e d i a l s e p t a l - d i a g o n a l band r e g i o n . A l l neurones were s p o n t a n e o u s l y a c t i v e and e x h i b i t e d d i s c h a r g e f r e q u e n c i e s r a n g i n g between 2 and 50 Hz. On t h e b a s i s of t h e i r d i s c h a r g e p a t t e r n s s e p t a l neurones were c l a s s i f i e d as e i t h e r i r r e g u l a r (I -neurones) or b u r s t i n g (B-neurones). I-neurones always d i s c h a r g e d i n an i r r e g u l a r or random p a t t e r n which d i d not e x h i b i t any c o n s t a n t phase r e l a t i o n s h i p t o the p a t t e r n s o f hippocampal e l e c t r i c a l a c t i v i t y (Fig.,3-3 A-C). B-neurones, i n a d d i t i o n t o f i r i n g i n an i r r e g u l a r manner which was always a s s o c i a t e d w i t h t h e d e s y n c h r o n i z e d p a t t e r n o f hippocampal a c t i v i t y ( F i g . 3-3 E) were a l s o observed t o d i s c h a r g e i n r h y t h m i c b u r s t s of 2-10 s p i k e s ( F i g . 3-3 D) . T h i s b u r s t i n g p a t t e r n , which o c c u r r e d e i t h e r s p o n t a n e o u s l y o r as a r e s u l t o f s e n s o r y s t i m u l a t i o n , e x h i b i t e d a c o n s t a n t f r e q u e n c y and phase r e l a t i o n s h i p t o hippocampal • t h e t a ' waves. A g i v e n B-neurone may d i s c h a r g e d u r i n g any phase of the ' t h e t a ' wave and f o r each c e l l t h e d i s c h a r g e 91 3- 3_i P . i ffergnces In The Discharge Patterns Of Septal Neurones Recorded During R M£hmical Or Desynchronized Hi££ocamDal A c t i v i t y AxB^DxIl Single oscilloscope sweeps i n which the top trace i s hippocampal e l e c t r i c a l a c t i v i t y and the lower trace the simultaneously recorded septal unit discharges. A and D were recorded during RSA whereas C and E were recorded during i r r e g u l a r a c t i v i t y . C x f i Inter-spike i n t e r v a l histograms (corresponding to A and D) indicating i r r e g u l a r discharge of the I-neurone (C) and the bursting bimodal discharge pattern of the B-neurone (F). Number of spikes in each histogram i s 2000 and the bin width i s 3 msec. 92 :93 ZIILL 2 - 4.1 Local i z at ion of B-neurones In - The-Septal-Area. Ai Coronal sections from stereotaxic atlas showing the d i s t r i b u t i o n of neurones which were i d e n t i f i e d as B-neurones.. B xc H i s t o l o g i c a l v e r i f i c a t i o n of the position of the recording electrode by ejection of pontamine sky blue dye (indicated by arrows) . Note that the top deposit was l e f t at the s i t e of non-bursting neurones i n the dosal septum and the lower deposit was l e f t where bursting neurones were f i r s t recorded in a track through the medial septal region. 00 CD < 95 d i s t r i b u t i o n as revealed by s e r i a l correllograms remained f a i r l y constant (see also Petsche et a l , 1962 Fig. 2). In contrast to the conclusions drawn by Petsche et a l (1962) for septal bursts in the rabbit, a small proportion (20-30%) of B-neurones i n the rat display an amplitude decrement within a burst. However, i n t r a c e l l u l a r recordings are necessary for a guantitative analysis and comparison of t h i s decrement to the 'inactivation response* of hippocampal pyramidal c e l l s described by von Euler and Green (1960) . Similar patterns of neuronal discharges have been described i n the rabbit. (Petsche et a l , 1962; Stumpf et a l , 1962; Apostol and Creutzfeldt, 1974), Of the 431 neurones recorded throughout the septal region in t h i s series of experiments 202 neurones (47%) were c l a s s i f i e d as I-neurones and 229 (53%) as B-neurones. However, there was a regional d i s t r i b u t i o n of B-neurones whereby a high proportion was recorded i n penetrations that were in the ventro-medial aspects of the septum (Fig. 3-4) . In addition to t h e i r c h a r a c t e r i s t i c discharge patterns, septal neurones were c l a s s i f i e d on the basis of t h e i r response to stimulation of the dentate gyrus. Of 51 B-neurones tested, 24 were antidromically activated following single pulse stimulation of the dentate at the same electrode position from which RSA a c t i v i t y was recorded (Fig. 3-5). In contrast, only one 96 f I S i . 2z S i . C h a r a c t e r i s t i c s Of A Septal B^neurone.. hz Single oscilloscope sweep displaying the spontaneous bursting pattern. B£ Antidromic a c t i v a t i o n of the same neurone i n response to stimulation of the dentate gyrus (DG). C o l l i s i o n - e x t i n c t i o n with spontaneously occurring spike i s i l l u s t r a t e d in middle trace. Ci Inhibitory response of the B-neurone following antidromic activation of DG (25 superimposed sweeps), Arrows indicate stimulus a r t i f a c t s . ^Jo.2 mV 200 msec M"0.2 mV 2 msec 0.1 mV 50 msec 98 of 96 I-neurones examined could be antidromically activated. Several c r i t e r i a were used to ve r i f y that spikes were antidromically evoked: (a) short and constant latency {1.0-3,0 msec) at threshold stimulation (3-5 v o l t s ) , (b) c o l l i s i o n - e x t i n c t i o n between a spontaneous and the evoked spike at c r i t i c a l i n t e r v a l s up to 4.0 msec, (c) the presence of IS-SD break in the configuration of the evoked spike and (d) a b i l i t y to follow high freguency stimulation (100-150 Hz), Single pulse stimulation of the fimbria also resulted i n antidromic activation of the same septal neurones with a shorter latency (0.5-1,5 msec) than that observed during stimulation of the dentate gyrus. The spontaneous discharge of B-neurones was invariably i n h i b i t e d for periods of 30-100 msec following dentate or fimbria 1 stimulation even when they were not antidromically evoked. (Fig. 3-5). This i n h i b i t i o n i s sim i l a r to that reported by McLennan and M i l l e r (1974a) and used for the i d e n t i f i c a t i o n of medial septal neurones. Two additional c h a r a c t e r i s t i c s of MS response patterns following stimulation of the fimbria or the dentate gyrus were brief low amplitude burst discharges having low amplitudes preceding the period of i n h i b i t i o n and a high probability that MS neurones would begin to burst at the end of the in h i b i t o r y period (Fig. 3-6). Although I-neurones were not antidromically activated following stimulation of the fimbria, they 99 IIS*. 3z §k Bursting pis charge Pattern Of Septal M§U£ones Following Hippocampal Stimulation. ; Az rastered display of spontaneously occurring bursts (upper raster) which are synchronized by single pulse stimulation (arrow) i n the dentate (lower raster) . Bi post stimulus time histogram of the c e l l shown i n A indicating a rhythmical discharge pattern. 001 101 could be synaptically activated (37%) or inhibited (42$) by stimulation of s i t e s that antidromically activated B-neurones. SiEEScamrjal Response Patterns After E l e c t r o l y t i c And Kainic Acid Lesions Qf The. Septal Area Fig. 3-7 compares the e l e c t r i c a l a c t i v i t y recorded from DG of unoperated control rats and rats having e l e c t r o l y t i c or kainate lesions of the septal area. In control rats, the two d i s t i n c t spontaneously occurring patterns of e l e c t r i c a l a c t i v i t y , desynchronized and BSA, were recorded. RSA was also i n i t i a t e d by somatosensory stimulation such as pinching the animal*s t a i l (T.P. in Fig. 3-7B) or e l e c t r i c a l stimulation {1-40,100 Hz) in the region of the locus coeruleus. In contrast, spontaneous or evoked RSA was not recorded in e l e c t r o l y t i c or kainate lesioned rats. However, ir r e g u l a r hippocampal a c t i v i t y was not s i g n i f i c a n t l y altered by kainate and e l e c t r o l y t i c lesions of the medial septal region (Fig. 3-7D,G), In f a c t , there were no easily detectable differences between the e l e c t r i c a l a c t i v i t y recorded from the dentate of rats having kainic or e l e c t r o l y t i c lesions of the HS. The extent of kainate and e l e c t r o l y t i c lesions that resulted in the disruption of BSA are shown i n Fig. 3-8A,B . In both cases, the damage was r e s t r i c t e d to the medial septal-diagonal band region. L o c a l i z a t i o n 102 FIG... 3- Effects Of E l e c t r o l y t i c And Kainate Lesions Of The Septal Area On fliP£2£aJ2Eal E l e c t r i c a l Act.iyity.i_ A xD AGi spontaneously occurring a c t i v i t y showing the periodic appearance of RSA only i n controls. Dit,EfH;_ effect of pinching the animal's t a i l (T.P.) during the period indicated by s o l i d l i n e s . CjJgxLl e l e c t r i c a l stimulation of the brainstem evokes RSA i n control rats only. spont. E L E C T R O L Y T I C K A I N I C G j* J 1 S E C . 0.2 mV 104 of the kainic acid l e s i o n , as indicated by degeneration of neuronal perikarya and the marked p r o l i f e r a t i o n of g l i a (Fig. 3-9) , revealed that i t extended from 0.5-0.7 mm anterior to bregma to 0.7-1.0 mm posterior to bregma. Fig. 3-9 also shows that the number of normal perikarya i n the MS of a kainate lesioned rat i s s i g n i f i c a n t l y less than in a control section taken at the same l e v e l . Inasmuch as microscopic investigation reveals l o s s of perikarya, the p o s s i b i l i t y that presynaptic terminals or axons of passage were also damaged by kainic i n j e c t i o n s can not be ruled out. In order to eliminate t h i s p o s s i b i l i t y , the efficacy of the projection from the median raphe nucleus to the dentate gyrus was examined. This projection was selected since previous anatomical studies have shown that a proportion of i t s axons project through the septal area to terminate in the dentate (Halaris et a l , 1976). As previously reported (Assaf and M i l l e r , 1978c), e f f i c a c y of transmission was the same in control and kainate lesioned rats but was markedly attenuated in rats with e l e c t r o l y t i c destruction of the septal area. RSA was only blocked by h i s t o l o g i c a l l y v e r i f i e d i n j e c t i o n s of k a i n i c acid into the septal area. Control i n j e c t i o n s into the l a t e r a l v e n t r i c l e s , which caused neuronal degeneration i n a r e s t r i c t e d region of CA3, did not abolish RSA recorded in the dentate gyrus. 105 j r 8« Extent Of E l e c t r o l y t i c And Kainic-indjuced Lesions In The Sep_tal Ar ea Ai coronal section of the forebrain at the widest extent of the e l e c t r o l y t i c lesion showing loss of tissue r e s t r i c t e d to the septal area. Bz_ section taken at the same l e v e l following the intraseptal i n j e c t i o n of k a i n i c acid {2.0 "9) • 107 3- 9^ Neuronal Degeneration Following Kainic Injections Into The Septal Area^-Aj. a photomicrograph of the medial septal nucleus taken from a control rat showinq abundance of neurones. Bj. medial septal nucleus followinq Kainic i n j e c t i o n i n d i c a t i n g loss of normal perikarya and p r o l i f e r a t i o n of g l i a . Sections were stained with cresyl v i o l e t and magnified 1 0 0 X . Control 109 T h e r e f o r e the e f f e c t s o f i n t r a s e p t a l i n j e c t i o n s were not due t o d i f f u s i o n of k a i n i c a c i d i n t o t h e v e n t r i c l e s . D i s c h a r g e P a t t e r n Of S e p t a l Neurones I n The I s o l a t e d -F o r e b r a i n In t h e p r e v i o u s s e c t i o n i t was shown t h a t an i n t a c t s e p t o - h i p p o c a m p a l a x i s i s n e c e s s a r y f o r the g e n e r a t i o n of BSA. I n o r d e r t o i n v e s t i g a t e whether t h e b u r s t i n g p a t t e r n o f s e p t a l neurones i s s y n a p t i c a l l y r e l a y e d by p h a s i c , b r a i n s t e m i n f l u e n c e s , t h e d i s c h a r g e p a t t e r n o f B-neurones was r e c o r d e d b e f o r e (n=43) and a f t e r (n=22) a complete t r a n s e c t i o n of t h e d i e n c e p h a l o n at t h e l e v e l o f the hypothalamus ( F i g . .. 3-10) . . S t a b l e r e c o r d i n g s were o b t a i n e d up t o 4 h r s a f t e r the t r a n s e c t i o n . In 4 o f 6 a n i m a l s t h e i n t e g r i t y o f t h e s a g i t t a l s i n u s and the b a s i l a r a r t e r y was p r e s e r v e d and o n l y d a t a from t h e s e a n i m a l s were i n c l u d e d i n the a n a l y s i s . B e f o r e the k n i f e c u t 31 of 43 (72%) s p o n t a n e o u s l y a c t i v e neurones r e c o r d e d i n t h e medi a l a s p e c t s of n u c l e u s m e d i a l i s s e p t i were observed t o d i s c h a r g e i n r h y t h m i c a l b u r s t s a t an average f r e g u e n c y o f 5.3±0.5 Hz. A f t e r t h e k n i f e c u t o n l y 5 of 22 neurones (23%) were r e c o r d e d i n the b u r s t i n g mode of d i s c h a r g e at a f r e g u e n c y (4,3±0.4 Hz) which i s s i g n i f i c a n t l y { p <0.01) lower than t h a t o b s e r v e d b e f o r e the k n i f e c u t . 110 EISA. I z l Q L Acute T rjinsee t i on Of F oj:ebra_i n.. hz A schematic showing the location of the knife cut (dotted line) i n r e l a t i o n to indicated anatomical landmarks. B: a saggital section of brain at about 200 um from the midline. The arrow indicates the l e v e l of the recording electrode i n the septal region. Abbreviations CA1 Pyramidal c e l l region of hippocampus. CC Corpus Callosum AC anterior Commissure CP Caudate Nucleus DG Dentate Gyrus (Fascia Dentata) FM Fimbria FX Fornix HTA Hypothalamus LM Medial Lemniscus LV Lateral V e n t r i c l e B Red Nucleus S Septal Area I l l 112 Direct a c t i v a t i o n of the brainstsm or sensory stimulation did not alter the discharge pattern of septal neurones recorded i n the is o l a t e d forebrain preparation (Fig. 3-11). On two occasions recordings were obtained from the same c e l l before and after the knife cut and as shown in Fig. 3-11C, one of those c e l l s continued to burst a f t e r the knife cut. In cases where bursting was observed i n the i s o l a t e d forebrain, the spikes within a burst were not regular as shown by the wide i n t e r v a l histogram shown i n Fig. 3-11B. Since i t has been shown that physostigmine (eserine) enhances the burst a c t i v i t y of septal neurones (Petsche et a l , 1962; Assaf and M i l l e r , unpublished r e s u l t s ) , the response of the 22 neurones i n the isolated forebrain was recorded following an intravenous i n j e c t i o n of physotygmine (0.1 mg/kg). As shown in Fig. 3-11A, 13 of the 22 neurones (591) ex i h i b i t e d the bursting mode of discharge following the in j e c t i o n . The response to eserine had a latency of 90-100 seconds and was maximal 15-20 minutes post i n j e c t i o n . 113 EIG±.. : I r l l l " DiSSkarae Patterns Of Septal Neurones Before And After Transection Of The Forebrain Aj_ The number of medial septal neurones (expressed as a percentage of the t o t a l number . of c e l l s recorded) displaying the bursting mode of discharge. E_i Interval histogram of a neurone recorded before and a f t e r the knife cut. Bursting discharge as indicated by the bimodal d i s t r i b u t i o n continued after the cut, C A discharge pattern of a neurone (bottom sweeps) and a corresponding slow potentials recorded from the same electrode (top sweeps) before and at the indicated times aft e r a transection of the forebrain, ,, Eserine was injected 15 minutes afte r the transection to re-instate bursting discharge. 115 3j__£ Discussion These data indicate a relationship between the discharge pattern of septal neurones and e l e c t r i c a l a c t i v i t y of the hippocampus in the urethane anaesthetized rat s i m i l a r to that previously shown in the rabbit (Petsche et a l , 1962; Apostol and Creutzfeldt, 1974). In addition, at least two c e l l populations i n the medial septal region may be d i f f e r e n t i a t e d on the basis of t h e i r discharge patterns and whether or not they project to the hippocampus. Discharge Pattern Of Septal Neurones I-neurones, which are not antidromically activated by hippocampal stimulation, discharge in a random or irregular manner that i s unrelated to hippocampal a c t i v i t y . In contrast B-neurones discharge i n bursts which are phase related to e l e c t r i c a l a c t i v i t y of the hippocampus and are antidromically activated by stimulation of s i t e s shown to generate BSA., Since antidromic activation of B-neurones i s followed by i n h i b i t i o n an axon c o l l a t e r a l may be postulated to activate i n h i b i t o r y interneurones or perhaps those c e l l s presently c l a s s i f i e d as I-neurones. The observation that evoked discharges precede the i n h i b i t i o n of B-neurones and that a proportion of I-neurones are synaptically activated by hippocampal stimulation supports the l a t t e r suggestion. 116 Correlations With Mor£ho103i c a 1 Studies Studies of the h i s t o l o g i c a l organization of the medial septal region (Tombol and Petsche, 1969; Andy and Stephan, 1964) also indicate a heterogenous c e l l population consisting of two fundamentally d i f f e r e n t c e l l types (1) small ovoid neurones with profusely branching axon c o l l a t e r a l s and (2) medium to large neurones whose axons leave the medial septum i n a dorsal dire c t i o n . Since the terminals of the small ovoid c e l l s could be traced to the dendrites of the larger c e l l type, Tombol and Petsche (1969) suggest these neurones are i n t r i n s i c to the septal area. On the other hand the large neurones are presumably output c e l l s projecting to hippocampal regions. The close relationship between the discharge of B-neurones and hippocampal RSA observed i n t h i s and other studies (Petsche, et a l , 1962; Gogalak et a l , 1962; Apostol and Creutzfeldt, 1974) together with the fact that only these c e l l s could be antidromically activated by stimulation of RSA generating zones i n the dentate gyrus suggests that these neurones correspond to the output c e l l s described by Tombol and Petsche (1969). The i r r e g u l a r f i r i n g I-neurones may r e f l e c t a c t i v i t y in the i n t r i n s i c c e l l population. 117 Regulation Of HiEP.ocjirap.al Activity. Petsche et a l (1962) emphasized the importance of the medial septal region as a relay station transforming influences of the brainstem into a discontinuous pulse t r a i n which i n i t i a t e s RSa in the hippocampus. The present data tends to support t h i s suggestion since destruction of the septal area abolished rhythmical e l e c t r i c a l a c t i v i t y i n the hippocampus. The a d d i t i o n a l observation that kainate i n j e c t i o n s also block RSA suggests that i n t a c t septal c e l l bodies and not d i r e c t brainstem afferents to the hippocampus are absolutely necessary for the generation of rhythmical hippocampal a c t i v i t y . Neurones in the midbrain and pons may synaptically relay bursting patterns to septal neurones. However, the observation that in the i s o l a t e d forebrain preparation septal neurones continued to f i r e in rhythmical bursts suggests that mechanisms within the forebrain, possibly the septal area i t s e l f , are capable of generating *theta* rhythm. Removal of tonic brainstem influences would explain the lower incidence of bursting discharge i n the i s o l a t e d forebrain and the lack of responses to peripheral stimulation. ; Recent observations that hippocampal RSA p e r s i s t s i n cats with chron i c a l l y isolated forebrains (Olmstead and V i l l a b l a n c a , 1977) provides further support for the proposal that generation of rhythmical a c t i v i t y i s 118 dependent on f o r e b r a i n mechanisms. B o l e Of C h o l i n e r g i c System., The i n t r a v e n o u s i n j e c t i o n o f t h e a n t i c h o l i n e -e s t e r a s e , p h y s o s t i g m i n e , e l i c i t e d t h e b u r s t i n g d i s c h a r g e p a t t e r n o f B-neurones i n i n t a c t (See a l s o P e t s c h e e t a l , 1962) and i s o l a t e d f o r e b r a i n p r e p a r a t i o n s . L i k e w i s e , a n t i c h o l i n e s t e r a s e s have p o t e n t a c t i o n s on the p r o d u c t i o n o f hippocampal BSA i n i n t a c t , c e r v e a u i s o l e and i s o l a t e d f o r e b r a i n p r e p a r a t i o n s ( B r a d l e y and N i c h o l s o n , 1962; Monnier and Bomanowski, 1962; Olsmstead' and V i l l a b l a n c a , 1977; V a n d e r w o l f , 1975). E s e r i n e c o u l d be p o t e n t i a t i n g t h e a c t i o n of c h o l i n e r g i c a f f e r e n t s t o the s e p t a l a rea or the a c t i v i t y of s e p t a l c h o l i n e r g i c neurones. The o b s e r v a t i o n t h a t f o l l o w i n g s e p t a l l e s i o n s e s e r i n e i s i n e f f e c t i v e i n p r o d u c i n g BSA ( T o r i i , 1966 Stumpf, 19 65) s u g g e s t s t h a t the a c t i o n of e s e r i n e i s dependent on s e p t a l mechanisms. S i n c e e s e r i n e p o t e n t i a t e s the b u r s t i n g o f s e p t a l neurones i n the i s o l a t e d f o r e b r a i n , i t may be enhancing t h e a c t i v i t y o f t h e c o l l a t e r a l s of B-neurones onto n e i g h b o u r i n g s e p t a l neurones. The a d d i t i o n a l o b s e r v a t i o n t h a t a t r o p i n e b l o c k s hippocampal BSA i n u r ethane a n a e s t h e t i z e d r a t s ( Vanderwolf, 1975; A s s a f and M i l l e r u n p u b l i s h e d r e s u l t s ; c h a p t e r 5) emphasizes the r o l e o f c h o l i n e r g i c systems i n the p r o d u c t i o n of r h y t h m i c a l a c t i v i t y . 119 3«_5. Summary A summary of the proposed rel a t i o n s h i p s between the septal area and the hippocampus based on the re s u l t s of the present experiments i s shown in Fig. 3-12, Stimulation of a RSA generating zone i n the dentate gyrus antidromically activates only B-neurones ind i c a t i n g that these c e l l s project to the hippocampus. Furthermore, i t i s the discharge of B-neurones but not I-neurones which shows a phase and freguency rela t i o n s h i p to RSA waves. Since the antidromic activation of B-neurones i s followed by i n h i b i t i o n , an axon c o l l a t e r a l system may be postulated to activate i n h i b i t o r y interneurones or perhaps those c e l l s presently c l a s s i f i e d as I-neurones. Recent evidence that gamma-aminobutyric acid antagonists block the bursting of septal neurones (McLennan and M i l l e r , 1974; Segal, 1976} and the i n h i b i t i o n of septal neurones e l i c i t e d by hippocampal stimulation (McLennan and M i l l e r , 1974a) suggests that t h i s compound may be the i n h i b i t o r y transmitter of the interneurones. The observations that e l e c t r o l y t i c or kainate induced lesions of the medial septal diagonal band area abolish hippocampal RSA suggests that the septal area i s c r i t i c a l for the generation of rhythmical e l e c t r i c a l a c t i v i t y i n the hippocampus. Furthermore the bursting discharge pattern of septal neurones may be mediated by l o c a l forebrain mechanisms and i s not synaptically relayed by midbrain afferents. 121 FIG.. 3-12:, Schematic I l l u s t r a t i o n Of The Proposed £Slationshijg Bet ween The Brainstem x Septal Area And The Hi££ocam£al Formation The septal area transforms randomly a r r i v i n g signals from the brainstem into rhythmical bursts which are relayed to the hippocampal formation by B-neurones and in turn generate BSA. , Stimulation of the dentate ant idromically activates B-neurones and subsequently i n h i b i t s t h e i r spontaneous discharge via recurrent c o l l a t e r a l s onto interneurones in the septal area. o v A i -Neuronel O B-Neurone +V V -6 Hippocampal Formation Septal Area Brainstem 123 CHAPT EE 4 i ROLE OF A BAPHJzSEROTONIN SYSTEM IN SIS-CONTROL OF SEPTAL HIPPOCAHPAL ACTIVITY I n t r o d u c t i o n I n t h e p r e v i o u s c h a p t e r i t was shown t h a t the s e p t a l a r e a , p a r t i c u l a r l y the m e d i a l r e g i o n , p l a y s a s i g n i f i c a n t r o l e i n t h e c o n t r o l of hippocampal e l e c t r i c a l a c t i v i t y . E a r l i e r e x p e r i m e n t s have s u g g e s t e d t h a t e l e c t r i c a l s t i m u l a t i o n o f a s c e n d i n g systems f r o m t h e hypothalamus and b r a i n s t e m e l i c i t c o n t r a s t i n g p a t t e r n s o f hippocampal a c t i v i t y ( T o r i i , 1961; Yokoto and F u j i m o r i , 1964; A n c h e l and L i n d s l e y , 1972; Macadar et a l , 1974), I n an attempt t o l o c a l i z e the b r a i n s t e m o r i g i n s o f t h e s e s y s t e m s , Macadar e t a l (1974) demonstrated t h a t e l e c t r i c a l s t i m u l a t i o n o f a n a t o m i c a l l y d i s t i n c t m e s encephalic and p o n t i n e r e g i o n s e l i c i t s t h e c h a r a c t e r i s t i c hippocampal p a t t e r n s . Some of t h e s e r e g i o n s c o r r e s p o n d t o h i s t o c h e m i c a l l y i d e n t i f i e d monoamine-containing n u c l e i . S i n c e r e c e n t n e u r o a n a t o m i c a l data have shown t h a t t h e s e c e l l groups have e x t e n s i v e t e r m i n a l p r o j e c t i o n s upon the septum and hippocampus (see C h a p t e r 1 f o r review) # t h e p o s s i b i l i t y a r i s e s t h a t t h e s e p h a r m a c o l o g i c a l l y d i s t i n c t systems may form the n e u r a l s u b s t a t e s t h r o u g h which p a t t e r n s o f a c t i v i t y i n t h e septum and hippocampus a r e c o n t r o l l e d . Of p a r t i c u l a r i n t e r e s t i n t h i s c h a p t e r i s t h e s e r o t o n i n f i b r e system whose c e l l b o d i e s are l o c a t e d i n 124 t h e m e s e n c e p h a l i c raphe n u c l e i . The p r e s e n c e of a s u b s t a n t i a l p r o j e c t i o n o r i g i n a t i n g i n t h e median raphe n u c l e u s (MR) and i n n e r v a t i n g t h e septum and hippocampus (Conrad,Leonard and P f a f f , 1974; H a l a r i s e t a l , 1976} t o g e t h e r w i t h t h e o b s e r v a t i o n s t h a t l e s i o n s o f t h e MR r e s u l t i n a s i g n i f i c a n t d e p l e t i o n of f o r e b r a i n s e r o t o n i n (Lorens and G u l d b e r g , 197 4) suggest t h a t a s e r o t o n e r g i c i n n e r v a t i o n o f t h e septum may be i n v o l v e d i n the c o n t r o l of sep tal-hippoc'ampal f u n c t i o n . The aim of the p r e s e n t c h a p t e r i s t o 1) examine the e f f e c t s o f e l e c t r i c a l s t i m u l a t i o n o f the MR on n e u r o n a l d i s c h a r g e s i n t h e s e p t a l a r e a and r e l a t e d hippocampal a c t i v i t y and 2) determine whether t h e observ e d e f f e c t s are dependent on s e r o t o n i n c o n t a i n i n g systems o r i g i n a t i n g i n the MR, , 4^2 E x p e r i m e n t a l P r o c e d u r e s Experiments were performed on 54 r a t s a n a e s t h e t i z e d w i t h u r e t h a n e (1.0g/kg i . p . ) . S u r g i c a l p r o c edures and r e c o r d i n g e l e c t r o d e placements i n the hippocampus and t h e s e p t a l a r e a were i d e n t i c a l t o those d e s c r i b e d i n t h e p r e v i o u s c h a p t e r . I n a d d i t i o n , a c o n c e n t r i c b i p o l a r e l e c t r o d e was s t e r e o t a x i c a l l y lowered i n t o the r e g i o n of the median raphe n u c l e u s (0.8 mm a n t e r i o r t o s t e r e o t a x i c z e r o ; 0.0-0.5 mm l a t e r a l t o the m i d l i n e ; 6.0-6,7 mm below the s u r f a c e o f t h e c o r t e x ) . E l e c t r i c a l s t i m u l a t i o n of t h e MR c o n s i s t e d 125 o f e i t h e r s i n g l e p u l s e o f 0.05-0,1 msec d u r a t i o n and 10-20 msec d u r a t i o n and 10-20 V i n t e n s i t y o r r e p e t i t i v e v o l l e y s o f 40-100 Hz f o r 1-3 s e c at i n t e n s i t i e s of 1-9V. The s i g n a l s r e c o r d e d were f i l t e r e d (0.1-3 KHz f o r s e p t a l u n i t a c t i v i t y and 2-50 Hz f o r hippocampal a c t i v i t y ) and d i s p l a y e d on a d u a l beam o s c i l l o s c o p e and a n a l y s e d u s i n g the method o u t l i n e d i n Chapter 2. S p e c i a l c a r e had t o be ta k e n t o ensure t h a t t h e d i s c r i m i n a t o r window e l i m i n a t e d t h e c o n t r i b u t i o n c f s t i m u l u s a r t i f a c t s i n the a n a l y s i s of s e p t a l u n i t d i s c h a r g e d u r i n g r e p e t i t i v e p u l s e s t i m u l a t i o n . I n o r d e r t o d e p l e t e 5-HT, a t o t a l of 6 r a t s were p r e t r e a t e d by p a r a - c h l o r o p h e n y l a l a n i n e (p-CPA). An a d d i t i o n a l e v a l u a t i o n o f t h e r o l e o f s e r o t o n e r g i c mechanisms was made f o l l o w i n g the i n t r a v e n o u s i n j e c t i o n of t h e 5-HT p r e c u r s o r 5 - h y d r o x y t r y p t o p h a n (5-HTP) 5-60 mg/kg and the presumed a g o n i s t g u i p a z i n e (0.5-2.0 mg/kg; 2-(1-p i p e r a z i n y l ) - g u i n o l i n e s u p p l i e d by P f i z e r I n c . ) . , T h i s compound was p r e v i o u s l y shown t o a c t i v a t e c e n t r a l s e r o t o n i n - c o n t a i n i n g systems ( R o d r i g u e s , fiojas-Ramirez and D r u c k e r - C o l i n , 1973). 126 R e s u l t s S i n c e i n the p r e v i o u s c h a p t e r i t was shown t h a t a t l e a s t two p o p u l a t i o n s of s e p t a l neurones can be c l a s s i f i e d on t h e b a s i s o f t h e i r d i s c h a r g e p a t t e r n and i t s r e l a t i o n s h i p t o hippocampal RSA, t h e e f f e c t s o f MR s t i m u l a t i o n were a n a l y z e d s e p a r a t e l y f o r I-neurones and B-neurones. Response Of I-Neurones To j^aphe S t i m u l a t i o n The d i s c h a r q e p a t t e r n of a t o t a l of 202 I-neurones were examined f o l l o w i n g s i n g l e p u l s e s t i m u l a t i o n o f the MR a t s t i m u l u s i n t e n s i t i e s r a n g i n g between 10-15 v o l t s . One hundred and f i f t y - n i n e c e l l s (78.7%) were i n h i b i t e d f o r p e r i o d s o f 30-200 msec(mean=75.2±U0.5 S.D.). The d u r a t i o n o f the i n h i b i t i o n c o u l d be i n c r e a s e d c o n s i d e r a b l y i f t h e MR was s t i m u l a t e d w i t h two or more s u c c e s s i v e p u l s e s (100 Hz) a t t h r e s h o l d i n t e n s i t y or s i n g l e p u l s e s at s u p r a t h r e s h o l d (1.5 x T) i n t e n s i t y . The l a t e n c y t o the onset of the observed i n h i b i t i o n was always l e s s than 25 msec f o l l o w i n g t h e s t i m u l u s (mean=12.5 msec) and i n some cases t h e i n h i b i t i o n began i m m e d i a t e l y a f t e r the s t i m u l u s a r t i f a c t (e.g. c e l l on the r i g h t i n F i g . 4-1). H i s t o l o g i c a l v e r i f i c a t i o n of s t i m u l a t i o n s i t e s which e l i c i t e d t h e i n h i b i t o r y r e sponse i n d i c a t e d t h a t t h e y were l o c a l i z e d t o t h e MR ( F i g . 4-2B). E l e c t r o d e s i t e s i n the r e g i o n of the m e d i a l l o n g i t u d i n a l f a s c i c u l u s and the d o r s a l raphe 127 FIG.. Hz I I Response Of Septal I r n e u r o n e s To Raphe Stimulation,.. i x M i response of two neurones recorded i n the medial septal nucleus of the same animal. The top records show 25 superimposed oscilloscope sweeps and bottom records are rastered PST responses of the corresponding c e l l s . Note that i n A the onset of i n h i b i t i o n i s immediately after the stimulus a r t i f a c t ( v e r t i c a l l i n e s i n the rasters) while in B the onset latency i s longer. syood3 129 n u c l e u s were e i t h e r i n e f f e c t i v e o r sometimes e l i c i t e d s h o r t l a t e n c y (3-8 msec) s p i k e a c t i v a t i o n w i t h a s m a l l a m p l i t u d e f i e l d r e s p o n s e . When the same s t i m u l a t i n g e l e c t r o d e was low e r e d i n t o the MR p r o p e r , s e p t a l neurones p r e v i o u s l y u n a f f e c t e d were t h e n i n h i b i t e d ( F i g . 4-2 A). When the s t i m u l a t i n g e l e c t r o d e was l o c a t e d on t h e p e r i m e t e r of t h e MR, a m i x t u r e of both a c t i v a t i o n and i n h i b i t i o n seguences was observed. The t h r e s h o l d f o r the i n h i b i t o r y component o f t h e s e responses was lower t h a n t h a t f o r a c t i v a t i o n and was r e l a t e d t o the d i s t a n c e between t h e s t i m u l a t i n g e l e c t r o d e and the c e n t r e o f t h e MR . The e f f e c t s of s t i m u l a t i n g the r e g i o n of t h e d e n t a t e gyrus and CA3 a r e a o f t h e hippocampus were examined on a t o t a l of 96 I-neurones t h a t were i n h i b i t e d by MR s t i m u l a t i o n . Some neurones (41.7%) were i n h i b i t e d by both MR and hippocampal s t i m u l a t i o n , an a p p r o x i m a t e l y e g u a l p r o p o r t i o n (35.5%) were i n h i b i t e d by t h e MR and a c t i v a t e d by s t i m u l a t i o n of the hippocampus, and the remainder (20.8%) were not i n f l u e n c e d by hippocampal s t i m u l a t i o n . 130 I ISx,.- U- 2z Localization Of Stimulating Electrodes In Thelegion Of The Raphe And The Corresponding Response Of I-neurones. Az photomicrograph showing small iron deposits made at the s i t e of an i n e f f e c t i v e placement i n the dorsal raphe (top arrow) and a placement i n the median raphe that produced i n h i b i t i o n (lower arrow). Bz_ f r o n t a l sections through the raphe nucleus indicating the d i s t r i b u t i o n of stimulating s i t e s and th e i r effects on septal unit a c t i v i t y : (0) i n e f f e c t i v e , (•*) act i v a t i o n , {-) i n h i b i t i o n . ABBREVIATIONS! DR Dorsal Raphe Nucleus LH Medial Lemniscus MR Median Raphe Nucleus PCS Superior Cerebellar Peduncle 1 3 1 A 160/J 1 ^ \ P 100/J 132 Response Of B-neurones To Baghe Stimulation In contrast to the inhib i t o r y responses described above, single pulse stimulation of MB rarely influenced the discharge of i d e n t i f i e d B-neurones (Fig. 4-3). Inh i b i t i o n was observed i n only 5 of 229 B-neurones recorded during the i r r e g u l a r or bursting mode of discharge . I n c r e a s i n g the stimulus i n t e n s i t y (20-35 V) did not a l t e r the proportion of B-neurones influenced. Likewise, double pulse stimulation of the MB did not alt e r the proportion of B-neurones that were i n h i b i t e d . However, repetetive stimulation of the MR at low inten s i t y (2-9 V) and freguencies of 4 0-100 Hz resulted in the disruption of bursting which was replaced by an irregular f i r i n g pattern (Fig. 4 -4B) . Of 220 B-neurones recorded during the bursting mode, 185 (84.1%) began to f i r e in an i r r e g u l a r pattern. In the majority of cases t h i s response did not outlast the period of stimulation, however, some neurones (4 of 220) continued to f i r e in an irregular manner a f t e r the termination of MB stimulation (e.g. c a l l in Fig. 4-4 C) . The average discharge rate during the r e p e t i t i v e t r a i n was not consistently altered i n that a given neurone may f i r e with an increased or a decreased freguency during subseguent stimulus trains. The response of B-neurones described above was unigue to stimulation of MB nucleus since stimulation of adjacent areas did not result i n disruption of burst a c t i v i t y (Fig. 4-5) and sometimes changed the i r r e g u l a r 133 FIG a- 3z Characteristics Of A B^neurone During Single Pulse Stimulation Of MRA• A A rastered display of 60 successive stimulus presentations during both non-bursting (epochs 1-25) and bursting (epochs 26-60) modes of discharge, §xQl single oscilloscope sweeps showing the relationship of the same unit (lower traces) to hippocampal e l e c t r i c a l a c t i v i t y (top traces). 1 3 4 Epochs o o - i 3 00 CD O o O O * . • • • • • • • • * • i • • * • • * * • till • I | \ t" • • ... • • • « i •• • i 5 !' 'ii i ! I * • I • ««••*• I . : • ,«•: ... : «• 'j: ; - '.. ' i 1.1! * 4 : . J J * o 3 < O en 3 < 135 discharge of B-neurones into rhythmical bursts. The rhythmical bursts often outlasted the period of stimulation, persisting u n t i l the sta r t of the next t r i a l . Stimulation of s i t e s such as the medial longitudinal fasciculus or the superior cerebral peduncle also activated 1-neurones which were recorded i n the medial septal area (Fig. 4-2 and 4-5). Effe c t Qf Rajahe Stimulation On Hi2£OGameal^Jlestrijsal-•• Activity. The spontaneous e l e c t r i c a l a c t i v i t y of the hippocampus was recorded simultaneously with the discharge of septal neurones. As shown i n chapter 3, the bursting pattern of B-neurones was accompanied by BSA, Single pulse stimulation of MB which did not influence B-neurones also did not e l i c i t any detectable changes i n ongoing e l e c t r i c a l a c t i v i t y recorded from CA1, or the dentate gyrus. However, r e p e t i t i v e stimulation of MB caused a s h i f t from ongoing RSA to a fast low voltage desynchronized hippocampal a c t i v i t y (Fig. 4-4 A, 4-6C, D). The disruption of B-neurone bursting and desynchronization of e l e c t r i c a l a c t i v i t y were observed within 200 msec (average burst interval) of the stimulus presentation. However, i t was not possible to determine which occurred f i r s t . , I t i s important to note that low voltage (less than 5.0 V) stimulation of MB results i n hippocampal desynchronization independent of the mode of the 136 £22* 4- Hi III gets Qf Bejjetitiye MR Stimulation On The Bursting Discharge Pattern Of B^neuronesin Control And £:CPAtrgated. fiatsA~ A tB :_ top traces are hippocampal e l e c t r i c a l a c t i v i t y and lower traces are septal unit discharges.„MR stimulation occurs between the two arrows and the stimulus a r t i f a c t s obscure a portion of the spikes i n the lower traces. In (A) r e p e t i t i v e stimulation (3 V, 100 Hz, 600 msec) results in a desynchronization of the hippocampal a c t i v i t y and blockade of the c e l l u l a r bursting pattern. In (B) the MR stimulation i s i n e f f e c t i v e . Cz a rastered display of the discharge pattern of a B-neurone recorded i n a normal rat before and after a t r a i n of stimuli (3 V, 100 Hz, 3. 2 sec) indicated by the s o l i d white bar. Mote that, i n contrast to response i n {A), the ir r e g u l a r discharge pattern e l i c i t e d by MR outlasts the stimulus t r a i n . 138 p r e c e d i n g s p o n t a n e o u s l y o c c u r r i n g e l e c t r i c a l a c t i v i t y . S t i m u l a t i o n o f a d j a c e n t areas and sometimes the p e r i m e t e r of MR r e s u l t e d i n RSA a t low s t i m u l u s i n t e n s i t y , but at h i g h e r i n t e n s i t i e s (5-7 V) a d e s y n c h r o n i z a t i o n s y n c h r o n i z a t i o n seguence o c c u r r e d . E f f e c t s Of P-chlorqphe n y l a n i n e On Segt a l - Higp,oeam p a l A c t i v i t y P r e t r e a t m e n t with p - c h l o r o p h e n y l a n i n e (p-CPA) 3 days p r i o r t o e l e c t r o p h y s i o l o g i c a l e x p e r i m e n t s r e s u l t e d i n 60-80% d e p l e t i o n o f f o r e b r a i n 5-HT. I n t h e t r e a t e d r a t s (n=4) o n l y 7 of 24 (29.2%) I-neurones examined were i n h i b i t e d by MR s t i m u l a t i o n , compared t o 159 o f 202 (78.7%) i n c o n t r o l a n i m a l s (Table 1). The p-CPA p r e t r e a t m e n t d i d not a b o l i s h t h e s h o r t l a t e n c y e x c i t a t o r y r e s p o n s e s e l i c i t e d from s t i m u l a t i o n p o i n t s i n the v i c i n i t y of MR. Of 46 B-neurones r e c o r d e d i n t r e a t e d r a t s o n l y 9 (19.6%) were i n f l u e n c e d fay r e p e t i t i v e p u l s e s t i m u l a t i o n of t h e MR a t t h e same i n t e n s i t i e s (2-5v) which r e s u l t e d i n d i s r u p t i o n o f b u r s t i n g d i s c h a r g e i n t h e m a j o r i t y (84.1%) o f B-neurones recorded i n c o n t r o l s ( T a b l e 1) . As i n d i c a t e d by t h e b i n o m i a l d i s t r i b u t i o n , both the o c c u r r e n c e o f i n h i b i t i o n of I-neurones (p<0.001) and d i s r u p t i o n of the b u r s t i n g of B-neurones (p<0.05) were s i g n i f i c a n t l y l ower than t h o s e observed i n c o n t r o l r a t s . As shown i n F i g . 4-5, p-CPA p r e t r e a t m e n t d i d not 139 TABLE 1 COMPARISON OF SEPTAL NEURONES INFLUENCED BY MR STIMULA-TION IN CONTROL AND p-CPA TREATED RATS CELL TYPE CONTROL (n=29) p-CPA (n=4) I-NEURONES 159/202 (78.7%) 7/24 (29.2%) B-NEURONES 185/229 (84.1%) 9/46 (19.6%) p-CPA (400 mg/kg) was administered i n t r a g a s t r i c a l l y 3 days before recording experiments. Fluoremetric assays for forebrain serotonin levels were performed immediately following the recording sessions. Entries indicate number of c e l l s influenced/number of c e l l s tested (and percentages of t o t a l ) . 140 Elis. .Hz 5 i Effects Of Repetitive MR Stimulation On-MifiROcamgal E l e c t r i c a l A c t i y i t j . In Control And P.C.PA Treated Rats spontaneously occurrinq e l e c t r i c a l a c t i v i t y showing that RSA i s recorded i n both control and pCPA treated r a t s , I x f i ~ RSA recorded following a t a i l pinch <T.P.) . Cxi2.tO4.Ii effects of 4 and 5 volt stimulation {100 Hz) of the MR on hippocampal e l e c t r i c a l a c t i v i t y i n dicating that pCPA eliminates the MR evoked desynchronization. CONTROL SPONT. MM T.P. MR 4V MR 5V pCPA SPONT. rtWi^Ugiillili T.P. MR 4V H 0.6 mV MR 5V 2 Sec 142 s i g n i f i c a n t l y a l t e r t h e spontaneous e l e c t r i c a l a c t i v i t y o f u r e t h a n e a n a e s t h e t i z e d r a t s . As i n c o n t r o l r a t s 8SA o c c u r r e d s p o n t a n e o u s l y o r as a r e s u l t of somatosensory s t i m u l a t i o n such as t a i l p i n c h ( t . p . i n F i g . 4-5). However, MR s t i m u l a t i o n d i d not r e s u l t i n the low v o l t a g e d e s y n c h r o n i z e d a c t i v i t y which was observed i n c o n t r o l r a t s . I n a d d i t i o n MR s t i m u l a t i o n i n p-CPA t r e a t e d r a t s d i d not a b o l i s h ongoing RSA (Fig.4-5). I n a s e p a r a t e group o f 9 r a t s which r e c e i v e d a c e n t r a l i n j e c t i o n o f the c a t e c h o l a m i n e - s p e c i f i c n e u r o t o x i n , 6-hydroxydopamine, MR s t i m u l a t i o n r e s u l t e d i n d e s y n c h r o n i z a t i o n of RSA s u g g e s t i n g t h a t the e f f e c t s produced by p-CPA were due t o t h e l o s s of 5-HT and not n o r a d r e n a l i n e or dopamine. E f f e c t s Of Q u i p a z i n e On S e p t a l - H i p p o c a m p a l A c t i v i t y . S i x r a t s r e c e i v e d an i n t r a v e n o u s i n j e c t i o n of g u i p a z i n e (0.5-2.0 mg/kg) i n s a l i n e s o l u t i o n . I n 4 of them 0.6-1.0 mg/kg r e l i a b l y r e s u l t e d i n d i s r u p t i o n of s p o n t a n e o u s l y o c c u r r i n g RSA and a s h i f t of hippocampal a c t i v i t y t o low v o l t a g e d e s y n c h r o n i z a t i o n ( F i g , 4-6). In t h e r e m a i n i n g 2 r a t s a dose of 2.0 mg/kg was ne c e s s a r y t o produce any e f f e c t s . The o n s e t of d e s y n c h r o n i z a t i o n o c c u r r e d 2-4 minutes f o l l o w i n g t h e i n j e c t i o n and p e r s i s t e d f o r 1-3 hours. F u l l r e c o v e r y of RSA was demonstrated i n 3 r a t s which were s t u d i e d f o r up 6 hours f o l l o w i n g the i n j e c t i o n . In a l l a n i m a l s s e n s o r y - e v o k e d RSA r e c o v e r e d b e f o r e the appearance of 143 Z l ^ i . 4 - 6z Effects Of J2sA£§zine- On Hipp oca j£al E l e c t r i c a l A c t i v i t y kz_ spontaneous occurring a c t i v i t y prior to guipazine i n j e c t i o n . Bx£i a c t i v i t y recorded following the intravenous i n j e c t i o n of guipazine (1.0 mg/Kg). E>i recovery of BSA recorded 40 minutes following the i n j e c t i o n . C O N T R O L B QUIPAZINE +2min. +5min. +40min. (Recovery) 500 uV 1 sec. 145 spontaneously occurring RSI. An injection of the saline vehicle did not procduce any eff e c t s s i m i l a r to those of guipazine and a subsequent in j e c t i o n of eserine through the same cannula resulted in the appearance of RSA, The l a t t e r i n j e c t i o n v e r i f i e s that guipazine does not i r r e v e r s i b l y damage mechanisms which mediate RSA, The e f f e c t s of systemic injections of guipazine on the discharge pattern of 4 HS neurones(2 I-neurones and 2 B-neurones) were examined, Quipazine s i g n i f i c a n t l y lowered the spontaneous discharge of the I-neurones and attenuated the burst a c t i v i t y of B-neurones, The eff e c t of guipazine on a l l septal units was overcome by somatosensory stimulation. Effects Of 5hydroxy tryptophan On Hippocampal E l e c t r i c a l A c t i v i t y Some rats (n=5) received an intravenous i n j e c t i o n of the serotonin precursor 5-HTP (5-80 mg/kg) which, unlike 5-HT, passes the blood-brain barrier (Doner and Longo, 1962). Doses lower than 20 mg/kg did not result in any detectable change i n the spontaneously occurring or evoked patterns of hippocampal e l e c t r i c a l e l e c t r i c a l a c t i v i t y . Higher doses (40-80 mg/kg) produced a gradual desynchronization which occurred 5-7 min followinq the sta r t of the i n j e c t i o n . At 15-20 minutes post i n j e c t i o n the amplitude of the e l e c t r i c a l a c t i v i t y was greatly reduced (Fig. 5-8E). Recovery to the normal amplitude 146 I i Jff§£is 2 f 5-HT On Hippocampal Elecjtrieal-Actiyity.. A: spontaneously occurring a c t i v i t y recarded from the dentate gyrus before in j e c t i o n . BACJL records obtained 2-5 minutes after the in j e c t i o n of 5-HTP (60 mg/Kg). P i low voltage a c t i v i t y recorded 40 minutes following the i n j e c t i o n and persisting u n t i l the animal's death at 55 minutes post-i n j e c t i o n . CONTROL 5 HTP +2 min. +5 min. +40min. 500 uV 148 during the remainder of the recording session was not observed. Three of the four animals that received the higher doses ( i . e . >40 mg/kg) of 5-HTP died within 30-180 minutes post i n j e c t i o n . 4j_4 Discussion These data suggest that e l e c t r i c a l stimulation of the MR results i n i n h i b i t i o n of i r r e g u l a r l y f i r i n g septal neurones (I-neurones), disruption of rhythmical a c t i v i t y of bursting septal neurones (B-neurones) and desynchronization of e l e c t r i c a l a c t i v i t y of the hippocampus. Effect Of MR Stimulation On • I-neurones. -, Single pulse stimulation resulted i n short latency (mean=12. 5 msec) i n h i b i t i o n of I-neurones and considering the distance between stimulating and recording s i t e s (8-10 mm) allows for a conduction velocity of between 0.5 and 1.5 m/s. These v e l o c i t i e s are compatible with previously reported conduction v e l o c i t i e s of unmyelinated 5-HT f i b r e s (Wang and ighajanian, 1977) . The r e l a t i v e constancy of the onset of i n h i b i t i o n together with anatomical evidence f o r a dir e c t projection from the MB to the medial septum (Conrad, Leonard and P f a f f , 1974; Halaris et a l . , 1976) suggests that t h i s response i s mediated by a monosynaptic input d i r e c t l y onto I-neurones. The 149 absence of clear i n h i b i t o r y responses following stimulation of l o c i outside the MR indicates that the observed i n h i b i t i o n i s not mediated by an extra-raphe system. Effect Of MR Stimulation On B-neurones-In contrast to the response of I-neurones, B-neurones were unaffected by single pulse stimulation of the MR regardless of whether they were in bursting or non-bursting mode of discharge (See Fig. 4-3). However, the observation that r e p e t i t i v e stimulation disrupts the bursting of B-neurones without s i g n i f i c a n t l y a l t e r i n g their f i r i n g rate indicates a more complex relationship between the MB and those neurones than that suggested for I-neurones. Repetitive s t i m u l i may temporally summate to i n h i b i t l o c a l c i r c u i t s , involving septal interneurones (Tombol and Petsche, 1968; Mclennan and M i l l e r , 1974a) or neurones presently c l a s s i f i e d as I-neurones. I t i s interesting that seme septal neurones that are monosynaptically activated by stimulation of RSA generating zones i n the hippocampus are I-neurones which are i n h i b i t e d by MR stimulation. It i s conceivable that a proportion of I-neurones are septal interneurones that are excited by axon c o l l a t e r a l s of B-neurones which project to the hippocampus. An alternate explanation f o r the effectiveness of repetitve stimulation i s that mechanisms within the MR i t s e l f may be i n i t i a t e d only 150 by r e p e t i t i v e stimulation. However, th i s i s unlikely since single pulse stimulation of the ME r e s u l t s i n i n h i b i t i o n of I-neurones and a l t e r s neuronal transmission in the dentate gyrus (See Chapter 8). Regulation Of Hiepocampal a c t i v i t y Chapter 3 has emphasized the importance of the medial septal region as a relay station transforming influences of the brainstem into a discontinuous pulse t r a i n which i n i t i a t e s hippocampal RSA. The results of the present chapter show that a system o r i g i n a t i n g i n the MS i s capable of disrupting the bursting response of septal neurones which i n turn re s u l t s i n a desynchronized hippocampal a c t i v i t y . Wilson et al (1975) demonstrated a s i m i l a r effect on septal neurones and hippocampal a c t i v i t y following stimulation of the l a t e r a l hypothalmus. Since projections from the ME to the medial septum are known to course through the medial forebrain bundle, i t i s possible that desynchronization observed by these authors was due to the activation of f i b r e s of passage of neurones or i g i n a t i n g i n the MB . Alternate mechanisms mediating a desynchronization of hippocampal a c t i v i t y have been proposed. Anchel and Lindsley (1972) observed that lesions of the fornix blocked synchronization but not the desynchronization e l i c i t e d from hypothalamic s i t e s thereby concluding 151 that the l a t t e r effect was mediated by an extra-septal system. McLennan and M i l l e r (1976) proposed a frequency qating mechanism in the l a t e r a l septum which controls the discharge pattern of medial septal neurones such that either bursting or i r r e g u l a r f i r i n g w i l l r e s u l t depending on the l e v e l of hippocampal output mediated by the fimbrial pathway. At the present time the mechanisms regulating the a c t i v i t y of septal neurones and patterns of hippocampal a c t i v i t y have not been s u f f i c i e n t l y elucidated to eliminate any of the above proposals., Possible Role Of Serotonin The present re s u l t s suggest that a serotonergic system mediates the responses e l i c i t e d by MR stimulation. This suggestion i s based on the observation that p-CPA pretreatment which lowers forebrain 5-HT blocks the i n h i b i t i o n of I-neurones e l i c i t e d by MR stimulation. In addition, r e p e t i t i v e stimulation of MR i n p-CPA treated rats does not disrupt the bursting of B-neurones or desynchronize hippocampal a c t i v i t y . The additional observations that guipazine, a presumed agonist of serotonin (Rodriques et a l , 1973) desynchronizes hippocampal a c t i v i t y strengthens the proposal that a serotonergic system mediates the observed desynchronization. , The observation that the e f f e c t s of guipazine are 152 r e v e r s i b l e inasmuch as RSA can be subsequently produced by eserine i n j e c t i o n or somatosensory stimulation suggests that desynchronization i s not due to nonspecific and permanent destruction of BSA generators. Likewise the observation that p-CPA does not abolish BSA indicates an i n t a c t septal-hippocampal axis. The proposal that serotonin i s not es s e n t i a l for the generation of BSA i s also supported by the f a i l u r e to block BSA i n freely moving animals following the systemic i n j e c t i o n of the 5-HT antagonist methysergide (Robinson, 1978). The r e s u l t s following the intravenous i n j e c t i o n of 5-HTP were not as clea r cut as the desynchronization produced by guipazine i n j e c t i o n . Although desynchronization of hippocampal e l e c t r i c a l a c t i v i t y was observed following the i n j e c t i o n of 5-HTP, very large doses were reguired to produce r e l i a b l e e f f e c t s . Similar observations were previously made by Domer and Longo (1962) in the rabbit. Thus, i t i s possible that these doses reguired to e l i c i t desynchronization produced systemic effects which ultimately k i l l the animals. Previous studies have also implicated a serotonergic system i n the control of septal unit a c t i v i t y . Stefanis (1964) reported that iontophoretic application of 5-HT i n h i b i t s the discharge of septal neurones. More recently, Segal (1976) observed that 153 raphe stimulation r e s u l t s in a long latency (20-50 msec) i n h i b i t i o n of septal neurones. In the present experiments no i n h i b i t i o n with latencies longer than 25 msec was observed. Although i t i s d i f f i c u l t to account for t h i s discrepancy between the latencies to i n h i b i t i o n , the fact that i n both instances p-CPA pretreatment blocked the i n h i b i t i o n suggests that these ef f e c t s were mediated by serotonin. The observation that s t i m u l i of low i n t e n s i t y i n h i b i t e d I-neurones only i f the stimulating electrode was in the centre of the ME provides further support for the suggestion that the observed i n h i b i t i o n i s mediated by a serotonergic system originating i n the MB . Furthermore, the preliminary observation that systemic i n j e c t i o n of guipazine decreases the f i r i n g rate of I-neurones and attenuates the bursting of B-neurones also implicate 5-HT i n the control of septal unit a c t i v i t y . However, these effects can be more d i r e c t l y evaluated by the iontophoretic application of guipazine and 5-HT i t s e l f on i d e n t i f i e d septal neurones. 154 4.5 Summary A model o f the proposed r e l a t i o n s h i p between ME, s e p t a l a r e a and t h e hippocampus based on the r e s u l t s of t h i s and the p r e v i o u s c h a p t e r i s shown i n F i g . 4-8, S i n g l e p u l s e s t i m u l a t i o n o f MR causes d i r e c t i n h i b i t i o n of non b u r s t i n g I-neurones w i t h o u t i n h i b i t i n g B-neurones s u g g e s t i n g t h a t o n l y I-neurones r e c e i v e a monosynaptic i n p u t from MR. The d e s y n c h r o n i z a t i o n of the b u r s t i n g of B-neurones and RSA may be mediated by t e m p o r a l summation o f i n h i b i t e d I-neurones. S i n c e B-neurones a r e a n t i d r o m i c a l l y a c t i v a t e d by s t i m u l a t i o n of RSA g e n e r a t i n g zones they p r o j e c t t o the hippocampal f o r m a t i o n . The subseguent i n h i b i t i o n o f B-neurones s u g g e s t s t h a t t h e y g i v e o f f axon c o l l a t e r a l s t o n e i g h b o u r i n g i n h i b i t o r y i n t e r n e u r o n e s . The o b s e r v a t i o n t h a t some I-neurones which a re i n h i b i t e d by MR are s y n a p t i c a l l y a c t i v a t e d by hippocampal s t i m u l a t i o n s u g g e s t s t h a t a p o p u l a t i o n of I-neurones may i n f a c t be th e i n h i b i t o r y i n t e r n e u r o n e s . Recent e v i d e n c e t h a t antagonism o f the i n h i b i t o r y t r a n s m i t t e r gamma-a m i n o b u t y r i c a c i d d i s s r u p t s t h e b u r s t i n g of MS neurones (McLennan and M i l l e r , 1974a) emphasizes t h e im p o r t a n c e o f i n h i b i t o r y mechanisms i n t h e g e n e r a t i o n of r h y t h m i c i t y . M a n i p u l a t i o n s t h a t i n i t i a t e RSA i n c l u d i n g somatosensory or f i m b r i a l s t i m u l a t i o n (McLennan and M i l l e r , 1974a) may p r o v i d e a net e x c i t a t o r y d r i v e onto I-neurones. 155 F I G 4 - 8j_ A Schematic I l l u s t r a tign Of The Proposed Synaptic Arrangements Between The MR Segtumj, And Hippocampal Formation MB neurones d i r e c t l y i n h i b i t septal I-neurones which i n turn may i n h i b i t B-neurones, Stimulation of the dentate gyrus antidromically activates B-neurones and i n h i b i t s t h e i r spontaneous discharge v i a recurrent c o l l a t e r a l s innervating interneurones i n the septum. Black c i r c l e s represent i n h i b i t i o n , white c i r c l e s excitation. • — MR I B- neurone I-neurone Septum Hippocampus 157 CHAPTER 5_l THE DOE SAL NC^ADBENERGI C SYSTEM AND HIPPOCAMPAL ELECTRICAL- ACTIVITY 5.S.I Introduction Previous studies (reviewed i n chapter 1) have shown that stimulation of diffuse brainstem regions e l i c i t s rhythmical e l e c t r i c a l a c t i v i t y in the hippocampus. Further characterization of these ascending pathways and the l o c a l i z a t i o n of these c e l l s of o r i g i n i s e s s e n t i a l for understanding the mechanisms which underlie hippocampal and c o r t i c a l activation. The observation that e l e c t r i c a l stimulation i n the v i c i n i t y of the locus coeruleus, the s i t e of o r i g i n of noradrenaline containing neurones which innervate the forebrain , e l i c i t s RSA (Macadar et a l , 19 74) suggests that t h i s pharmacologically d i s t i n c t system may i n i t i a t e the generation of rhythmical a c t i v i t y i n the septum and hippocampus. This suggestion i s supported, i n part, by the observations that amphetamine in j e c t i o n s r e l i a b l y e l i c i t RSA (Vanderwolf, 1975) and that e l e c t r o l y t i c lesions of the locus coeruleus r e s u l t i n a decrease in e l e c t r o c o r t i c a l arousal (Jones et a l , 1973) . However, a detailed analysis of the role of t h i s pathway in the control of rhythmical hippocampal a c t i v i t y has not yet been car r i e d out., The aim of the present chapter i s to 1) test the e f f e c t s of e l e c t r i c a l stimulation of LC on hippocampal 1 5 8 e l e c t r i c a l a c t i v i t y , 2) determine whether these e f f e c t s are dependent on the dorsal noradrenergic pathway and 3) determine whether t h i s pathway mediates the well known effects of amphetamine on hippocampal e l e c t r i c a l a c t i v i t y . 5_j.2 Experimental Procedures E l e c t r i c a l stimulation: Bipolar stimulating electrodes were s t e r e o t a x i c a l l y positioned i n the v i c i n i t y of the locus coeruleus (1.5 mm posterior to stereotaxic zero; 0.9-1.3 mm l a t e r a l ; 5.5-7.0 mm below c o r t i c a l surface) in urethane anaesthetized rats. Stainless s t e e l recording electrodes were lowered into the CA1 pyramidal c e l l layer or the upper blade of the dentate gyrus to a position which r e l i a b l y produced spontaneous or sensory evoked RSA, The LC was then stimulated with a 3-10 second train of pulses (0,1 msec duration, 1-5 V) at a frequency of 10-300 Hz and the e f f e c t s on hippocampal e l e c t r i c a l a c t i v i t y were determined. The positions of the stimulating and recording electrodes were marked by producing a small lesion for subsequent h i s t o l o g i c a l v e r i f i c a t i o n . Lesion experiments: In order to examine i f the observed responses were mediated by the ascending NA projection, 9 rats received a b i l a t e r a l i n j e c t i o n of 6-hydroxydopamine(6-OHDA, 6 ug) d i r e c t l y into the dorsal NA bundle (AP=+2.6; L=+1.1; V=-4.5). In addition the LC 159 of 5 r a t s was e l e c t r o l y t i c a l l y l e s i o n e d b i l a t e r a l l y u s i n g 1.0-1.5mA a n o d a l c u r r e n t f o r 10-20 seconds. At l e a s t 5 days were a l l o w e d f o r r e c o v e r y a f t e r which the r a t s were a n a e s t h e t i z e d w i t h u r ethane f o r r e c o r d i n g e x p e r i m e n t s . The o c c u r r e n c e o f RSA i n c o n t r o l and l e s i o n e d r a t s was no t e d d u r i n g spontaneous c o n d i t i o n s and f o l l o w i n g p e r i p h e r a l s e n s o r y s t i m u l a t i o n , i n t r a v e n o u s i n j e c t i o n o f amphetamine (0.5*2.0 mg/kg), e s e r i n e (0.08-0.1 mg/kg) and a t r o p i n e s u l p h a t e (1-5 mg/kg). 5_s.3 R e s u l t s SRO&taneous And Sensory. Evoked P a t t e r n s Of E l e c t r i c a l A c t i v i t y . . ; The spontaneous e l e c t r i c a l a c t i v i t y of the hippocampus i n r a t s w i t h e l e c t r o l y t i c l e s i o n o f t h e LC or 6-OHDA l e s i o n o f t h e d o r s a l NA bundlewas not d i f f e r e n t from t h a t r e c o r d e d i n c o n t r o l r a t s . A m i x t u r e o f f a s t and slow waves w i t h t h e p e r i o d i c appearance of RSA was r e c o r d e d i n a l l t h r e e groups ( F i g . 5-1). The freg u e n c y o f RSA f o l l o w i n g s ensory s t i m u l a t i o n i n c o n t r o l (mean=5.6±0.2) r a t s d i d not d i f f e r from t h a t r e c o r d e d i n e l e c t r o l y t i c (mean=5.3±0.4) or 6-OHDA t r e a t e d (mean=5. 3 + 0. 2, ) r a t s ( F i g . 5-2). I n a d d i t i o n t h e r e were no d e t e c t a b l e d i f f e r e n c e s i n t h e a m p l i t u d e or shape of s p o n t a n e o u s l y o c c u r r i n g and sensory-evoked RSA. 160 5- I i Rhythmical E l e c t r i c a l A c t i v i t y Recorded In The Dentate Gyrus Of Control An£Treated Bats A RSA was e l i c i t e d i n each animal by a t a i l pinch during the period indicated by the s o l i d l i n e . NORMAL 6-OHDA LC LESION 1 sec. 16 2 FIGJS. 5- 2 i Frecjuency Of S p o n t a n e o u s l y O c c u r r i n g And-E l i c i t e d RSA A-Each h i s t o g r a m r e p r e s e n t s the mean fr e g u e n c y o f RSA r e c o r d e d i n normal (n=7), 6-OHDA-treated (n=9) and L O l e s i o n e d (n = 5) groups. The v e r t i c a l bars a r e s t a n d a r d e r r o r s of t h e mean. There a r e no s t a t i s t i c a l l y s i g n i f i c a n t d i f f e r e n c e s between e x p e r i m e n t a l groups. However, RSA r e c o r d e d d u r i n g t a i l p i n c h (T.P.) o r f o l l o w i n g e s e r i n e was f a s t e r than s p o n t a n e o u s l y o c c u r r i n g RSA. 164 I n o r d e r t o v e r i f y t h a t r h y t h m i c a l e l e c t r i c a l a c t i v i t y i s g e n e r a t e d i n each group of r a t s the fr e g u e n c y of RSA was examined (2-60 min.), f o l l o w i n g t h e i n t r a v e n o u s i n j e c t i o n o f e s e r i n e (0.08-0.1 mg/kg). T h i s t r e a t m e n t r e s u l t e d i n t h e c o n t i n u o u s appearance of RSA i n a l l t h r e e groups o f r a t s f o r p e r i o d s r a n g i n g between 20 min and 2 h r s . As i n t h e case o f spontaneous and sensory-evoked RSA, t h e r e were no s i g n i f i c a n t d i f f e r e n c e s i n t h e fr e g u e n c y o f RSA between c o n t r o l and l e s i o n e d r a t s ( F i g . 5-2) . E f f e c t Of LC s t i m u l a t i o n On Hi£j20camj_al A c t i v i t y ^ In c o n t r o l r a t s e l e c t r i c a l s t i m u l a t i o n (1-6 V) i n the r e g i o n o f the LC r e l i a b l y e l i c i t e d RSA h a v i n g a fregue n c y of 4-7 Hz. As shown i n F i g . 5-3, t h e fr e g u e n c y o f e l i c i t e d RSA was r e l a t e d t o t h e i n t e n s i t y o f s t i m u l a t i o n . At lower s t i m u l a t i o n i n t e n s i t i e s (1-2 V) t h e evoked RSA was s t i m u l u s bound , but at h i g h e r i n t e n s i t i e s RSA p e r s i s t e d f o r up t o 30 seconds f o l l o w i n g t e r m i n a t i o n of the t r a i n . The s t i m u l u s f r e g u e n c y was s e l e c t e d by k e e p i n g t h e i n t e n s i t y and d u r a t i o n o f t h e t r a i n c o n s t a n t and a n a l y z i n g RSA a t a range of s t i m u l u s f r e q u e n c i e s (10-300 Hz). A s t i m u l u s f r e g u e n c y o f 100 Hz was found t o be o p t i m a l a t 3-4 V i n t e n s i t y and t h i s f r e q u e n c y was used i n t h e remainder of the a n a l y s i s . 165 fi^Ls. 5- J i - E f f e c t s Of Stimulation In Thg Region Of The. Locus Coeruleus On E l e c t r i c a l A c t i v i t y Of The Dentate Gyrus The top record shows RSA e l i c i t e d by a t a i l pinch (T.P) and the remaining records show the effects of stimulating LC with the voltages indicated on the l e f t during the period shown by the s o l i d bar. 167 As shown i n Fig. 5-4, e l e c t r i c a l stimulation of diffuse regions i n the v i c i n i t y of LC also i n i t i a t e d RSA. An analysis of the relationship between the h i s t o l o g i c a l l y v e r i f i e d s i t e of the stimulating electrode and the current reguired to i n i t i a t e BSA revealed several regions which were at l e a s t as ef f e c t i v e as LC proper. These regions include the mesencephalic nucleus of the f i f t h nerve (MBS V) and more ventral placements in nucleus pontis caudalis. Placements in the area of MES V also e l i c i t e d f a c i a l movements which could i n d i r e c t l y i n i t i a t e RSA through sensory-motor pathways (Vanderwolf, 1972). Effects Of 62OHDA On RSA I n i t i a t e d By LC Stimulation-In control rats the concentration of NA i n the hippocampal formationwas 0.41.t0.0 5 ug/kg. Following a b i l a t e r a l i n j e c t i o n of 6-OHDA into the dorsal bundle the concentration was reduced to 0.02±0.3ug/kg (94% depletion) which i s s i g n i f i c a n t l y lower (p<0.01) than normal. E l e c t r i c a l stimulation of LC i n 6-OHDA treated rats (Fig, 5 - 5 ) e l i c i t e d RSA of comparable freguency and amplitudes to that recorded i n control rats. As shown in Fig. 5 - 6 , the stimulus-response curve of 6-OHDA rats i s not s i g n i f i c a n t l y d i f f e r e n t from the normal curve, despite less than 10% NA remaining i n the hippocampal formation. 16.8S FIG._ 5- L o c a t i o n Of S t i jmulat ing E l e c t r o d e s In The Region Of LC And I n t e n s i t y T h r e s h o l d For The Genera t i on Of RSA A i Drawings of c o r o n a l s e c t i o n s showing t h e e f f e c t i v e n e s s of s t i m u l a t i n g at each s i t e as i n d i c a t e d by the key on the lower l e f t . B i Corona l s e c t i o n o f the pons at about P 2.0. The arrow i n d i c a t e s the t i p o f the s t i m u l a t i n g e l e c t r o d e on the pe r imete r of LC. A b b r e v i a t i o n s Cer Cerebe l lum DTN Dorsa l Tegmental Nucleus NV mesencephal ic nuc leus of the f i f t h nerve LC Locus Coeru leus RPOC R e t i c u l a r i s P o n t i s O r a l i s C a u d a l i s IV Fourth C e r e b r a l V e n t r i c l e 169 170 Z1Q.3. 5- 5: E l e c t r i c a l A c t i v i t y Recorded In The Dentate Gyrus Of A 6-OHD_A Treated Rat Rhythmical a c t i v i t y was evoked by t a i l pinch (T.P.) and stimulation in the reqion of LC with the indicated voltaqes (B and C). D shows continous RSA recorded 5 minutes following eserine i n j e c t i o n . 172 Effects Of Amphetamine In control rats the intravenous i n j e c t i o n of D-amphetamine (2-5 mg/kg) evoked RSA in a dose-dependent manner. This response (Fig, 5-7) had an onset latency of 2-5 minutes and persisted for 2-4 hrs. The frequency of amphetamine induced RSA ranged between 5-7 Hz and was remarkably constant for each animal. The i n j e c t i o n did not r e s u l t in any overt movement, although the rate of v e n t i l a t i o n often increased during the f i r s t 5 min post i n j e c t i o n . Amphetamine also e l i c i t e d RSA i n rats which had an e l e c t r o l y t i c lesion of the LC or a 6-OHDA i n j e c t i o n into the dorsal bundle. The latency to onset and the freguency of BSA produced by amphetamine did not d i f f e r s i g n i f i c a n t l y between groups. Effects Of Atropine On Spontaneous And - Evoked- Act i v i t y • It has been suggested that under some conditions RSA i s blocked by atropine whereas in cases such as those related to concurrent movement RSA i s resis t a n t to blockage by atropine (Vanderwolf, 1975). In the present series of experiments, using urethane anaesthetized r a t s , RSA was always blocked by 2,0—5.0 mg/kg atropine independent of whether i t was spontaneously occurring, e l i c i t e d by a t a i l pinch or i n i t i a t e d by stimulation i n the region of LC. In addition, atropine pretreatment 2-5 minutes p r i o r to 173 F I G ^ 5- 6.1 Freguency Of RSA As A F u n c t i o n Of LC S t i m u l u s I n t e n s i t y In C o n t r o l And 6-OHDA Les i o n e d Rats The v e r t i c a l e r r o r s of t h e mean. ba r s r e p r e s e n t s t a n d a r d 174 I Stimulus Intensity (Volts) 175 fJG-A 5- 7t I f f gets Of Amphetamine On E l e c t r i c a l Activity Of The Dentate Gyrus The top sweep of each record shows the ongoing e l e c t r i c a l a c t i v i t y 5.0 minutes following a saline vehicle i n j e c t i o n and the lower sweep shows the a c t i v i t y 5.0 minutes following the intravenous i n j e c t i o n of amphetamine (2.0 mg/Kg). Note that amphetamine i n i t i a t e d rhythmical e l e c t r i c a l a c t i v i t y in a l l three cases. CONTROL Saline Amph. 6-OHDA Sali me Amph. ELECTROLYTIC Saline Amph. US sec. 177 amphetamine i n j e c t i o n was found t o c o m p l e t e l y prevent RSA f o r p e r i o d s of up t o 2,0 h r s , 5jJ5 D i s c u s s i o n Does The D o r s a l NA Bundle U n d e r l y RSA ? The r e s u l t s o f t h e p r e s e n t c h a p t e r demonstrate t h a t t h e d o r s a l n o r a d r e n e r g i c bundle o r g i n a t i n g i n the l o c u s c o e r u l e u s does not p l a y an e s s e n t i a l r o l e i n the g e n e r a t i o n o f RSA r e c o r d e d i n ure t h a n e a n a e s t h e t i z e d r a t s . I f t h e d o r s a l n o r a d r e n e r g i c bundle were c r i t i c a l f o r t h e g e n e r a t i o n o f r h y t h m i c a l a c t i v i t y one would expect t h a t 1) d e s t r o y i n g t h i s p r o j e c t i o n would a b o l i s h e r h y t h m i c a l a c t i v i t y and 2) d i r e c t e l e c t r i c a l s t i m u l a t i o n of the c e l l b o d i e s o f o r i g i n would i n i t i a t e r h y t h m i c a l a c t i v i t y a t t h r e s h o l d i n t e n s i t i e s l o w e r than t h o s e r e q u i r e d at a d j a c e n t s i t e s . However, 6-OHDA l e s i o n s o f the d o r s a l bundle which r e s u l t e d i n 90-95% d e p l e t i o n o f NA i n t h e hippocampus d i d not a l t e r the spontaneous o r sensory-evoked p a t t e r n s of hippocampal e l e c t r i c a l a c t i v i t y . I n a d d i t i o n 6-OHDA l e s i o n s d i d not i n c r e a s e t he s t i m u l u s i n t e n s i t y r e q u i r e d t o i n i t i a t e BSA from e l e c t r o d e placements l o c a l i z e d t o LC proper r e l a t i v e t o placements s i t u a t e d i n a d j a c e n t are a s . These r e s u l t s s u p p o r t t h e s i m u l t a n e o u s f i n d i n q s of Robinson e t a l (1977) which show t h a t t h e d o r s a l NA bundle i s not e s s e n t i a l f o r t h e q e n e r a t i o n of RSA i n 178 f r e e l y moving r a t s . These r e s e a r c h e r s d i d not f i n d any low t h r e s h o l d s i t e s w i t h i n t h e LC f o r the i n i t i a t i o n of RSA and were a l s o unable t o b l o c k RSA f o l l o w i n g d e p l e t i o n o f hippocampal NA. The r e s u l t s of the p r e s e n t c h a p t e r and t h o s e of Robinson et a l (1977) do not e l i m i n a t e t h e p o s s i b i l i t y t h a t NA c o n t a i n i n g axons o t h e r t h a n t h o s e c o u r s i n g through t h e d o r s a l bundle p l a y a r o l e i n t h e g e n e r a t i o n of RSA. I t i s p o s s i b l e t h a t NA c o n t a i n i n g c e l l groups c a u d a l t o LC ( i . e . A2 and A4 a c c o r d i n g t o D a hlstrom and Fuxe, 1964) may i n f l u e n c e e x t r a - h i p p o c a m p a l r e g i o n s such as t h e s e p t a l a r e a which are a s s o c i a t e d w i t h t h e g e n e r a t i o n o f RSA. The r e c e n t o b s e r v a t i o n o f Moore (1978) t h a t 6-OHDA l e s i o n s of d o r s a l NA bundle t o t a l l y d e p l e t e NA i n t h e hippocampus but o n l y p a r t i a l l y (48%) l o w e r s e p t a l NA c o n t e n t suggest t h a t c a u d a l groups may i n d i r e c t l y i n f l u e n c e RSA. The f a i l u r e t o a b o l i s h RSA by making s p e c i f i c l e s i o n s t a k e n t o g e t h e r w i t h t h e o b s e r v a t i o n t h a t d i f f u s e a r e a s i n b r a i n s t e m i n i t i a t e RSA s u g g e s t s t h a t m u l t i p l e a s c e n d i n g systems modulate RSA. The s e p t a l a r e a may be the s i t e of i n t e r a c t i o n s between th e s e v a r i o u s systems and may t h u s be t h e o n l y c r i t i c a l r e g i o n f o r the g e n e r a t i o n of RSA. 179 4£tipn Of Ampheta mine The intravenous i n j e c t i o n of amphetamine was shown to produce RSA i n rats which had e l e c t r o l y t i c destruction of LC, 6-OHDA lesion of the dorsal bundle and control rats. The mechanisms mediating t h i s action of amphetamine are not known and may be related to enhanced synaptic release of the catecholamines dopamine and noradrenaline (reviewed by Moore, 1977). The effect of amphetamine i s unlikely to be mediated by enhanced NA release from the terminals of LC axons since RSA was demonstrated following e l e c t r o l y t i c lesions of LC. Amphetamine could enhance the a c t i v i t y of the mesolimbic DA system which has been shown to innervate the l a t e r a l septal area (L i n d v a l l , 1975; Assaf and M i l l e r , 1977). The neurones of the l a t e r a l septum have been implicated in the generation of rhythmical burst discharge of medial septal neurones (McLennan and M i l l e r , 1974a, 1976). It i s not known to what extent the actions of amphetamine are dependent on septal-hippocampal cholinergic neurones. The preliminary observations that in urethane anaesthetized rats the action of amphetamine can be blocked by atropine emphasize the role of cholinergic systems. There i s also the p o s s i b i l i t y that amphetamine injec t i o n s enhance the release of acetycholine ( Moore, 197 7). However, there i s no direct evidence that Ach i n the septal area 180 i s enhanced by amphetamine. P o s s i b l e C o n t r i b u t i o n Qf Orethane A n a e s t h e s i a The r e s u l t s of t h e p r e s e n t c h a p t e r were o b t a i n e d i n u rethane a n a e s t h e t i z e d r a t s and t h e p o s s i b i l i t y a r i s e s t h a t t h e s e c o n d i t i o n s mask the p o t e n t i a l i n f l u e n c e of LC on RSA. As p o i n t e d out i n c h a p t e r 3, u r e t h a n e narrows t h e range o f RSA from 3-12 Hz i n f r e e l y moving r a t s down to 3-7 Hz and any c o n t r i b u t i o n of LC i n d r i v i n g f r e q u e n c i e s above 7.0 Hz w i l l not be seen. The r e s u l t s o f Gray, HcNaughton, James and K e l l y (1975) do i n f a c t suggest t h a t NA l o w e r s the t h r e s h o l d f o r ' s e p t a l d r i v i n g * o f RSA at an o p t i m a l f r e q u e n c y of 7.7 Hz. However, s e p t a l d r i v i n g has not y e t been r e l a t e d t o p h y s i o l o g i c a l o c c u r r e n c e of RSA and t h e r e s u l t s o f Robinson e t a l (1977) i n f r e e l y moving r a t s s u g g e s t t h a t t h e f u l l f r e q u e n c y range of RSA ( i . e . 3-12 Hz) o c c u r s i n t h e absence o f hippocampal NA. However the e v i d e n c e t h a t t h e f u l l freguency band o c c u r s i n the absence of LC n o r a d r e n e r g i c system does not e l i m i n a t e t h e p o s s i b i l i t y t h a t t h i s system modulates RSA by l o w e r i n g the t h r e s h o l d f o r c e r t a i n f r e g u e n c i e s . , 181 5._5 Summary The results of the present chapter eliminate the p o s s i b i l i t y that the LC noradrenergic system plays plays an e s s e n t i a l role in generation of BSA. It i s proposed that amphetamine e l i c i t s BSA by mechanisms other than enhanced NA release in the hippocampus. 182 CHAPTER 6: CHARACTERIZATION OF THE PER FOR ANT PATH-DENTATE PSOJECTION 6j_1_ Introduction As reviewed i n chapter 1, h i s t o l o g i c a l studies of the hippocampal formation reveal two major c e l l types, the pyramidal c e l l s of the' hippocampus proper and the granule c e l l s (G-cells) of the dentate gyrus. The G-c e l l s form a curved sheet of c e l l bodies with a p i c a l l y oriented dendrites reaching the obliterated hippocampal f i s s u r e (chapter 1, F i g . 1). In addition to t h i s clear segregation of dendrites and c e l l bodies, there i s a highly organized laminar d i s t r i b u t i o n of the termination of the major afferent f i b r e s . As shown i n Fig. 6-1, afferents from the i p s i l a t e r a l entorhinal cortex terminate in the cuter molecular layer onto d i s t a l dendrites wheras commissural and associational afferents terminate more proximal to the c e l l bodies in the inner molecular layer. While the precise organization of other e x t r i n s i c afferents i s not yet known, histochemical studies suggest that monoamine-containing systems terminate i n the v i c i n i t y of the G-c e l l bodies and the dentate hilus (reviewed in section 1. 1. 1) The most massive of the above inputs i s that from the entorhinal cortex which projects via the perforant path (PP) to the outer two-thirds of the d e n d r i t i c 183 ll L arain_a.E Organization Of Afferents - To The-Dentatgi, Afferents from the l a t e r a l and medial regions of the entorhinal cortex project via the perforant path onto the outer two-thirds of of the dentate molecular layer which contains the dendrites of G-cells, Commissural and associational (not shown here) fibres from the cont r a l a t e r a l and i p s i l a t e r a l CA3, respectively, terminate i n the inner molecular layer. Cholinergic and monoamine-containing fibers project to more dif f u s e regions i n the v i c i n i t y of the c e l l body layer and dentate h i l u s . > Entorhinal (PP) Commissural Septal (Ach) • Raphe (5 -HT) L C (NA) 185 t r e e . A c c o r d i n g t o N a f s t a d ( 1967), t h e s m a l l d i a m e t e r axons of the PP make en-passage s y n a p t i c c o n t a c t w i t h many s p i n e s o f t h e p r o f u s e l y b r a n c h i n g d e n d r i t e s of G-c e l l s . E l e c t r o p h y s i o l o g i c a l a n a l y s i s o f t h i s s t r i c t l y l a m i n a t e d system r e v e a l e d t h a t s t i m u l a t i o n of PP r e s u l t s i n e x t r a c e l l u l a r l y r e c o r d e d f i e l d p o t e n t i a l s h a v i n g a t o p o g r a p h i c a l o r g a n i z a t i o n ( G l o o r e t a l , 1964; Andersen and Lomo, 1966; Lomo, 1971a; McNaughton and B a r n e s , 1977) E l e c t r o d e s s i t u a t e d i n t h e o u t e r m o l e c u l a r l a y e r of t h e d e n t a t e r e c o r d a n e g a t i v e e x t r a c e l l u l a r f i e l d which r e f l e c t s t h e s y n a p t i c c u r r e n t generated by d e n d r i t e s and i s termed the p o p u l a t i o n EPSP (Lomo, 1971a) . At t h e c e l l body l a y e r t h e evoked response c o n s i s t s o f a p o s i t i v e wave w i t h a superimposed n e g a t i v e s p i k e r e p r e s e n t i n g the synchronous f i r i n g o f n e u r o n a l a c t i o n p o t e n t i a l s and i s t h e r e f o r e r e f e r r e d t o as the p o p u l a t i o n s p i k e (Lomo, 1971a), An i m p o r t a n t f e a t u r e o f t h e above p r o j e c t i o n system i s t h e p l a s t i c i t y o f s y n a p t i c t r a n s m i s s i o n which i s observed when t h e PP i s s t i m u l a t e d a t p a r t i c u l a r f r e g u e n c i e s . R e l a t i v e l y b r i e f t e t a n i c s t i m u l a t i o n r e s u l t s i n an enhancement of both the p o p u l a t i o n EPSP and s p i k e r e s p o n s e which p e r s i s t s f o r hours and on o c c a s s i o n s f o r s e v e r a l days ( B l i s s and Lomo, 1973; B l i s s and Gardner-Medwin, 1973; Douglas and Goddard, 1975). In c o n t r a s t , a s i n g l e c o n d i t i o n i n g p u l s e t o the 186 PP r e s u l t s only i n short term (less then 1 second) potentiation of the response to a subsequent test pulse (Lomo, 1971b; Steward et a l , 1976). The f i r s t mentioned paradigm, commonly referred to as post-tetanic potentiation (PTP) as well as the l a t t e r , known as paired pulse f a c i l i t a t i o n , have also been confirmed i n i n v i t r p hippocampal s l i c e s (Dudek et a l , 1976; M i l l e r , unpublished observations), The mechanisms which mediate the above forms of functional p l a s t i c i t y are not known. Andersen (1975) suggests that they may r e f l e c t the a b i l i t y of the presynaptic terminal to release more transmitter. However, the p o s s i b i l i t y of an altered responsiveness of the postsynaptic elements i s just as l i k e l y (Dunwiddie et a l , 1978). In order to evaluate these p o s s i b i l i t i e s , t h i s part of the thesis examines the role of e x t r i n s i c afferents to the dentate in modifying the response of G-cells to stimulation of the perforant path. Such analysis i s e s s e n t i a l for determining whether changes in the e f f i c a c y of the c o r t i c a l input via the PP occur as a r e s u l t of the a c t i v i t y of afferents to the postsynaptic c e l l i n addition to those postulated to occur i n the presynaptic elements of the test pathway. The purpose of the present chapter i s to characterize the entorhinal-dentate projection i n the urethane anaesthetized rat and analyze paired pulse f a c i l i t a t i o n of the PP-evoked responses. The following chapters examine the e f f e c t s of stimulating e x t r i n s i c afferents 187 o r i g i n a t i n g i n t h e c o n t r a l a t e r a l hippocampus ( c h a p t e r 7 ) , median raphe n u c l e u s ( c h a p t e r 8) and t h e l o c u s c o e r u l e u s ( c h a p t e r 9) on t h e s e response, 6K2 E x p e r i m e n t a l Procedures E x t r a c e l l u l a r r e c o r d i n g e x p e r i m e n t s were performed on a t o t a l o f 94 urethane a n a e s t h e t i z e d r a t s . C o n c e n t r i c b i p o l a r s t i m u l a t i n g e l e c t r o d e s were p o s i t i o n e d i n the a n g u l a r bundle (8.1 mm p o s t e r i o r t o bregma; 4.2-4.4 mm l a t e r a l ; 3.0-3.5 below c o r t i c a l s u r f a c e ) t o a c t i v a t e t h e p e r f o r a n t path a s i t l e a v e s t h e i p s i l a t e r a l e n t o r h i n a l c o r t e x o r i n the s u b i c u l u m a d j a c e n t t o t h e hippocampal f i s s u r e where t h e PP e n t e r s t h e d e n t a t e g y r u s . E x t r a c e l l u l a r p o t e n t i a l s were r e c o r d e d from t h e d o r s a l hippocampus and t h e d e n t a t e g y r u s u s i n g e i t h e r g l a s s m i c r o p i p e t t e s f i l l e d w i t h 4M NaCl (2-5Mohm r e s i s t a n c e ) or sharpened t u n g s t e n m i c r o e l e c t r o d e s (9-12 Mohm) , I n some ca s e s more tha n one r e c o r d i n g e l e c t r o d e were used t o r e c o r d s i m u l t a n e o u s l y from both t h e s y n a p t i c r e g i o n i n the m o l e c u l a r l a y e r and from the c e l l body l a y e r , The s i g n a l s were a m p l i f i e d and a n a l y z e d a s d e s c r i b e d i n c h a p t e r 2, T y p i c a l l y t h e average of 20-30 s t i m u l u s p r e s e n t a t i o n s ( I S I 2.5-4.0 seconds) were p l o t t e d . The e f f e c t s o f c o n d i t i o n i n g s t i m u l a t i o n o f the PP on a subseguent t e s t v o l l e y , d e l i v e r e d through t h e same e l e c t r o d e was a n a l y z e d u s i n g t h e p a i r e d - p u l s e paradigm. T h i s c o n s i s t e d of comparing the a m p l i t u d e s and r a t e s o f 188 r i s e of the response evoked by the f i r s t {conditioning) pulse to that evoked by the second (test) pulse. The i n t e r v a l between conditioning and test pulses (C-T interval) ranged between 0-1500 msec, 6^3 Results Fig. 6-2 shows the e x t r a c e l l u l a r f i e l d potentials evoked by low i n t e n s i t y (1-2V) stimulation of the PP and recorded at the indicated depths in the dentate gyrus. The l o c a t i o n of the electrode at each recording s i t e can be approximated by the corresponding schematic of the upper and lower blades of the dentate. As shown by Lomo (1971a) in the rabbit and Gloor et a l (1964) i n the cat, a short latency negative wave i s recorded in middle molecular layer with a maximum at the l e v e l of the PP synapses (Nafstad, 1967). At the c e l l body layer the response reverses to a positive wave on which the discharges of i n d i v i d u a l G-cells are superimposed. The mirror images of these responses occur i n the lower blade of the dentate. As shown in Fig. 6-3, increasing the stimulus i n t e n s i t y enhances the amplitude of the dendritic negative f i e l d and s i g n i f i c a n t l y increases the number of spikes recorded on the positive c e l l layer f i e l d . At higher stimulus i n t e n s i t i e s (3-7 V) the units discharge synchronously giving r i s e to a large negative spike which for the reasons outlined below i s referred to as 189 Z 2.L Dejoth £rofiles Of The F i e l d P o t e n t i a l s Evoked I n The Dentate Gyrus By A Weak P e r f o r a n t Path V o l l e y Az_ E x t r a c e l l u l a r f i e l d p o t e n t i a l s r e c o r d e d a l o n g a v e r t i c a l t r a c k a t the i n d i c a t e d depths below t h e c o r t i c a l s u r f a c e . Ej. Depth p r o f i l e o f t h e evoked p o t e n t i a l s measured 1.0-1.5 msec a f t e r t h e i r onset ( i n d i c a t e d by v e r t i c a l b a r s i n A). Cj_ Schematic r e p r e s e n t a t i o n of t h e approximate p o s i t i o n s of the upper and lower b l a d e s of t h e d e n t a t e g y r u s . 191 IISL*. 6- 3z_ F i e l d Potentials Evoked In The Dentate. By. PP Stiraulation At Various Intensities Ai F i e l d potentials recorded at the indicated distances above the granule c e l l body layer following a weak PP volley. Bi Depth p r o f i l e as in A with a stronger PP volley. > £ 92 193 the population spike response of dentate granule cells.... I d e n t i f i c a t i o n Of Dentate Responses^ KJL * Molecular Iai§£ - As shown i n Fig, 6-4, a large negative response i s recorded by an electrode situated in the middle molecular layer of the dentate. This negative response (N2 in Fig. 6-4B) i s maximal 150-190 um from G-cell body layer in the region of the d i s t a l dendrites where the PP synapses terminate (Blackstad, 1958; Nafstad, 1967). The onset of t h i s negative response occurs at 2-4 msec following stimulation of the PP and reaches a peak at 4-6 msec. The latency to the onset of t h i s response and i t s r i s e time are d i r e c t l y related to stimulus intensity (Fig. 6-4B,C). The time from the onset of t h i s f i e l d to the e a r l i e s t discharge of G-cells ranged between 1.5-2,5 msec. The above negative response was evoked monosynaptically since i t followed moderate (up to 50 Hz) but not high freguency (100-200 Hz) stimulation of the perforant path (Fig.6-4D). In addition, i t s onset latency was r e l a t i v e l y consistent and thus unlikely to r e s u l t from polysynaptic a c t i v a t i o n (Fig. 6-4A) . A s i m i l a r negative response was previously recorded i n the dentate molecular layer of cats (Gloor et a l , 1964) and rabbits (Lomo, 1971a). Lomo (1971a) observed that the onset of the e x t r a c e l l u l a r negative wave coincided with the onset of the i n t r a c e l l u l a r 194 OSA 6_r Hi I d e n t i f i c a t i o n Of The F i e l d Potentials Recorded In The Dentate Molecular Lay,e.rA; hi 5 superimposed oscilloscope sweeps showing the small i n i t i a l d eflection (N1) and the prominent negative response (N2) . Bi e f f e c t s of increasing the stimulus i n t e n s i t y on the above responses. Cz_ expanded plots showing the s i z e of N1 and rate of r i s e of N2 at various stimulus i n t e n s i t i e s . P i i n a b i l i t y of the large negative f i e l d (N2) to follow high freguency stimulation suggesting that i t i s produced synaptically. 195 196 EI6.*. £- 5z_ Relationship Between PP Stimuljus In tensity And Amplitude Of Responses Recorded In The Dendritic Region Of The. Dent ate Each point represents the average oftwenty responses recorded i n the dendritic region of the dentate gyrus. The amplitude of the EPSP was measured while increasing (0) and decreasing (,) stimulus intensity to evaluate the v a r i a b i l i t y of the response. 0.0 1.0 2.0 3.0 4.0 Stimulus Intensity (Volts) 198 EPSP. Since t h i s negative monosynaptic wave i s maximal i n the layer of the perforant path synapses, i t w i l l be c a l l e d the e x t r a c e l l u l a r EPSP, This interpretation i s in accord with that of Gloor et a l (1964) and Lomo (1971a) . In some e x t r a c e l l u l a r records obtained i n the molecular layer a small negative deflection (N1 in Fig. 6-4C) precedes the onset of the EPSP by 0.4-0.6 msec. This small deflection i s only recorded i n a r e s t r i c t e d zone i n the v i c i n i t y of the PP synapse and i s l i n e a r l y r e l a t e d to stimulus intensity (Fig. 6-5) . Unlike the EPSP, t h i s early component i s not attenuated by high freguency stimulation (Fig. 6-4D) and i s actually enhanced at a short pulse i n t e r v a l s when the EPSP i s refractory. It i s , therefore, concluded that the early negative component i s the PP f i b r e volley and thus does not show c h a r a c t e r i s t i c s of the synaptic response described above. A s i m i l a r interpretation of th i s response was previously proposed by Lomo (1971a)., In addition, Lomo (1971a) observed that the presynaptic volley was not as res i s t a n t to anoxia as the synaptic response. , 199 ISSt. §.Z 6± I d e n t i f i c a t i o n Of The Evoked Potential S§£2ES§d At G-cell Body Lai§£i A: Components of the response recorded at G-c e l l body layer following stimulation of PP with a strong (10 V) volley. The plot i s an average of 20 t r i a l s and peaks are l a b e l l e d according to their p o l a r i t y and latency. Bj. Relationship between the evoked EPS P recorded i n the dendritic layer and the mirror image posi t i v e wave recorded at c e l l body layer. Cj_ Rate of r i s e of P1 response and dendritic EPSP as a function of stimulus i n t e n s i t y . 201 Bj_ C e l l Layer : Fig, 6-6& shows the components of the ex t r a c e l l u l a r f i e l d potential evoked by a strong perforant path volley (10V) and recorded at the c e l l body layer. These components are labelled according to th e i r p o l a r i t y and latency to onset. P1 i s recorded simultaneously with the r i s e of the negative dendritic EPSP and appears as i t s mirror image (Fig. 6-6B). Discharge of G-cells r a r e l y occurs at the start of P1 but rather 1.5-2.0 msec after i t s onset. The rate of r i s e of the P1 i s si m i l a r to that of the EPSP (Fig. 6-6D) and i s most l i k e l y the ex t r a c e l l u l a r r e f l e c t i o n of the dendritic EPSP recorded at the c e l l body layer. The observation that i t i s the opposite polarity to the EPSP and i s not caused by the discharge of G-cells suggests that i t acts as a source providing current to the dendritic EPSP sink (Lomo, 1 971 a) . Component N2 i s maximal i n the region of G - c e l l bodies and i s recorded only following stimulation of the perforant path with higher i n t e n s i t y . Only a positive wave i s recorded by the same electrode following low stimulus i n t e n s i t i e s (0.5-1.0 V) s u f f i c i e n t to evoke the dendritic EPSP. As the stimulating intensity i s gradually increased (1.0-2.0 V) single units are recorded on the po s i t i v e wave. The number of units increase with higher stimulus i n t e n s i t i e s (Fig. 6-7) u n t i l a prominent N2 component i s recorded. The latency to the peak of N2 corresponds to the mean latency to discharge of in d i v i d u a l G-cells 202 II£A,; 6 - 7z Development Of The Dentate jPjJfiulation Sjgikej, Responses recorded in the G - c e l l layer following PP stimulation with the indicated voltages. Note recruitment of single unit discharge to form a compound spike at higher stimulus i n t e n s i t y . 204 8z_ E f f e c t s Of Stimulus Intensity On Am£litude M S MtSliSx Of %k§ O-cell Layer response The peak latency and amplitude of the N2 component of the c e l l layer response i s plotted as a function of stimulus i n t e n s i t y . 2 0 5 " 5.0 Stim. Intensity (V) 206 (Andersen, Skrede and B l i s s , 1971) and i s i n v e r s e l y r e l a t e d t o s t i m u l u s i n t e n s i t y (Fig.,6-8).„ S i n c e the N2 component i s maximal i n the G - c e l l l a y e r ( F i g . 6-9) and i s r e l a t e d t o t h e d i s c h a r g e of G-c e l l s , i t most l i k e l y r e p r e s e n t s the monosynaptic a c t i v a t i o n o f a p o p u l a t i o n o f G - c e l l s ( G l o o r e t a l , 1964; Lomo, 1971a). F u r t h e r s upport of the s y n a p t i c n a t u r e o f N2 i s t h a t i t does not f o l l o w h i g h f r e g u e n c y (>100 Hz) s t i m u l a t i o n and shows l a t e n c y s h i f t s c h a r a c t e r i s t i c of monosynaptic responses ( F i g . 6-10). A d d i t i o n a l e v i d e n c e t h a t t h e N2 component i s caused by G - c e l l d i s c h a r g e was o b t a i n e d by a n t i d r o m i c a l l y a c t i v a t i n g t h e same u n i t s f o l l o w i n g s t i m u l a t i o n o f the mossy f i b r e (MF) pathway. As shown i n F i g . 6-11, low i n t e n s i t y s t i m u l a t i o n o f the MF pathway r e s u l t s i n a s harp n e g a t i v e s p i k e l o c a l i z e d t o t h e G - c e l l body l a y e r . U n l i k e the s y n a p t i c a l l y evoked N2 wave, t h i s s p i k e i s not i n f l u e n c e d by h i g h f r e g u e n c y s t i m u l a t i o n (>150 Hz) and does not show an a p p r e c i a b l e s h i f t i n l a t e n c y ( F i g . , 6-11D). I n a d d i t i o n the a m p l i t u d e of the s y n a p t i c N2 component c o u l d be enhanced by low f r e q u e n c y (5-7 Hz) s t i m u l a t i o n whereas the a n t i d r o m i c s p i k e c o u l d not be. The N2 component may t h u s r e p r e s e n t the monosynaptic a c t i v a t i o n o f a number of d e n t a t e G - c e l l s and i n t h e t e r m i n o l o g y of Lomo (1971a) w i l l be h e r e a f t e r r e f e r r e d t o as the p o p u l a t i o n s p i k e . 207 6- 9i The C e l l Layer Response Recorded At Various Depths In The Dentate^ Each point and corresponding potentials are the averages of 20 responses recorded along the same dorsal-ventral electrode track. Distance above G-Cell Layer O CO 209 FIG._ 6-1 Oj. Presynaptic And Postsynaptic Responses Following Stimulation Of The Perforant Path x~ Aj. e f f e c t of increasing stimulus freguency on PP evoked population spike response. B: f i b r e volley recorded from the PP (top sweep) and the G - c e l l layer response (lower sweep) following stimulation of the entorhinal cortex at 175 Hz. Note that, unlike the population spike, the PP volley i s not attenuated by high freguency stimulation. Ci 5 superimposed oscilloscope sweeps indicating v a r i a t i o n of the population spike latency c h a r a c t e r i s t i c of a monosynaptic response. p i histogram showing the d i s t r i b u t i o n of peak latencies following PP stimulation (ISI=2.5 seconds). The f i r s t v e r t i c a l bar indicates constancy of the stimulus a r t i f a c t . o o o X 1 1 5 "> 8 X M o Spikes CJl o J O - i 3 CD O 012 211 Component P2 i s recorded at the G-cell body layer at threshold i n t e n s i t i e s for tne production of a dendritic EPSP. Since P2 i s always recorded with the P1 component i f may also be interpreted as an image of the dendritic EPSP. However P2 i s not always symmetrical with P1 and may have neurones discharging anywhere along i t s length (Fig. 6-12). Since some of those evoked neurones which appear on the P2 component are not antidromically evoked but are synaptically activated by MF stimulation (Fig, 6-12), they are unlikely to represent G-cells. The neurones that discharge on the P2 component are frequently recorded i n the dentate h i l u s and may represent interneurones that were activated by G-cell axon c o l l a t e r a l s . In t h i s regard, Lomo (1 968) has suggested that P2 component s i g n i f i e s the s t a r t of IPSP's on granule c e l l s which are i n i t i a t e d by i n h i b i t o r y interneurones. On the basis of the above data i t may be suqqested that P2 may be in part the r e f l e c t i o n of dendritic EPSP and i n part the st a r t of i n h i b i t o r y mechanisms. The l a t e neqative component (N3) has not yet been i d e n t i f i e d . I t i s recorded i n the region of the c e l l body layer and i s maximal i n the region of the dentate hilu s at approximately 2.0 mm from the midline. Stimulating i n t e n s i t i e s below threshold for the production of a population spike produce a r e l i a b l e S3 component and as the stimulating i n t e n s i f y i s increased 212 F i g A 6 - J J i IntidromiG And Orthodromic Activation Of Dentate Granule C e l l s , Aj. depth p r o f i l e s i n d i c a t i n g a relationship between the PP population spike (left) and a similar negative response evoked by mossy f i b r e (MF) stimulation (right) . Bi the e f f e c t s of r e p e t i t i v e stimulation (100 Hz) on the MF (top trace) and PP (bottom trace) evoked responses recorded at the G - c e l l layer. Note that the amplitude of response evoked by MF stimulation i s not attenuated and thus may be antidromic. C: antidromic a c t i v a t i o n of G - c e l l following MF stimulation. The top trace indicates relationship between antidromically activated c e l l (obligue arrow) and f i e l d . Bottom trace indicates that both resonses followed 200 Hz stimulation of MF., DJJ. MF evoked response at fast sweep speed indicating the consistent latency and amplitude c h a r a c t e r i s t i c of antidromic responses CIS I = 1.0 sec), 2\3 214 the amplitude of N3 also increases. The amplitude of 133 was dependent on the position of the stimulating electrode but no stimulating positions were found that evoked a population spike without the N3 component., The evoked discharges of neurones were not usually superimposed on the N3 component, although in seme cases small spikes were recorded near the peak of this l a t e negative wave (Fig. 6-12). The latency to the onset of these spikes was 9-15 msec and was always longer than that for the monosynaptic population spike. However, i t i s d i f f i c u l t to assess whether these i n d i v i d u a l spikes were related to P2 or N3. The same neurones that are evoked near N3 are synaptically activated by mossy f i b r e stimulation (Fig. 6-12) suggesting that they are not G-cells. Furthermore the amplitude of N3 was s i g n i f i c a n t l y reduced by low freguency stimulation (<20 Hz) which potentiated the population spike response and often resulted i n the complete disappearance of N3 and the appearance of new negative spikes resembling the N2 component (Fig. 6-12D). The long latency of the N3 suggests that i t may have resulted from stimulating commissural projections coursing through the angular bundle to terminate i n the entorhinal cortex. However, a complete transection of the commissural projections i n 6 rats (see chapter 7) 10-20 days before the recording experiments did not 215 FIG t 6-12.1 I d e n t i f i c a t i o n Of The Late Negative Componen t 1N3L Recorded In The Denote Following Stimulation Of The Perforant Path. Ai relationship between the latency of the population spike (N2) and the l a t e r negative component (N3). Note also that the population spike amplitude i s more variable than the presynaptic volley (N1)., Bz_ a single unit evoked i n the v i c i n i t y of N3 following stimulation of PP. This unit was f i r i n g spontaneously during the experiment. Cz evoked discharge of a neurone recorded i n the dentate hilus following stimulation of PP (2 V) and MF (2.5 V) . Note that t h i s neurone i s synaptically activated by MF stimulation indicating that i t i s not a G - c e l l . Pi e f f e c t s of frequency stimulation (10 Hz) on the PP-evoked responses indicating fr a c t i o n a t i o n of late negative component and enhancement of the population spike response. The numbers of pulses giving r i s e to the corresponding response are indicated on the l e f t . 217 eliminate the N3 component. On the basis of the above data, i t i s proposed that N3 represents the polysynaptic activation of neurones i n the dentate hilus. The observations that i t had a longer latency than the population spike and i s maximal i n the region of the hi l u s support t h i s suggestion. The least conspicuous component of the response recorded at the G - c e l l body layer i s the e a r l i e s t negative component (N1 i n Fig. 6-4). Dnlike a l l the components described above, N1 i s not always observed. When present,the H1 component was recorded 0.4-0.8 msec before the onset of P1 component and followed stimulation frequencies greater than 200 Hz which blocked the monosynaptic activation of G-cells (Fig. 6-10D). I t i s l i k e l y that N1 component i s the e x t r a c e l l u l a r r e f l e c t i o n of the perforant path volley recorded i n the region of the G - c e l l layer. This interpretation i s i n accord with the terminology of Lomo (1971a) and Steward et a l (1976).s The e x t r a c e l l u l a r f i e l d responses recorded in the dentate following stimulation of the PP are summarized in Fig. 6-2. Stimulation of the angular bundle produces a PP f i b r e volley in the presynaptic terminals. This i s followed by the depolarization of G-cell dendrites which i s reflected by a large negative f i e l d referred to as the population EPSP. The depolarization 218 £IG*. 6-13i Discharge Pattern Of fientate Granule C e l l s x Ai i n t e r v a l histogram and corresponding oscilloscope sweep i n d i c a t i n g bursting discharge pattern of a unit recorded in the G-c e l l layer. I i r elationship between the discharge pattern of a dentate c e l l and slow e l e c t r i c a l a c t i v i t y recorded in the v i c i n i t y of the microelectrode. The same c e l l i s recorded during BSA (top sweeps) and irr e g u l a r activity(lower sweeps). , C i a non-bursting dentate unit recorded during sensory stimulation (between the two arrows). 2/9 > E IT) — b d 220 d i s c h a r g e s a p r o p o r t i o n of the g r a n u l e c e l l s c a u s i n g t h e n e g a t i v e p o p u l a t i o n s p i k e . F o l l o w i n g G - c e l l d i s c h a r g e s neurones i n the d e n t a t e h i l u s are a c t i v a t e d by mossy f i b r e axon c o l l a t e r a l s g i v i n g r i s e t o the l a t e components of the PP-evoked response. The G - c e l l s p r o j e c t v i a MF on t o p y r a m i d a l c e l l s o f CA3. Qa. S i n g l e U n i t D i s c h a r g e Dentate g r a n u l e c e l l s ( G - c e l l s ) r e c o r d e d i n u r e thane a n a e s t h e t i z e d r a t s f i r e s p o n t a n e o u s l y i n e i t h e r an i r r e g u l a r or b u r s t i n g p a t t e r n a t r a t e s between 2 and 90 Hz. Somatosensory s t i m u l a t i o n such as a t a i l p i n c h i n c r e a s e d t h e f i r i n g r a t e and evoked a b u r s t i n g d i s c h a r g e p a t t e r n i n a p r o p o r t i o n (approx.20%) of G - c e l l s ( F i g . .6-13). T h i s b u r s t i n g d i s c h a r g e p a t t e r n was r e l a t e d to r h y t h m i c a l slow a c t i v i t y r e c o r d e d s i m u l t a n e o u s l y from the v i c i n i t y of the m i c r o e l e c t r o d e o r from b i p o l a r e l e c t r o d e s i n t h e o v e r l y i n g CA1 r e g i o n o f the hippocampus ( c h a p t e r 3 ) . The r e s p o n s e o f s p o n t a n e o u s l y f i r i n g G - c e l l s c o n s i s t s o f s i n g l e s p i k e a c t i v a t i o n a t a l a t e n c y of 3-7 msec f o l l o w i n g s t i m u l a t i o n o f the p e r f o r a n t path ( F i g . 6-14). T h i s s h o r t l a t e n c y a c t i v a t i o n d i d not f o l l o w h i g h f r e g u e n c y (>100 Hz) s t i m u l a t i o n and c o u l d v a r y a p p r o x i m a t e l y 0.5-1.0 msec between c o n s e c u t i v e s t i m u l i a t t h r e s h o l d i n t e n s i t y (ISI=1.0 s e c ) . These m o n o s y n a p t i c a l l y a c t i v a t e d neurones were i d e n t i f i e d as 221 fISi S z l i i i The Response Of An Id e n t i f i e d Dentate Granule C e l l To Stimulation Of The Perforant A: top trace indicates spontaneous discharge and bottom trace shows a c t i v a t i o n - i n h i b i t i o n sequence following single pulse stimulation of PP. BA f a s t sweep speed showing single spike activation which preceded the i n h i b i t i o n . C:. corresponding PST histograms of the above unit following stimulation of PP at two diffe r e n t i n t e n s i t i e s (bin width = 5.0 msec; 50 stimulus presentations i n each case)., 222 223 IiSi. 6zl5^ I dent i f .icat i on Of Dentate Ne urones A Aj. schematic arrangement of recording electrode in the G - c e l l layer and stimulating electrodes in PP and G - c e l l axons (MF). Ei response of a dentate c e l l to stimulation of the PP (left) and MF ( r i g h t ) . Note that the response to PP had a variable latency indicating an interposed synapse, whereas, there i s minimal latency s h i f t following MF stimulation i n d i c a t i n g that i t i s antidromic. , Each record consists of 5 superimposed sweeps. Cj_ the l e f t photo shows a dentate c e l l recorded in the same experiment which was synaptically evoked by MF stimulation i n d i c a t i n g that t h i s particular unit i s not a G-c e l l . The ri g h t photo shows a G-cell which i s evoked by MF stimulation on the antidromic f i e l d and follows high freguency (200 Hz) stimulation. 224 PP • L MF L MF IMF l m s e c 225 G-cells since 1) they were recorded i n the G - c e l l layer, 2) they discharged at the same latency as the population spike and 3) they were often antidromically activated by MF stimulation {Fig, 6-15). C e l l s which were not evoked near the population spike and were synaptically activated by MF stimulation were tent a t i v e l y i d e n t i f i e d as h i l a r neurones {Fig. 6-16). The short latency activation of spontaneously f i r i n g G-c e l l s was always followed by a period of i n h i b i t i o n l a s t i n g 50-300 msec {Fig..6-14). Individual G-cells which were not activated by low i n t e n s i t y stimulation were s t i l l i n h i b i t e d . In a l l cases duration of the i n h i b i t i o n could be lengthened considerably (up to 700 msec) by suprathreshold stimulation of the perforant path (Fig. 6-14), Antidromic activation of G-c e l l s was also followed by i n h i b i t i o n having a similar duration. The i n h i b i t i o n of G-cells i s postulated to occur as a r e s u l t of c o l l a t e r a l a c t i v a t i o n of i n h i b i t o r y interneurones (Gloor, 196 3; Andersen et a l , 1964) . Multiple discharge of singl e G-cells i n response to PP stimulation was r a r e l y encountered. However, i n 3 of 94 experiments the same granule c e l l s were observed to f i r e twice on the positive f i e l d . , I n these cases low in t e n s i t y (1-3 V) stimulation evoked a single spike at 3-7 msec latency whereas higher i n t e n s i t y stimulation resulted i n the appearance of a second spike 2-3 msec following the f i r s t spike. The occurrence of a second 226 FIG^ 6-J6_i Response Of Hilar Neurones Following Stimulation Of The Perforant Path And The Mossy Fibres^ Ai stimulation of either PP or MF synaptically activate the same neurone recorded i n the h i l a r region. The 4 superimposed oscilloscope sweeps in each case indicate that the unit i s evoked after the primary response <N2) and shows a variable latency. Bj. a h i l a r neurone which i s evoked by PP stimulation and appears on the P2 component of the positive wave. The lower trace at slow sweep speed indicates i n h i b i t i o n of the spontaneous discharge following the i n i t i a l a ctivation response. 2Z 7 228 IISA 6-J7: Paired Pulse Potentiation Of The Extrac e l l u l a r EPSP Aj. the peak amplitudes (open c i r c l e s ) and rate of r i s e of the te s t response measured 1.0 msec after i t s onset ( f i l l e d c i r c l e s ) are plotted as a percentage of control responses at the indicated C-T i n t e r v a l s . Bz the negative EPSP recorded i n the dendri t i c layer following conditioning (lower arrow) and test (top arrow) stimulation of the PP at an in t e r v a l of 35 msec. C: superimposed oscilloscope sweeps of the condition ( f i r s t deflection) and the test EPSP*s at i n t e r v a l s of 10-100 msec. 230 spike was dependent on the position of the stimulating electrode in the angular bundle. Multiple discharge of G-cells may thus be a re s u l t of act i v a t i o n of separate afferents from the medial and l a t e r a l entorhinal cortex (McNaughton and Barnes, 1977). Potent ration of Dentate E x t r a c e l l u l a r fie§fiSQ§§§ P2ES2§tipn EPSP Fig. 6-17 shows the e f f e c t s of a conditioning PP volley on the amplitude and rate of r i s e of the population EPSP produced by a subsequent test stimulus to the same pathway. Measurements of the EPSP were made 1.0 msec after i t s onset to eliminate contamination associated with G-cell discharges (see chapter 2). The response to a single test pulse was compared to that e l i c i t e d by a conditioning pulse at the indicated C-T int e r v a l s . Potentiation of the test response was observed at C-T i n t e r v a l s of 20-200 msec with the maximum increase of approximately 150% of control was obtained at i n t e r v a l s of 30-50 msec. At short C-T interva l s (less than 5.0 msec) the size of the test EPSP was much lower than control. This attenuation may be due to refractoriness of the postsynaptic element. I t i s in t e r e s t i n g to note that at the shortest C-T i n t e r v a l s the test presynaptic volley i s actually enhanced at a time when the test EPSP i s depressed. An enhanced presynaptic volley at these short i n t e r v a l s i s 231 FIG_. 6-_18i S t a t i s t i c a l Analysis Of EPSP Potentiation^ A i d i s t r i b u t i o n of EPSP amplitudes evoked by conditioning and test perforant path volleys. Note that there i s less variance i n the amplitude of the test EPSP and that the modal amplitude of the test EPSP i s greater than the control EPS P. Bz_ j o i n t amplitude density plot of EPSP amplitudes evoked by conditioning and test perforant path volleys.,Note that there are no responses i n the f i r s t guadrant in d i c a t i n g that the test EPSP was always larger than the control EPSP.See Chapter 2 f o r description of the analysis. 50 -t JITest O c 3 CT CD o-J 4.0 High CL E < Low '—-Condition Ii Low 5.0 6.0 Amp. (mV) 1L IV Amp. 7.0 High 233 most l i k e l y due to temporal summation of stimulating currents., As shown i n Fig. 6-18, at a given C-T i n t e r v a l the conditioning and test EPSP exhibited unique amplitude d i s t r i b u t i o n s . The fact that at optimal C-T i n t e r v a l s (35 msec in Fig. 6-18) the test EPSP i s r e l i a b l y larger than the condition EPSP i s further demonstrated i n the joint amplitude density plot (described i n Chapter 2) shown i n Fig. 6-188. The 4th guadrant contains the number of t r i a l s (76 out of 84) on which a te s t EPSP was larger than a condition EPSP. In contrast the f i r s t guadrant shows that there were no occasions i n which a conditioning EPSP was larger than a test EPSP. In two separate experiments i n which the variance of amplitudes and latencies of EPSP*s evoked by 500 condition and te s t PP volleys were measured, more variance was observed i n the amplitude and latency of the conditioning volleys. The modal amplitude was greater and the latency of EPSP was shorter (by 0.3-0.5 msec) following the test volley. 233a Leaves 234-235 do not exist . 236 Bj_ Population Spike Fig, 6-19 shows the time course of potentiation of the EPSP recorded by one barrel of an electrode which i s situated i n the synaptic layer and the population spike simultaneously recorded by a second barrel located i n G-cell body layer. At short C-T i n t e r v a l s (0-20 msec) the amplitude of the test population spike i s depressed whereas at longer i n t e r v a l s i t was potentiated 2-3 times control value. It i s important to note that the amplitude of the test spike was depressed at short i n t e r v a l s (10-20 msec) when the test EPSP i s potentiated. In most experiments the population spike was potentiated for much longer C-T i n t e r v a l s (200-400 msec) while the test EPSP was not s i g n i f i c a n t l y d i f f e r e n t at these periods. The magnitudes of depression and potentiation of the population spike were dependent on the in t e n s i t y of the conditioning PP volley. Paired pulse stimulation with low i n t e n s i t y did not give r i s e to the i n i t i a l period of depression but produced marked potentiation (Fig. 6-20). In some cases conditioning stimulation did not evoke a population spike response whereas a test volley of the same intensity resulted i n a large population spike. The magnitude of potentiation of the population spike was up to 4 times greater than that of the population EPSP. This difference in the magnitude of 237 6^19: Paired-pulse Potentiation Of The EPSP And The Population Spike Rj.spqnse.__ ft: the population spike response evoked by a conditioning ( f i r s t response in each sweep) and test (second response) PP pulses. Note depression of of the test population spike at short C-T in t e r v a l s (< 20 msec) and potentiation at longer i n t e r v a l s . Bz amplitude of the tes t pop spike ( f i l l e d c i r c l e s ) and rate of r i s e of EPSP (open c i r c l e s ) plotted as function of C-T i n t e r v a l . Each point i s the average of 20 t r i a l s . 2 3 9 F I G . 6 - 2 O L E f f e c t s Of Stimulus I n t e n s i t y On Paired-p.ulse Potentiation Of The PojguJLation Sjgijse,. Amplitude of the t e s t population spike i s plotted as a function of condition-test (C-T) i n t e r v a l . The intensity of both the conditioning and test pulse was increased. ° i — i 1 1 1 1 1 1 1 1 1 - 2 0 0 +&0 160 240 3 20 C-T Interval (msec) 241 potentiation was not due to reaching a maximal EPSP amplitude since reducing the intensity of the conditioning and test PP volleys also did not re s u l t in potentiation of the EPSP to the same degree as that of the population spike. Furthermore, a larger EPSP than that evoked by the test pulse could be evoked by increasing the i n t e n s i t y of the test volley. Therefore i t i s unlikely that the lesser degree of potentiation of the EPSP r e l a t i v e to the population spike i s due to maximal activation of the postsynaptic elements. £t sLAaale Unit Discharge As shown in Fig, 6-21, paired pulse stimulation of the perforant path also resulted i n potentiation of single spike discharge. This potentiation occurred either as an increase i n the number of neurones e l i c i t e d by the test volley or a reduced latency to discharge of i n d i v i d u a l neurones. As shown i n the middle trace of Fig. 6-21, spontaneously f i r i n g neurones which were not evoked by a conditioning volley of low in t e n s i t y appear on the positive f i e l d produced by the test pulse. As Fig. 6-21 shows potentiation of i n d i v i d u a l spike discharge occurred at a time when the population spike was also potentiated inasmuch as a G-cell that was not evoked on the control population spike was evoked coincidently with the potentiated population 242 kz2.ll Potentiation Of G-cell JiscJaj§£ge---By.-Pai£S^ --Pulse Stimulation Of The Perforant Path A A. j_ spontaneously f i r i n g G-cell which i s inhib i t e d but not excited by a singl e conditioning PP volley. Bz response of the above neurone to double pulse stimulation i n d i c a t i n g that i t i s evoked by the test and not the conditioning volley. Cj; c orrelation between potentiation of the population spike and singl e unit discharge (arrow) . The unit was not evoked on the conditioning population spike (top sweep) but appears on the test population spike (lower sweep). £ 4 J 50 msec. J|_Jo.2mV 10 msec. •Control Test T 2.0 mV 10 msec. 244 s p i k e . As i n t h e case o f t h e p o p u l a t i o n s p i k e r e s p o n s e , s t r o n g c o n d i t i o n i n g v o l l e y s which evoked a s i n g l e s p i k e r e s u l t e d i n i n h i b i t i o n o f t h e t e s t s p i k e a t s h o r t (5-20 msec) C-T i n t e r v a l s . I t i s noteworthy t h a t at C-T i n t e r v a l s l o n g e r than 20 msec p o t e n t i a t i o n o f s i n g l e s p i k e d i s c h a r g e o c c u r s at a t i m e when t h e spontaneous d i s c h a r g e of t h e same G - c e l l i s c o m p l e t e l y i n h i b i t e d by t h e c o n d i t i o n i n g v o l l e y . , T h i s s u g g e s t s t h a t mechanisms u n d e r l y i n g p o t e n t i a t i o n of G - c e l l d i s c h a r g e can overcome the s i m u l t a n e o u s l y o c c u r r i n g i n h i b i t i o n of G-c e l l s . However a t s h o r t C-T i n t e r v a l s the p o t e n t i a t i o n i s n ot s u f f i c i e n t t o overcome t h e p o t e n t i n h i b i t i o n . 6 g_ 4 D i s c u s s i o n F i e l d A n a l y s i s Of The P e r f o r a n t Path I n p u t To Th.e D entate Gyrus S t u d y i n g f i e l d p o t e n t i a l s i n an area t h a t has a s i m p l e l a m i n a r o r g a n i z a t i o n a l l o w s a comparison between a n a t o m i c a l l y v e r i f i e d t e r m i n a l a r e a s and e l e c t r o p h y s i o l o g i c a l r e s p o n s e s . E l e c t r i c a l s t i m u l a t i o n o f PP r e s u l t s i n an e x t r a c e l l u l a r n e g a t i v e f i e l d which i s maximal i n t h e m i d d l e m o l e c u l a r l a y e r . The same zone c o n t a i n s the d e n s e s t t e r m i n a l d e g e n e r a t i o n f o l l o w i n g e n t o r h i n a l l e s i o n s as o b s e r v e d i n l i g h t ( B l a c k s t a d , 1956) and e l e c t r o n ( N a f s t a d , 1967) m i c r o s c o p i c i n v e s t i g a t i o n s i n t h e r a t . T h i s e x t r a c e l l u l a r n e g a t i v e f i e l d {shown i n F i g . 6-22) may r e p r e s e n t t h e summed 245 EPSP's r e s u l t i n g from the d i r e c t action of PP terminals on the dendrites of G-cells since 1) i t follows the presynaptic volley by 0.4-0.8 msec which i s within one synaptic delay, 2) i t follows moderate but not high freguency stimulation and 3} i t has been recorded simultaneously with i n t r a c e l l u l a r EPSP's of i n d i v i d u a l G-cells (Lomo, 1971a). This e x t r a c e l l u l a r EPSP may be due to an active sink r e s t r i c t e d to the l e v e l of the PP synapses drawing current from passive sources in the area of the c e l l body. Previous studies have also interpreted the negative response recorded in the molecular layer as synaptic current r e f l e c t i n g the dir e c t action of PP transmitter on the dendrites of G-c e l l s (Lomo, 1971a; Steward et a l , 1 976). In the region of the granule c e l l body PP stimulation evokes a positive waveform which, i n part, appears as the mirror image of the e x t r a c e l l u l a r EPSP for which i t i s a source. Individual G-calls are evoked on the p o s i t i v e f i e l d and increased stimulating current synchronously discharges a large number of G-cells giving r i s e to a large negative spike which i s referred to as the population spike response. The observations that the units giving r i s e to the population spike are G-cells i d e n t i f i e d by antidromic a c t i v a t i o n and h i s t o l o g i c a l l o c a l i z a t i o n of the recording electrode supports the suggestion that the N2 component shown in Fig. 6-4 i s the population spike of G-cells. These conclusions are supported by previously published 246 experiments i n rabbits and cats (Gloor et a l , 1964; Lomo, 1971a), In addition to the monosynaptic activation of G-c e l l s , polysynaptic activation of interneurones via axon c o l l a t e r a l s of G-cells give r i s e to the late components of the f i e l d recorded i n the area of the c e l l body layer (N3). The observation that s i m i l a r l a t e components are recorded following activation of mossy f i b r e s or stimulation Of the commissural pathway (see chapter 7), suggests that the late components are not a unigue feature of the PP input onto G-cells but may r e f l e c t the a c t i v i t y of neurones i n the area of the dentate h i l u s . This may explain the f a i l u r e to f i n d stimulating s i t e s in the angular bundle which were not associated with late responses and the f a c t that the l a t e negative component (N3 in Fig. 6-4) was not abolished by lesioning commissural pathways. Response Of Spontaneously Firing. G-cells,,-The spontaneous discharge pattern of G-cells consists of both i r r e g u l a r and bursting discharge patterns. The bursting discharge pattern may underly slow rhythmical e l e c t r i c a l a c t i v i t y recorded i n the dentate (See chapter 3, Bland et a l , 1976)., Single pulse stimulation of the PP r e s u l t s i n single spike a c t i v a t i o n of G-cells at short latency (3-7 msec) which corresponds to the population spike response, This 247 6-22: Interpretation Of The Responses Recorded In Th§ Dentate Following PP Stimulation, r A stimulating electrode i n the PP activates axons o r i g i n a t i n g i n the entorhinal cortex (EC) and r e s u l t s i n a stimulus a r t i f a c t recorded i n the dentate at time (T)=0. The stimulus propogates to PP terminals and r e s u l t s in a small f i b r e volley (T=1). The release of PP transmitter causes EPSP's on G - c e l l dendrites and the associated inward current results in a negative e x t r a c e l l u l a r potential (T=2) i n the molecular layer (top profile) and a positive potential at the c e l l body layer (middle p r o f i l e ) . The subseguent discharge of G-cells (bottom schematic) re s u l t s in a negative population spike . (T=3) ... Activation of interneurones (T=4) by G - c e l l axon c o l l a t e r a l s i n h i b i t s the discharge of G-cells and gives r i s e to the long latency components of the f i e l d responses (T=5). 249 a c t i v a t i o n i s f o l l o w e d by p r o l o n g e d i n h i b i t i o n which may be caused by r e c r u i t m e n t of i n h i b i t o r y i n t e r n e u r o n e s l o c a t e d i m m e d i a t e l y below t h e G - c e l l l a y e r (Andersen e t a l , 1964). The o b s e r v a t i o n t h a t some c e l l s which a r e not a c t i v a t e d by t h e PP a r e s t i l l i n h i b i t e d s u g g e s t s t h a t t h e s e i n h i b i t o r y i n t e r n e u r o n e s p r o j e c t t o n e i g h b o u r i n g c e l l s . T h i s i s s u p p o r t e d by G o l g i s t u d i e s i n which t h e f u s i f o r m c e l l s below the g r a n u l e c e l l s were observed t o have l o n g axons which make c o n t a c t w i t h G - c e l l s up t o 1.0 mm away ( C a j a l , 1911). T h i s p o t e n t i n h i b i t i o n which i n p o s t - s t i m u l u s h i s t o g r a m s appears from 0-5 msec f o l l o w i n g t h e a c t i v a t i o n o f G - c e l l s may p r e v e n t the m u l t i p l e d i s c h a r g e of G - c e l l s i n response to a p e r f o r a n t path v o l l e y . The few cases i n w h i c h t h e same c e l l was observed t o f i r e t w i c e on the p o s i t i v e f i e l d may be a r e s u l t of a c t i v a t i n g two s e p a r a t e a f f e r e n t s c o u r s i n g t h r o u g h the a n g u l a r bundle. McNaughton and Barnes (1977) have r e c e n t l y shown t h a t a f f e r e n t s from the l a t e r a l and the m e d i a l e n t o r h i n a l c o r t e x which p r o j e c t t o the o u t e r and middle m o l e c u l a r l a y e r , r e s p e c t i v e l y , may be a c t i v a t e d by a n g u l a r bundle s t i m u l a t i o n . The second s p i k e which o c c u r r e d 2-3 seconds a f t e r t h e i n i t i a l s p i k e may be t h e r e s u l t o f EPSP* s g e n e r a t e d on the most d i s t a l d e n d r i t e s and would t h u s r e a c h the soma a f t e r mere p r o x i m a l s y n a p t i c c u r r e n t s ( B a l l and R i n z e l , 1973) . 250 The D i s c h a r g e o f Neurones Other Than G - c e l l s The d i s c h a r g e o f neurones o t h e r than G - c e l l s may have been r e c o r d e d i n t h e area o f t h e G - c e l l body l a y e r and a d j a c e n t h i l a r zone. U n f o r t u n a t e l y , r e c o r d i n g s were h a r d t o o b t a i n from t h e s e n e i g h b o u r i n g neurones because they generate s m a l l a m p l i t u d e s p i k e s and may be o b s c u r e d by t h e G - c e l l p o p u l a t i o n r e s p o n s e s . I n a few c a s e s when s t a b l e r e c o r d i n g s were o b t a i n e d from t h o s e neurones, c o n v e r g i n g e x c i t a t o r y i n p u t s from PP and mossy f i b r e s at l a t e n c i e s l o n g e r t h a n t h o s e o f G - c e l l s were r e a d i l y demonstrated. Those neurones may t h u s be a c t i v a t e d by G - c e l l axon c o l l a t e r a l s . S i n c e G o i g i s t u d i e s r e v e a l a heterogeneous p o p u l a t i o n of c e l l t y p e s i n t h e r e g i o n o f t h e h i l u s t h e s e neurones a r e u n l i k e l y t o produce synchronous p o p u l a t i o n s p i k e s s i m i l a r t o those g e n e r a t e d by t h e u n i f o r m d i s c h a r g e of G - c e l l s . I t i s p o s s i b l e t h a t the c e l l s which were a c t i v a t e d by both PP and MF s t i m u l a t i o n c o r r e s p o n d t o the a n a t o m i c a l l y d e s c r i b e d f u s i f o r m c e l l s which p r o j e c t t o t h e G - c e l l b o d i e s . These c e l l s would thus be t h e i n h i b i t o r y i n t e r n e u r o n e s p o s t u l a t e d by Andersen et a l (1966) and s c h e m a t i c a l l y shown i n F i g , 6-22. 251 Potentiation 9_f The Extrace 11 ular EPSP The above discussion has presented evidence that the f i r s t 1-1.5 msec of the negative dendritic response r e f l e c t s the synaptic current generated by the perforant path. An increase in the amplitude and rate of r i s e of t h i s response occurs 5-200 asec following a conditioning pulse to the same pathway. Such potentiation could occur either as a r e s u l t of an increase i n the size of the afferent test volley or due to an increase in e f f i c i e n c y of synaptic transmission between the perforant path and the G-cell dendrites. The f i r s t mentioned p o s s i b i l i t y of presynaptic involvement i s unlikely since the amplitude of the PP volley which r e f l e c t s presynaptic current was not potentiated i n t h i s and previous experiments (Lomo, 1971b). However, t h i s argument i s based on the assumption that the amplitude of the presynaptic volley r e f l e c t s the a c t i v i t y of presynaptic terminals i n control and potentiated states. The data presented in Fig, 6-4 shows that t h i s i s the case i n control s i t u a t i o n s since an increase in stimulus i n t e n s i t y enhances the amplitude of the presynaptic volley and the EPSP, Perhaps i n the potentiated state terminals other than those which are activated in control states and which do not give rise to the presynaptic volley are activated. On the basis of the present data one cannot eliminate t h i s p o s s i b i l i t y . 252 An i n c r e a s e i n t h e e f f e c t i v e n e s s of t r a n s m i s s i o n between the p e r f o r a n t path t e r m i n a l s and t h e d e n d r i t e s of G - c e l l s may be mediated by an augmented r e l e a s e of t r a n s m i t t e r and/or changes i n the p o s t s y n a p t i c element which s u b s t a n t i a l l y i n c r e a s e the s y n a p t i c c u r r e n t g e n e r a t e d by the same amount of t r a n s m i t t e r . The mechanisms t h a t c o u l d g i v e r i s e t o an i n c r e a s e i n e x t r a c e l l u l a r s y n a p t i c c u r r e n t w i t h o u t i n v o l v i n g changes i n t r a n s m i t t e r r e l e a s e i n c l u d e enhanced s e n s i t i v i t y o f t h e p o s t s y n a p t i c membrane and m o r p h o l o g i c a l changes i n d e n d r i t i c s p i n e s which a f f e c t t h e i r c u r r e n t g e n e r a t i n g c h a r a c t e r i s t i c s ( B a l l , 1970). However, the above changes i n the p o s t s y n a p t i c elements must be s p e c i f i c t o t h e a r e a a d j a c e n t t o PP synapses s i n c e c o n d i t i o n i n g s t i m u l a t i o n o f c o m m i s s u r a l a f f e r e n t s which t e r m i n a t e a d j a c e n t t o t h e PP synapses does not a l t e r the r a t e of r i s e of a t e s t EPSP evoked by the PP (Steward et a l , 1977; See a l s o Chapter 7 ) . P o t e n t i a t i o n Of The P o p u l a t i o n S p i k e On the b a s i s o f t h e p r e s e n t d a t a and t h a t of Andersen e t a l (1971) the a m p l i t u d e of t h e p o p u l a t i o n s p i k e r e s p o n s e i s r e l a t e d t o t h e number o f i n d i v i d u a l G - c e l l s which are a c t i v a t e d f o l l o w i n g s t i m u l a t i o n o f t h e p e r f o r a n t p a t h . Double p u l s e s t i m u l a t i o n of PP r e s u l t s i n a marked i n c r e a s e i n the a m p l i t u d e o f t h e PP s p i k e evoked by t h e t e s t v o l l e y r e l a t i v e t o t h a t evoked by t h e c o n d i t i o n i n g s t i m u l u s . Such p o t e n t i a t i o n c o u l d 253 o c c u r e i t h e r as a r e s u l t o f an i n c r e a s e i n s y n a p t i c c u r r e n t t h u s p r o d u c i n g a g r e a t e r d e p o l a r i z a t i o n o f G-c e l l s o r by an enhanced p r o b a b i l i t y t h a t a g i v e n G - c e l l i s evoked by the t e s t v o l l e y . The l a t t e r c o u l d be due t o a l o w e r i n g o f t h e t h r e s h o l d f o r t h e g e n e r a t i o n of an a c t i o n p o t e n t i a l by an i n d i v i d u a l G - c e l l or due t o an i n c r e a s e i n t h e e f f e c t i v e n e s s of d e n d r i t i c d e p o l a r i z a t i o n s r e a c h i n g the soma. As d i s c u s s e d i n t h e p r e v i o u s s e c t i o n , t h e r e i s an i n c r e a s e i n the s y n a p t i c c u r r e n t g e n e r a t e d by the t e s t PP v o l l e y which c o u l d i n p a r t account f o r p o t e n t i a t i o n o f the p o p u l a t i o n s p i k e response. However, t h e magnitude o f p o t e n t i a t i o n o f t h e EPSP ( l e s s t h a n 150%) was u s u a l l y much l e s s t h a t the degree o f p o t e n t i a t i o n o f the p o p u l a t i o n s p i k e (up t o 700%). F u r t h e r m o r e i n some a n i m a l s p o t e n t i a t i o n of the p o p u l a t i o n s p i k e r e s p o n s e was o b s e r v e d i n t h e absence o f apparent changes i n t h e r a t e of r i s e of the EPSP (See a l s o Lomo, 1971b). A comparison between the time c o u r s e of p o t e n t i a t i o n o f t h e EPSP and the p o p u l a t i o n s p i k e ( F i g , 6-19) r e v e a l s t h a t t h e a m p l i t u d e o f t e s t p o p u l a t i o n s p i k e i s g r e a t e r than c o n t r o l a t l o n g C-T i n t e r v a l s (over 200 msec) when t h e t e s t EPSP i s not d i f f e r e n t from c o n t r o l . I t i s a l s o noteworthy t h a t at s h o r t C-T i n t e r v a l s (0-20 msec) the a m p l i t u d e o f t h e t e s t PP s p i k e i s dep r e s s e d at a t i m e when the EPSP i s 254 potentiated., Depression of the population spike amplitude at short C-T i n t e r v a l may be due to potent recurrent i n h i b i t i o n i n i t i a t e d by the conditioning volley (Lomo, 197 1b; Steward et a l , 1 976). Perhaps potentiation of the population spike at long C-T in t e r v a l s i s due to the action of polysynaptic mechanisms on G-cells. The re l a t i o n s h i p between paired-pulse potentiation discussed above and long-lasting potentiation resulting from tetanic stimulation of the PP i s not cl e a r . B l i s s and Lomo (1973) have shown that the rate of r i s e of the EPSP and amplitude of the population spike could be potentiated for periods ranging from 30 minutes to 10 hours following a brief t r a i n (3-4 sec) of high frequency (100 Hz) stimulation. B l i s s , Gardner-Medwin (1973) reported that long-lasting potentiation was not produced on a l l occasions when there was paired pulse stimulation. Furthermore, these studies (Bliss and Lomo, 1973; Bliss and Gardner-Medwin, 1973) observed increases in the amplitude of the population spike with no change in the synaptic current. This observation and the fact that the magnitude and time course of potentiation of the population spike d i f f e r from those of the EPSP suggest that at lea s t a component of long-l a s t i n g potentiation may be due to changes in tonic influences acting on the G-cells. The above suggestions imply that potentiation of 255 the population spike could occur i n absence of changes in transmitter release and may be dependent on the a c t i v i t y of the postsynaptic elements. In order to examine t h i s p o s s i b i l i t y , the effects of stimulating e x t r i n s i c afferents to the dentate on PP-evoked responses w i l l be studied in the remaining chapters. 256 CHAPTER 7.. NEURONAL I S I E S M I S S T O N IN THE DE N TA TE -GIBUS: ROLE OF THE COSHISSURAL INPUT Z*JL XlLtrgduct ion The previous chapter characterized the f i e l d responses recorded in the dentate following stimulation of the perforant path (PP), Homosynaptic potentiation of t h i s c o r t i c a l input involved both an increase in synaptic current and amplitude of the population spike response. I t was suggested that the increased e f f i c a c y of the PP input may could be a re s u l t of changes in the postsynaptic c e l l i n addition to those postulated to occur i n the presynaptic elements of th i s pathway. Since the termination of the commissural (COMM) input to the dentate gyrus i s r e s t r i c t e d to a zone extending 50-100 um above the G - c e l l layer and just below the PP synapses (Blackstad, 1956; Gottlieb and Cowan, 1 9 7 3 ) , activation of the COMM input may generate synaptic current at s i t e s adjacent to PP synapses. This arrangement allows an examination of the p o s s i b i l i t y that conditioning COMM stimulation potentiates PP-evoked responses. The aim of the present chapter i s to: 1) determine the laminar p r o f i l e of synaptic currents recorded in the dentate following stimulation of the COMM input. 2) examine whether conditioning stimulation of the COMM input a l t e r s the f i e l d responses evoked by a test PP 257 volley. Zi.2 2x2erimental Procedures. Extra c e l l u l a r recordings were performed on a t o t a l of 15 urethane anaesthetized rats. Concentric bipolar electrodes were positioned in the angular bundle to activate the perforant path and in the co n t r a l a t e r a l CA3-CA4 area of the hippocampus to activate the commissural projection at i t s or i g i n . , In some experiments an additional bipolar electrode was positioned i n the commissural pathway as i t courses through the ventral psalterium (1.5 mm posterior to bregma; 0.5-2.0 mm l a t e r a l ; 3.0 mm below the c o r t i c a l surf ace), Single unit a c t i v i t y and f i e l d potentials were recorded from the dentate as described i n the previous chapters. The responses to 20-30 stimulus presentations (ISI 2.5-4.0 seconds) were averaged and plotted at predetermined depths to allow a comparison between the f i e l d responses evoked by stimulation of PP and COM pathways. 258 7__3 Results Fig.,7-1 shows the extracellular f i e l d potentials recorded at indicated depths i n the dentate following stimulation (5 V) of the PP and Comm pathways. As shown i n the previous chapter, PP stimulation results i n a short latency negative wave which i s maximal 175-225 um above the G-cell layer and a positive wave with a superimposed population spike recorded at c e l l body layer. COMM stimulation also r e s u l t s i n a short latency (4-7 msec at i t s peak) f i e l d response in the dentate. When the recording electrode is situated in the most d i s t a l dendrites a low amplitude positive waveform i s recorded. This p o s i t i v e wave reverses to a negative wave at the l e v e l of the PP synapse and reaches a maximum negativity 100 um above the G - c e l l layer. As the recording electrode i s advanced further towards the G-cell layer the e x t r a c e l l u l a r negativity reverses to a positive wave at approximately 25 um above the c e l l bodies. 259 : l l Lamin ax P r o f i l e s Of The F i e l d Potentials J22&e4 la The Dentate By. Stimulation Of The Perforant Path A_n d Cgmmissural Pathway, F i e l d potentials evoked by PP (left) an COHM (right) volleys recorded i n the upper blade of DG along a v e r t i c a l track at the indicated depths below the surface of the cortex. Each record i s the average of 20 t r i a l s and the arrows indicate the stimulus a r t i f a c t s . 2GO Depth (JUM) , - J 2 M V t _ C O M M 5 V 5 msec 261 I d e n t i f i c a t i o n Of The Dentate Responses Aj_ M o l e c u l a r Layer F i g . 7-1 shows t h a t t h e most prominent response i n t h e m o l e c u l a r l a y e r i s a n e g a t i v e f i e l d which i s maximal i n the i n n e r m o l e c u l a r l a y e r a t a d i s t a n c e of 75-125uM above the G - c e l l l a y e r . The onset o f t h i s f i e l d o c c u r s 3-5 msec a f t e r s t i m u l a t i o n o f the c o n t r a l a t e r a l CA3 or 2-4 msec f o l l o w i n g d i r e c t a c t i v a t i o n o f t h e COMM pathway i n the v e n t r a l p s a l t e r i u m (1.5 mm from the m i d l i n e ) . , I n the same ani m a l t h e onset and peak l a t e n c i e s o f t h i s n e g a t i v e wave are c o n s i s t e n t and l a t e n c y s h i f t s g r e a t e r than 0.5 msec were r a r e l y e n c o u n t e r e d . T h i s n e g a t i v e response f o l l o w e d moderate (1-75 Hz) but not h i g h (>200 Hz) f r e g u e n c y s t i m u l a t i o n . S i n c e t h i s n e g a t i v e wave i s maximal i n t h e i n n e r m o l e c u l a r l a y e r a t the same l e v e l as the t e r m i n a t i o n o f the COMM pathway ( G o t t l i e b and Cowan, 1973; H j o r t h -Simonsen and L a u r b e r g , 1978) , i t may be t h e e x t r a c e l l u l a r r e f l e c t i o n of a monosynaptic EPSP r e f l e c t i n g t h e d e p o l a r i z a t i o n o f a p i c a l l y o r i e n t e d d e n d r i t e s . A l t h o u g h i n t r a c e l l u l a r d a t a i s l a c k i n g , t h e s i m i l a r i t y between t h i s n e g a t i v e d i p o l e and t h a t produced by the PP { d e s c r i b e d i n c h a p t e r 6) s u p p o r t s t h i s s u g g e s t i o n . S i n c e t h e maximal n e g a t i v i t y i s 75-125 um above t h e G - c e l l l a y e r , t h e d e p o l a r i z i n g 262 s y n a p t i c c u r r e n t may be generated by t h e p r o x i m a l d e n d r i t e s of G - c e l l s , A s i m i l i a r i n t e r p r e t a t i o n was p r e v i o u s l y proposed by Deadwyler e t a l (1975). However, the p o s s i b i l i t y t h a t t h e s e c u r r e n t s a r e generated by o t h e r s t r u c t u r e s such as t h e d i s t a l d e n d r i t e s of neurones whose c e l l b o d i e s are below t h e G - c e l l l a y e r can not e l i m i n a t e d . Long. L a t e n c y Responses As shown i n F i g , ,7-2, t h e above s h o r t l a t e n c y r e s p o n s e s were sometimes (6 of 15 anim a l s ) f o l l o w e d by secondary r e s p o n s e s h a v i n g onset l a t e n c i e s between 15 and 20 mSec, S i n c e these l a t e responses d i s p l a y a g r e a t e r s h i f t i n l a t e n c y than the e a r l i e r components and do not f o l l o w 20-50 Hz s t i m u l a t i o n ( F i g . 7-3B) they may be evoked p o l y s y n a p t i c a l l y . The l a m i n a r p r o f i l e of the l a t e component was not s i m i l a r t o the e a r l i e r r e sponses evoked by COMM s t i m u l a t i o n but was r e l a t e d t o th e PP-evoked p r o f i l e ( F i g . 7-2B). However, s i m i l a r l a t e components were never observed f o l l o w i n q s t i m u l a t i o n o f t h e PP i t s e l f . The l a t e components were c o m p l e t e l y e l i m i n a t e d by a t r a n s e c t i o n o f t h e i p s i l a t e r a l p e r f o r a n t path. As shown i n F i g . 7-3C, i f the k n i f e c u t i s made between the e n t o r h i n a l c o r t e x (EC) and the PP s t i m u l a t i n g e l e c t r o d e , the l a t e COMM response d i s a p p e a r s whereas the PP re s p o n s e i s normal. H j o r t h - S i m o n s e n (1973) has 262a Leaf 262 continued on 271 • 263 FIG*. ~Lz 2l Laminar P r o f i l e s Of Primary And Secondary I i§14 Potentials Recorded In The Dgntat-e-loilowiaa Stimulation Of The COMM Pathway..; depth p r o f i l e s recorded at the indicated distance above the granule c e l l layer following stimulation of the PP (lower oscilloscope sweeps) and COMM pathway i n the ventral psalterium (upper oscilloscope sweeps).,Note the appearance of secondary responses (obligue arrows) following COMM stimulation. B:, p r o f i l e of the amplitude of the early (COMM1) and l a t e (COMM 2) evoked potentials measured 1 msec a f t e r t h e i r onset. P r o f i l e of PP evoked potentials i s included for comparison., Note that the late COMM2 response p a r a l l e l s the PP p r o f i l e . 264 265 FIG.. 7- 3: Characterist ics Of Early. And Late f i e l d Responses Hecgrded In The Dentate Foilowing Stimulation Of The Commissural Pathway. hi effects of stimulus intensity oh the amplitude of responses recorded in the molecular layer. Each record i s 5 superimposed sweeps at the indicated stimulus i n t e n s i t y . Note that the l a t e components are only produced at higher voltages. , JBj. e f f e c t of freguency stimulation on the early and late responses i n d i c a t i n g the polysynaptic nature of the late response since i t does not follow moderate (40-80 Hz) stimulus frequencies. ; Cz_ elimination of the l a t e component but not the early response following transection of the PP suggesting that the late response i s polysynaptically relayed v i a the entorhinal cortex. 1\) (Ts 266a Leaves 267-270 do not exist. 271 provided anatomical evidence for a d i r e c t projection from CA3 to the i p s i l a t e r a l EC which gives r i s e to the PP. Thus the late responses could r e s u l t from activation of the entorhinal cortex and subsequent depolarization of G - c e l l dendrites. This p o s s i b i l i t y i s strengthened by the observations of Deadwyler et a l (1975) that stimulation of CA3 evokes unit discharges in EC which precede the late dentate responses fay 1,5 msec. Although no single EC units were recorded in the present investigation f i e l d responses which precede the dentate l a t e component by 5-7 msec were recorded i n the EC of 1 animal. Although Deadwyler et al (1975) suggest that the i p s i l a t e r a l entorhinal cortex was activated d i r e c t l y by contralateral hippocampus, i t i s also possible that antidromic activation of axon c o l l a t e r a l s of the i p s i l a t e r a l CA3 results in activation of the EC on the same side. The l a t t e r p o s s i b i l i t y i s more l i k e l y since there i s no direct anatomical evidence f o r a contalateral projection. ]3. G-cell Layer Fig. 7-4 shows the e x t r a c e l l u l a r f i e l d potentials evoked by COMM and PP stimulation and recorded i n the c e l l body layer. An early negative deflection i s recorded at the same latency as the presynaptic volley which was observed i n the molecular layer and 0.5-0.8 msec before the onset of the e a r l i e s t c e l l discharge or compound spikes (indicated by oblique 272 FIG t 7- 41 Fi e l d Potentials Recorded In The Granule C e l l Layer Following Stiffiulatisn Of The PjISfor&St Pa th And, £ogfflissu£a3, Pafeh»ay§A 4i comparison between the responses evoked by PP and COMM stimulation at the indicated voltages. In each case a negative population spike i s indicated by the arrow, Bi i n t e r a c t i o n between PP and COMM evoked responses at the indicated C-T in t e r v a l s . Note depression of the negative spike at the shorter C-T i n t e r v a l suggesting convergence of the two inputs on the units giving r i s e to thi s response. (2 7 5 274 arrows i n Fig. 7-4). I t may thus represent the presynaptic volley of the COMM pathway. The reason that i t i s more prominent than the PP f i b r e volley recorded from the same s i t e may be rela t e d to the observations that COMM f i b r e s pass through the G-cell layer en route to the inner molecular layer. In 2 of 15 animals a prominent spike was recorded at the same latency (2.5 msec) as the f i b r e v o l l e y . This spike was less than 0.2 msec i n duration and may thus be caused by a single axon. Following the f i b r e volley a large compound spike (oblique arrow) i s sometimes (in 4 of 15 animals) recorded in response to COMM stimulation. However, the stimulating voltages reguired to produce a compound spike was 3 times higher than those reguired to evoke a prominent dendritic potential. The amplitude of the compound spike was maximal at the same depth as the PP-evoked population spike in the G-cell layer. The COMfl-evoked spike was markedly depressed when preceded (1-20 mSec) by a PP-evoked population spike. On the other hand, i f a COMM volley precedes PP stimulation the PP-evoked population spike i s depressed. , The compound spikes recored following commissural stimulation may r e f l e c t the synchronous discharge of a population of dentate neurones and w i l l thereby be referred to as the COMM-evoked population spike. I t i s noteworthy that the COMM evoked population spike always occurs early on the descending limb of the positive wave, whereas, the PP Leaf 274 continued on 277. Leaves 275-276 do not exist. 277 evoked p o p u l a t i o n s p i k e may occur l a t e on the des c e n d i n g l i m b o r on the a s c e n d i n g l i m b . T h i s may be due t o a s h o r t e r c o n d u c t i n g d i s t a n c e between COMM synapses on p r o x i m a l d e n d r i t e s t h a n PP synapses on d i s t a l d e n d r i t e s . The l a s t n e g a t i v e component has not y e t been i d e n t i f i e d b u t i t s s i m i l a r i t y t o the N3 component f o l l o w i n g PP s t i m u l a t i o n s u g g e s t s t h a t i t a l s o r e p r e s e n t s the p o l y s y n a p t i c a c t i v a t i o n of neurones i n th e d e n t a t e h i l u s . Cj_ S i n g l e U n i t D i s c h a r g e The response of 60 s p o n t a n e o u s l y f i r i n g d e n t a t e c e l l s was s t u d i e d f o l l o w i n g s t i m u l a t i o n of both th e PP and COMM pathway. Of t h o s e , 11 (18%) were a c t i v a t e d 4-8 msec f o l l o w i n g s t i m u l a t i o n of the COMM pathway (5-10 V) and 21 (35%) were a c t i v a t e d by the p e r f o r a n t p a t h (3-5 V ) . The PP was always s t i m u l a t e d w i t h l o w e r i n t e n s i t y i n o r d e r t o a v o i d p o p u l a t i o n s p i k e s which even i n a f i l t e r e d r e c o r d o b s c u r e d t h e d i s c h a r g e of i n d i v i d u a l neurones. As shown i n F i g . 7-5, convergence o f COMM and PP i n p u t s on 9 neurones was demonstrated by p a i r i n g t he two i n p u t s at sub and s u p r a t h r e s h c l d i n t e n s i t i e s and a range of c o n d i t i o n - t e s t i n t e r v a l s . Two s u b t h r e s h o l d s t i m u l i p a i r e d a t 1-5 msec i n t e r v a l s r e s u l t e d i n a s p i k e f o l l o w i n g the second p u l s e ( F i g . 7.5B, bottom t r a c e ) s u g g e s t i n g t h a t 277a Leaf 277 continued on 282. 278 7- 5_i. Response Qf Spontaneously F i r Dentate Neurones Following Stimulation Of The Perforant Path And Commissura1 Inputs i i top: f a s t sweep speed i n d i c a t i n g s i n g l e spike a c t i v a t i o n of the same neurone following stimulation of PP ( l e f t arrows) and COMM (right arrows).bottom: same c e l l at slow sweep speed indicating i n h i b i t i o n following the a c t i v a t i o n . Note that COMM stimulation produced longer i n h i b i t i o n . B: interaction between PP and COMM stimulation indicating possible convergence of the two inputs on the same neurone shown i n A. ,, Note that both pulses at suprathreshold intensity e l i c i t a spike (top) but when the C-T i n t e r v a l i s shortened the response to the second pulse i s occluded (middle) . In contrast, a spike i s only e l i c i t e d following the second of two subthreshold pulses (bottom) . 17 7 PP , ,. > « * * w ESSE COMM " J o . 2 mV 5 msec •Jo.2 mV 50 msec B. P P - C O M M ( l.5xT) P P - C O M M (1.5 xT) P P - C O M M (0.5 xT) 5 msec LEAVES 280 AND 281 OMITTED IN PAGE NUMBERING. 282 temporal/spatial summation had occurred. However, pairing two suprathreshold pulses at the same c r i t i c a l i n t e r v a l resulted in a spike following the f i r s t but not the second pulse (Fig. 7-5; middle trace) due to either refractoriness of the neurone or recurrent i n h i b i t i o n which i s described below. The short latency activation of spontaneously f i r i n g neurones e l i c i t e d by COMM stimulation was always followed by a prolonged period of i n h i b i t i o n l a s t i n g 50-800 msec. Spontaneous units (49 out of 60) that were not activated by COMM stimulation were s t i l l i n h i b i t e d by low i n t e n s i t y stimulation (1-3 V) of the COMM pathway. The length of i n h i b i t i o n following stimulation of the COMM pathway at 1.5T inte n s i t y reguired to activate a neurone was usually 2-3 times longer that e l i c i t e d by stimulation of the PP at the same r e l a t i v e i n t e n s i t y (1.5 X threshold). Origin Of The Commissural Pathway, H i s t o l o g i c a l l y v e r i f i e d s i t e s for evoking the responses described above were located i n the CA3-CA4 subfield of the cont r a l a t e r a l dorsal hippocampus Responses having the shortest latency and maximum amplitude were obtained following stimulation s i t e s which were nomotopic to the recording electrode. If the stimulating electrode was moved 0.5-1.0 mm from this optimal s i t e a positive-going wave was recorded in the Leaf 282 continued on 287. Leaves 283-286 do not e x i s t . 287 d e n t a t e independent of the depth of the r e c o r d i n g e l e c t r o d e . .Placements which were 2.5-3.0 mm from the m i d l i n e were more e f f e c t i v e than temporal placements i n th e r e g i o n o f t h e f i m b r i a . E l e c t r o d e placements i n the v e n t r a l p s a l t e r i u m were more e f f e c t i v e i n e v o k i n g d e n t a t e f i e l d r e s p o n s e s than t h o s e s i t u a t e d i n the hippocampus p r o p e r . ill§£ts Of Commissural S t i m u l a t i o n On P£r§I§K5^ Responses F i g , 7-6 shows the e f f e c t s of low i n t e n s i t y (3-5 V) c o n d i t i o n i n g COMM v o l l e y on t h e a m p l i t u d e of the p o p u l a t i o n s p i k e and r a t e of r i s e of the EPSP evoked by a t e s t PP v o l l e y . Whereas t h e amplitude and r i s e time of the t e s t EPSP were not a l t e r e d , the t e s t p o p u l a t i o n s p i k e was markedly p o t e n t i a t e d a t l o n g C-T i n t e r v a l s (20-400 msec) and depressed a t s h o r t e r i n t e r v a l s (5-20 msec), As shown i n t a b l e I I , maximum p o t e n t i a t i o n of th e PP-evoked p o p u l a t i o n s p i k e (290±33%) was observed 50-70 msec a f t e r c o n d i t i o n i n g s t i m u l a t i o n of t h e COMM pathway. I t i s noteworthy t h a t p o t e n t i a t i o n of the p o p u l a t i o n s p i k e was observed i n e v e r y a n i m a l (n=9) as l o n g as t h e a m p l i t u d e o f t h e p o p u l a t i o n s p i k e evoked by a c o n t r o l PP v o l l e y was submaximal. I f t h e PP was s t i m u l a t e d w i t h h i g h i n t e n s i t y (20-30 V) t h e p o p u l a t i o n s p i k e was not markedly p o t e n t i a t e d ( i . e. L e s s than 125% of c o n t r o l ) by a p r e c e d i n g COMM p u l s e . The c o n d i t i o n i n g s t i m u l u s i n t e n s i t y was not as c r i t i c a l s i n c e 287a Leaf 287 continued on 294. 288 E l S t , lz 6.1 E f f e c t s Of Cpjmmissjar^l S t i m u l a t i o a On Dentate f i e l d Responses J y o k e d By A T e s t P e r f o r a n t P a t h V o l l e y ^ A i a m p l i t u d e of the p o p u l a t i o n s p i k e and r a t e of r i s e o f EPSP evoked by a t e s t PP v o l l e y preceded by c o n d i t i o n i n g s t i m u l a t i o n o f COMM pathway (lower graph) o r t h e PP i t s e l f (upper graph). Note the h e t e r o s y n a p t i c p o t e n t i a t i o n o f the p o p u l a t i o n s p i k e but not t h e EPSP. J i averages o f 20 p o p u l a t i o n s p i k e r e sponses evoked by s t i m u l a t i n g t h e p e r f o r a n t path (arrows) a l o n e and f o l l o w i n g c o n d i t i o n i n g s t i m u l a t i o n o f t h e p e r f o r a n t path i t s e l f o r commi s s u r a l pathway. C: EPSP evoked by s t i m u l a t i n g PP al o n e ( t r a c e #2) and f o l l o w i n g c o n d i t i o n i n g s t i m u l a t i o n o f PP ( t r a c e #3) or COMM ( t r a c e #1) a t the same C-T i n t e r v a l (35 msec) as i n B showing t h a t t h e r a t e o f r i s e o f the EPSP i s a l t e r e d o n l y a f t e r double p u l s e s t i m u l a t i o n of t h e p e r f o r a n t path ( i . e . PP-PP). A. P P - P P 300 i J " l 1 1 1 1 1 1 1 1 1 -40 0 +40 120 200 280 C-T Interval (msec.) B. 2 msec 00 2?o TABLE II EFFECTS OF CONDITIONING STIMULATION OF THE COMMISSURAL PATHWAY ON PP-EVOKED POPULATION SPIKES E X P T . NO. amplitude of population spike* (percent of control) duration (mSec) SC-74 275 260 SC-75 135 120 SC-76 240 160 SC-77 260 180 SC-92 500 300 SC-115 350 320 SC-116 250 280 SC-133 280 300 SC-137 325 400 T O T A L = 9 MEAN = 290±33 MEAN = 258+29 *Each value represents the average of 20 t r i a l s at a condition-test i n t e r v a l of 35 mSec. 290a Leaves 291-293 do not exist. 294 potentiation of PP-evoked population spike was observed following a wide range (3-40 V) of commissural s t i m u l i . However conditioning volleys at i n t e n s i t i e s higher than 10 7 caused a small but s i g n i f i c a n t (110% of cont r o l , p<0.05) increase in the amplitude of the PP-evoked EPSP recorded i n 2 animals and prolonged both the i n i t i a l period of depression and subseguent potentiation of the PP-evoked population spike in a l l cases. In the 2 animals i n which a small increase in EPSP amplitude was observed the conditioning COMM stimulation evoked prominent l a t e responses. Thus the observed potentiation of the EPSP i n those two cases may be due to polysynaptic activation of the test PP pathway. Evidence That Potentiation O f T h e Population Spike Was JSfit Due To Changes In The Test Pathway. An increase in the amplitude of the PP-evoked population spike could be caused by activation of the i p s i l a t e r a l entorhinal cortex by the conditioning commissural volley. This prior activation of the EC would resu l t i n homosynaptic potentiation of the PP-evoked responses. Although the f a i l u r e to observe potentiation of the PP-evoked EPSP does not support t h i s p o s s i b i l i t y , further precaution was taken by sectioning the outflow of the EC v i a the PP i n 2 animals. The experimental paradigm consisted of stimulating EC, the PP i t s e l f and COMM pathways while recording from the dentate. When the electrodes were Leaf 294 continued on 298. Leaves 295-297 do not exist. 298 p o s i t i o n e d so t h a t s t i m u l a t i o n o f EC evoked a f i b r e v o l l e y i n the PP and a p o p u l a t i o n s p i k e i n the d e n t a t e a s t e r e o t a x i c k n i f e c u t was made between the EC and PP. F o l l o w i n g the k n i f e c u t s t i m u l a t i o n o f EC no l o n g e r evoked a d e n t a t e r e s p o n s e , but PP s t i m u l a t i o n p r o x i m a l t o t h e c u t r e s u l t e d i n a prominent p o p u l a t i o n s p i k e . C ommissural c o n d i t i o n i n g v o l l e y s c o n t i n u e d t o p o t e n t i a t e t h e PP-evoked p o p u l a t i o n s p i k e s u g g e s t i n g t h a t the EC d i d n ot mediate the observed p o t e n t i a t i o n . 7_j_4 D i s c u s s i o n F i e l d A n a l y s i s Of The C o n i s s u r a l I n g u t T o The- Dentate A) M o l e c u l a r l a y e r : S t i m u l a t i o n o f t h e COMM pathway r e s u l t s i n a s h o r t l a t e n c y e x t r a c e l l u l a r n e g a t i v e f i e l d which i s maximal i n the i n n e r m o l e c u l a r l a y e r . The l a m i n a r p r o f i l e s u g g e s t s t h a t t h e zone from which n e g a t i v e f i e l d s were r e c o r d e d extends 50-100 urn above t h e G - c e l l l a y e r and c o r r e s p o n d s t o the a n a t o m i c a l l y v e r i f i e d t e r m i n a l a r e a s o f t h e COMM pathway ( B l a c k s t a d , 1956; G o t t l i e b and Cowan, 1973). T h i s r e g i o n o f t h e m o l e c u l a r l a y e r , which c o n t a i n s p r i n c i p a l l y t h e d e n d r i t i c s h a f t s of G - c e l l s , i s below the PP synapses on t h e d i s t a l d e n d r i t e s of G - e e l l s . I n a d d i t i o n t o th e s e s t r u c t u r e s , t h i s l a y e r c o n t a i n s t h e d e n d r i t e s and axons o f c e l l s i n the pol y m o r p h i c l a y e r . U n l i k e t h e n e g a t i v e d e n d r i t i c wave which i s evoked 299 by PP stimulation, the COMM negativity has not yet been related to in t r a c e l l u l a r l y recorded EPSP^ of ind i v i d u a l G - c e l l s (Lomo, 1971a). This makes i t d i f f i c u l t to determine precisely which postsynaptic elements are mediating the COMM evoked responses. However, the laminar p r o f i l e s recorded in the present and previous experiments (Deadwyler et a l , 1975) suggest that the short latency (4-7 msec) responses result from a direc t action of the COMM pathway on dentate G-cells since 1) the COMM negative f i e l d i s produced monosynaptically and i s preceded by a presynaptic volley, 2) as in the case of the PP input, the negative dipole produced by COMM stimulation i s accompanied by a positi v e wave recorded i n G - c e l l layer which may r e f l e c t a passive source for the active sink at the COMM synapses and 3) individual G-cells are sometimes evoked on the positive c e l l layer f i e l d . B) Responses recorded in G-cell layer: In the region of G-cell body layer COMM stimulation produces a positive waveform upon which i n d i v i d u a l spikes are superimposed. Increasing the stimulating voltage (from 2-20 V) increased the size of t h i s positive waveform, although i t was always smaller than the PP evoked positive f i e l d . High stimulating voltages (15-20 V) were reguired to evoke a f i e l d with an amplitude comparable to that e l e c i t e d by PP stimulation (3-5 V). In 4 of 15 experiments a compound spike s i m i l a r to the PP-evoked population spike was superimposed on the 300 p o s i t i v e waveform. Deadwyler e t a l (1975) were not a b l e t o e l i c i t p o p u l a t i o n s p i k e s a t a l l f o l l o w i n g s t i m u l a t i o n o f t h e c o n t r a l a t e r a l CA3. These d i f f e r e n c e s may be r e l a t e d t o the f a c t t h a t Deadwyler et a l (1975) used f i n e monopolar e l e c t r o d e s t o s t i m u l a t e the c e l l s o f o r i g i n of t h e COMM pathway which may not be as e f f e c t i v e as a c t i v a t i n g t h e pathway d i r e c t l y w i t h b i p o l a r e l e c t r o d e s . The number o f s i n g l e u n i t s evoked i n t h e G - c e l l l a y e r was always lower f o l l o w i n g COMM s t i m u l a t i o n than PP s t i m u l a t i o n . I n c o n t r a s t t o t h e low p r o p o r t i o n of de n t a t e u n i t s which a r e a c t i v a t e d , over 90% o f sp o n t a n e o u s l y f i r i n g G - c e l l s a r e i n h i b i t e d f o l l o w i n g COMM s t i m u l a t i o n , independent of whether the same u n i t s were a c t i v a t e d or not. T h i s i n h i b i t i o n may be mediated by c o l l a t e r a l a c t i v a t i o n of b a s k e t - t y p e i n t e r n e u r o n e s which have l o n g axons p r o j e c t i n g towards G - c e l l body l a y e r ( C a j a l , 1911; Andersen e t a l , 1964) . S i n c e t h e i n h i b i t i o n produced by COMM s t i m u l a t i o n i s 2-3 tim e s l o n g e r t h a n t h e i n h i b i t i o n observed f o l l o w i n g PP s t i m u l a t i o n a t th e same r e l a t i v e i n t e n s i t y (1.5 X T) i t becomes n e c e s s a r y t o p o s t u l a t e t h a t more i n h i b i t o r y i n t e r n e u r o n e s are a c t i v a t e d f o l l o w i n g COMM s t i m u l a t i o n . These i n t e r n e u r o n e s may be t h e h i l a r neurones h a v i n g d e n d r i t i c a r b o r i z a t i o n e x t e n d i n g t o the i n n e r m o l e c u l a r l a y e r i n the r e g i o n of t h e COMM t e r m i n a l s ( G o t t l i e b and Cowan, 1973; Hj o r t h - S i m o n s e n 301 and Laurberg, 1978). The observations that a substantial proportion of COMM terminals are found i n the h i l a r region (Gottlieb and Cowan, 1973; Hjorth-Simonsen and Laurberg, 1978) supports the suggestion that the COMM input may be on inhibitory interneurones in addition to that postulated on G-cells. This postulated feed-forward i n h i b i t i o n of G-cells by COMM fi b r e s would explain the potent i n h i b i t i o n without concomitant activation observed i n most cases following COMM stimulation. Since the axons of in h i b i t o r y interneurones ramify extensively within the G-ce l l layer only a few d i r e c t connections with those neurones are reguired to produce the prolonged i n h i b i t i o n observed i n t h i s and Deadwyler et al»s (1975) study. Comparison With The Termination Of The Perforant Path As described i n t h i s and the previous chapter, PP and COMM f i b r e s terminate upon the outer and inner molecular layers of the dentate, respectively. The delineation between the terminal areas of PP and COMM pathways i s clear with minimal overlap between the two zones (Lynch and Cotman, 1975). This delineation i s also apparent from the present f i e l d analysis. The functional significance of these adjacent terminal f i e l d s i s not yet cl e a r , however, they may underlie the differences in the amplitudes of f i e l d 302 responses and number of G-cells evoked. , The COMM ext r a c e l l u l a r negative f i e l d has a slower r i s e time and smaller amplitude than that evoked by the PP. This may be due to either differences i n the effectiveness of PP and COMM terminals to generate synaptic currents or to a greater number of PP synapses in the outer molecular layer. The l a t t e r i s more l i k e l y since the outer molecular layer has a large number of en passage synapses with dendritic spines which degenerate following PP lesions (Nafstad, 1967) . These synapses on dendritic spines would be expected to generate more concentrated synaptic current than COHM synapses on the main dendritic shafts. These differences i n synaptic arrangement may also underlie the greater effectiveness of the PP input i n discharging G-cells and producing population spikes. Perhaps the physiological role of the COMM system i s to a l t e r the effectiveness of the more d i s t a l PP input rather than to d i r e c t l y activate G-cells. 303 E f f e c t Of Commissural S t i m u l a t i o n On PP Evoked-Responses S i n g l e p u l s e s t i m u l a t i o n o f COMM pathway r e s u l t e d i n a marked i n c r e a s e i n t h e a m p l i t u d e o f the PP-evoked p o p u l a t i o n s p i k e w i t h o u t a l t e r i n g the a m p l i t u d e or r a t e of r i s e o f t h e e x t r a c e l l u l a r EPSP, In the same a n i m a l s i n which c o n d i t i o n i n g COMM s t i m u l a t i o n d i d not a l t e r t h e s i z e o f t h e EPSP produced by a t e s t PP v o l l e y , p a i r e d p u l s e s t i m u l a t i o n o f the PP i t s e l f r e s u l t e d i n a r e l i a b l e p o t e n t i a t i o n o f t h i s r e s p o n s e . The l a t t e r o b s e r v a t i o n i s i n agreement w i t h the f i n d i n g s o f Steward et a l (1978) who a l s o f a i l e d t o f i n d p o t e n t i a t i o n of the PP-evoked EPSP f o l l o w i n g c o n d i t i o n i n g s t i m u l a t i o n o f t h e COMM a f f e r e n t . The a d d i t i o n a l o b s e r v a t i o n o f Steward et a l (1978) t h a t p a i r e d p u l s e s t i m u l a t i o n o f e i t h e r t h e PP, COMM or a s s o c i a t i o n a l pathways r e s u l t i n homosynaptic p o t e n t i a t i o n of a t e s t EPSP s u g g e s t s t h a t p o t e n t i a t i o n o f s y n a p t i c c u r r e n t can on l y be produced by p r i o r s t i m u l a t i o n of t h e t e s t pathway i t s e l f . A s i m i l a r c o n c l u s i o n was r e a c h e d by Andersen et a l (1978) and Lynch e t a l (1978) who observed t h a t t e t a n i c s t i m u l a t i o n o f an a f f e r e n t t o one d e n d r i t i c zone o f a CA1 p y r a m i d a l c e l l r e s u l t s i n l o n g l a s t i n g p o t e n t i a t i o n o f t h e same a f f e r e n t b u t not a f f e r e n t s p r o j e c t i n g t o a d j a c e n t d e n d r i t i c l a y e r s . I n c o n t r a s t t o the EPSP, the a m p l i t u d e o f t h e PP-304 evoked p o p u l a t i o n s p i k e was markedly enhanced f o l l o w i n g c o n d i t i o n i n g COMM s t i m u l a t i o n . Thus u n l i k e p o t e n t i a t i o n of the EPSP tha t o f the p o p u l a t i o n s p i k e i s not r e s t r i c t e d t o c o n d i t i o n i n g s t i m u l a t i o n o f the t e s t pathway. The observed p o t e n t i a t i o n of the p o p u l a t i o n s p i k e i s dependent on the COMM pathway s i n c e i t was not seen f o l l o w i n g acute or c h r o n i c t r a n s e c t i o n of the v e n t r a l p s a l t e r i u m , Fur thermore , p o t e n t i a t i o n of the p o p u l a t i o n s p i k e by COMM s t i m u l a t i o n was not e l i m i n a t e d by s u r g i c a l t r a n s e c t i o n between the e n t o r h i n a l c o r t e x and the PP thereby e l i m i n a t i n g the invo lvement of the c e l l s of o r i g i n of the PP i n p u t . The o b s e r v a t i o n tha t i n some animals p o t e n t i a t i o n of the PP-evoked p o p u l a t i o n sp i ke was observed w i t h i n 10 msec f o l l o w i n g a COMM c o n d i t i o n i n g v o l l e y s e v e r e l y r e s t r i c t s the invo lvement of p o l y s y n a p t i c ex t rah ippocampa l systems. In any case t h e absence o f changes i n the r a t e of r i s e of the EPSP when the popu la t ion sp i ke i s markedly p o t e n t i a t e d argues aga ins t the p o s s i b i l i t y t h a t the h e t e r o s y n a p t i c p o t e n t i a t i o n i s mediated by p r e s y n a p t i c events l e a d i n g to i n c r e a s e d PP t r a n s m i t t e r r e l e a s e . 305 7__5 Summary I n summary t h e r e s u l t s o f the p r e s e n t c h a p t e r demonstrate t h a t s t i m u l a t i o n of the COMM pathway r e s u l t s i n s h o r t l a t e n c y f i e l d r e s p o n s e s i n the d e n t a t e gyrus. The l a m i n a r p r o f i l e and i t s r e l a t i o n s h i p t o the depth p r o f i l e f o l l o w i n g s t i m u l a t i o n of PP s u g g e s t s t h a t t h e COMM i n p u t t o the d e n t a t e d e p o l a r i z e s the d e n d r i t e s of G - c e l l s at a p o s i t i o n more p r o x i m a l t o th e c e l l body t h a n the d i s t a l l y l o c a t e d PP synapses. The response o f sp o n t a n e o u s l y f i r i n g G - c e l l s f o l l o w i n g CO MM s t i m u l a t i o n causes a c t i v a t i o n - i n h i b i t i o n seguences i n a few cas e s and pure i n h i b i t i o n i n t h e remainder. S i n g l e p u l s e s t i m u l a t i o n of the COMM pathway r e s u l t s i n marked p o t e n t i a t i o n o f t h e PP-evoked p o p u l a t i o n s p i k e w i t h o u t a l t e r i n g the EPSP produced by a t e s t PP v o l l e y . On t h e b a s i s o f t h e s e d a t a , h e t e r o s y n a p t i c p o t e n t i a t i o n o f the p o p u l a t i o n s p i k e may be an a d d i t i o n a l f e a t u r e o f n e u r o n a l t r a n s m i s s i o n t h r o u g h t h e d e n t a t e gyrus. 306 C H A P I H 8: THE HAPHE^SEBOTONIN AND NEURONAL TRANSMISSION IN THE DENTATE GYROS 8 j_1 Introduction The previous two chapters described the laminar p r o f i l e of the perforant path (PP) input to the dentate and the f e a s a b i l i t y of using the rate of r i s e of the ex t r a c e l l u l a r EPSP and amplitude of the population spike as indicators of synaptic current and synchronous neuronal discharges, respectively. The observation made in Chapter 7 that conditioning stimulation of the commissural pathway increases the amplitude of the population spike without a l t e r i n g the rate of r i s e of the EPSP prompted an examination of the ef f e c t s of stimulating known monoamine projections to the dentate. Unlike the commissural input the monoamine systems do not substantially innervate the molecular layer of the dentate and are thus unlikely to a f f e c t the PP synapses on the d i s t a l dendrites of granule c e l l s . The present chapter examines the eff e c t s of stimulating the median raphe nucleus (MR), the s i t e of o r i g i n of hippocampal serotonin on f i e l d potentials evoked in the dentate by test PP voll e y s . Whenever possible the response of single dentata units i s also examined i n order to r e l a t e observed changes i n f i e l d potentials to the a c t i v i t y of i n d i v i d u a l neurones recorded from the same s i t e . 307 8.2 Experimental Procedures The methods used were similar to those described in chapter 7, Bipolar stimulating electrodes were st e r e o t a x i c a l l y positioned i n the angular bundle to activate the perforant path and in the mossy f i b r e s or i p s i l a t e r a l CA3 (AP = -3.0 mm; L = +3.5 mm; V =-3.5 mm) to activate granule c e l l s antidromically. ,An a d d i t i n a l stimulating electrode was positioned i n the median raphe nucleus (AP = -8.0 mm; L = 0.0 mm; V = -6.0 mm to -6 ,5 mm). The experiments were performed on 15 normal rats and 5 rats pretreated with para-chlorophenylalanine (p-CPA, 400 mg/kg; i . p.). , Immediately following the electrophysiological experiments, some animals (5 normals and 5 p-CPA-treated) were decapitated and the hippocampal formation on both sides removed f o r subseguent 5-HT assay described i n chapter 2. 308 8JJ_3 Results Effects Of MR Stimulation On G-cells The response of 66 spontaneously f i r i n g G-cells was studied following stimulation i n the region of the median raphe nucleus. Dentate G-cells were i d e n t i f i e d on the basis of t h e i r response patterns following PP stimulation and their r e l a t i o n s h i p to PP-evoked f i e l d responses (Fig, 8-1). In some cases (n=15) c e l l s were further i d e n t i f i e d by antidromically activating them by mossy f i b r e stimulation. As shown i n Fig, 8-1D, single pulse stimulation (10-20V) of the MR inh i b i t e d a t o t a l of 56 (85%) G-c e l l s at latencies ranging between 10-25 msec (See also Fig, 8-2), The duration of i n h i b i t i o n lasted for periods of 25-175 msec (mode=43.0 msec)... The duration of inhibiton could be lengthened by stimulating the MR with higher i n t e n s i t y or increasing the number of pulses (Fig. 8-2). H i s t o l o g i c a l v e r i f i c a t i o n of e f f e c t i v e stimulating electrode placements indicated a d i r e c t r e l a t i o n s h i p between the length of i n h i b i t i o n and proximity of the electrode to the MR (Fig. 8-3). In addition stimulating s i t e s 1.0-2.0 mm posterior to the MB did not produce the observed i n h i b i t i o n i n d i c a t i n g a s p e c i f i c action of t h i s p a r t i c u l a r nucleus on G-cells. 309 I I S x 8- Xl E f f e c t s • Of MR S t i m u l a t i o n On Dentate Granule C,gll§» A i arrangement o f r e c o r d i n g e l e c t r o d e a t the l e v e l o f t h e c e l l body and s t i m u l a t i n g e l e c t r o d e s i n e x c i t a t o r y (open t r i a n g l e s ) and i n h i b i t o r y ( c l o s e d t r i a n g l e s ) a f f e r e n t s from the e n t o r h i n a l c o r t e x {EC) and median r a p h e n u c l e u s (MR), r e s p e c t i v e l y . C o l l a t e r a l of t h e G - c e l l axon, the mossy f i b r e (MF), a c t i v a t e s an i n h i b i t o r y i n t e r n e u r o n . Bx o s c i l l o s c o p e t r a c e s of the waveforms evoked by PP s t i m u l a t i o n evoked a t v a r i o u s i n t e n s i t y i n d i c a t i n g t h a t r e c o r d i n g s were from t h e G - c e l l l a y e r . C^p: p o s t s t i m u l u s h i s t o g r a m s ( b i n w i d t h = 5 msec) of a G - c e l l d i s c h a r g e f o l l o w i n g s i n g l e p u l s e s t i m u l a t i o n o f the PP (C) and MR (0). I n s e r t s show 25 superimposed o s c i l l o s c o p e sweeps. A r r o w s i n d i c a t e s t i m u l u s a r t i f a c t s and c a l i b r a t i o n s a r e 5 msec/1.0 mV i n B and 50 msec/. 1 mV i n C and D. I -Neurone - -'—F=»—«' B 5 V 10 V c. 50 n a. t • j D 50 , . ! . I . I i I ' I ' I ' . I ' I ' I ' 1 ' I ' I 1 I 1 I 1 I ' I ' I * _ „ . 750 0 A — r 1 0 0 0 311 FIG, 8- 2j_ • Inhibition Of Spgntaneously F i r i n g Dentate Cel l s By MB S^jfulatipnjs. A i e f f e c t s of increasing the stimulus i n t e n s i t y on the length of i n h i b i t i o n observed following MB stimulation, (10 superimposed oscilloscope sweeps) , .• B_j, effects of increasing the number of pulses delivered to the MB on the length of the observed i n h i b i t i o n . In the lower records the two pulses were repeated by 10 msec. (20 superimposed oscilloscope sweeps). Cj, PST histogram of the response of a G-cell to MB stimulation with single or r e p e t i t i v e (300 Hz) pulses. Bin width i s egual to 10 msec; 50 t r i a l s in each case. B. 313 Effects Of MR Stimulation On Pg-§voked Responses The response of 12 spontaneously f i r i n g G-cells which were excited by the perforant path were examined following stimulation of the MR. As shown i n Fig. 8-4, single pulse stimulation of MR, which inh i b i t e d the spontaneous discharge of a l l 12 c e l l s , also blocked the single spike activation of 8 c e l l s (6 6%) following stimulation of the PP at threshold i n t e n s i t y . However, stimulation of the MR did not r e l i a b l y block the activation produced by stimulating the perforant path at 2 X T. When the PP was stimulated with 3 x T i n t e n s i t y , thereby evoking a number of G-cells (Fig. 8-4C) , MR stimulation resulted in a more synchronous discharge of G - c a l l s which resembled the population spike response. Thus, i t became impossible to determine whether i n d i v i d u a l c e l l s which were evoked i n response to control PP stimulation were also evoked by the test PP volley since they may be masked by compound spikes such as that shown in Fig. 8-4C. In order to overcome this problem, the amplitude of the PP-evoked population spike was measured before and a f t e r stimulation of the MR. As shown in F i g . 8-5, a single conditioning volley to MR resulted in an increase in the amplitude of the PP-evoked population spike at c r i t i c a l i n t e r v a l s between 20-70 msec. The r i s e time of the PP-evoked EPSP was not altered by conditioning pulses that potentiated the amplitude of 314 D L G A , 8- 3z Belatignship. Between In h i b i t i o n 0£ Dentate C e l l s And Position Of The Stimulating Electrode In The Region Of hEx Ai schematic of a coronal section of the rat brain at the l e v e l of MR (A160) indicating the position of the stimulating electrode along a v e r t i c a l track which produced the corresponding responses shown i n B. Bi the response of a spontaneously f i r i n g dentate unit following stimulation i n the region of MR at the indicated positions. Each record consists of 10 superimposed oscilloscope sweeps.Abbreviations as on page 130. 3\5 316 FIG.. 8- j4z_ E f f e c t s Of MR Stimulation On PP Evoked Activation Of Dentate Cellsj, Ai spontaneously f i r i n g dentate units which were i n h i b i t e d by single pulse stimulation of MR (top) and activated by PP (bottom)., B- indicates that the PP-evoked spike (top) i s eliminated by a preceding pulse to the MR (bottom) . PP stimulus i n t e n s i t y at threshold and C-T i n t e r v a l i s 30 mSec., Single spike activation of the above unit., Note the elimination of the PP evoked spike i n the lower sweep. . Ci same experiment as in B except the PP i s stimulated with s u f f i c i e n t i n t e n s i t y to evoke a number of neurones. Note the appearance of a compound spike following MR conditioning stimulation i n the lower sweep. , {_ [ 0.2 mV 2 msec PP MR-PP t • T 0.2 mV 5 msec PP MR-PP T 0.2 mV 5 msec 3 1 8 8- 5:, E f f e c t s Of MS Conditionina Pulses OB The F i e l d Responses Evoked By The Perforant Path A kz_ graphs of the amplitude of population spike and rate of r i s e of the EPSP evoked by a PP volley (7.5 V) when preceded by a conditioning pulse (15 V) to PP (top graph) or to the MB (lower graph). C: oscilloscope traces of control PP-evoked population s p i k e s ( l e f t ) and following conditioning PP (top right) and MR (lower right) pulses. 320 t h e p o p u l a t i o n s p i k e r e c o r d e d i n t h e same a n i m a l s . P o t e n t i a t i o n of the p o p u l a t i o n s p i k e was obser v e d i n ev e r y animal t e s t e d w i t h t h e maximum p o t e n t i a t i o n r a n g i n g between 110% and 170% o f c o n t r o l . The degree o f p o t e n t i a t i o n was r e l a t e d t o t h e d i s t a n c e between t h e s t i m u l a t i n g e l e c t r o d e and t h e c e n t e r of the MR, C o n d i t i o n i n g s t i m u l a t i o n i n t h e DR and a d j a c e n t a r e a s d i d not r e s u l t i n r e l i a b l e p o t e n t i a t i o n . I t i s noteworthy t h a t a t the s t i m u l u s i n t e n s i t i e s used (10-20V) MR s t i m u l a t i o n d i d not r e s u l t i n s i g n i f i c a n t d e p r e s s i o n of the p o p u l a t i o n s p i k e s i m i l a r to t h a t which i n i t i a l l y f o l l o w s c o n d i t i o n i n g s t i m u l a t i o n of t h e PP i t s e l f . I S l a t i o n s h i p Between E f f e c t s Of MR S t i m u l a t i o n - - On-Eopjulatign S p i k e And I n h i b i t i g n Of S i n g l e Units.. : As i l l u s t r a t e d i n F i g . 8-6, the time c o u r s e o f t h e f a c i l i t a t i o n evoked by t h e MR c o n d i t i o n i n g p u l s e s c o i n c i d e d w i t h the p e r i o d of i n h i b i t i o n o f G - c e l l s r e c o r d e d at t h e same p o s i t i o n or i n t h e immediate v i c i n i t y . When the d u r a t i o n o f the i n h i b i t o r y component was a l t e r e d by changing i n t e n s i t i e s o f s t i m u l a t i o n , the p e r i o d of f a c i l i t a t i o n of the p o p u l a t i o n s p i k e was a l t e r e d i n t h e same d i r e c t i o n . I n F i g . 8-7, t h i s r e l a t i o n s h i p i s f u r t h e r i l l u s t r a t e d by p l o t t i n g t h e a m p l i t u d e of a r e p r e s e n t a t i v e p o p u l a t i o n s p i k e and the p r o b a b i l i t y t h a t a g i v e n G - c e l l i s i n h i b i t e d a t v a r i o u s 321 | I G i ; 8; 6: F a c i l i t a t i o n Of The Population Spike Response Following PP And MS Conditioning Pulses The time course of popoulation spike potentiation (upper graphs) following PP (A) and MR (B) conditioning pulses i s compared with the response of a spontaneously f i r i n g dentate neurone (PST histograms, bin width = 5 mSec) ) recorded i n the v i c i n i t y of the population spike..Note that the duration of potentiation i s related to the length of the i n h i b i t i o n e l i c i t e d by the conditioning pulse i n each case. 323 times following MB stimulation. The probability was computed by counting the number of c e l l s (n=66) which were i n h i b i t e d during 5,0 msec sample bins following MS stimulation. It i s noteworthy that the time period when the majority of G-cells were i n h i b i t e d by MR stimulation i s the optimal C-T i n t e r v a l (30-35 msec) for maximal f a c i l i t a t i o n of the population spike. E f f e c t s of P_;CPA On Bespones Ivoked JEjy. Stimulation Of MR As shown in table 3, p-CPA pretreatment resulted i n a s i g n i f i c a n t depletion (69%) of hippocampal 5-HT measured by the spectrofluorometric assay described i n chapter 2. The pretreatment also resulted i n a s i g n i f i c a n t attenuation of MR-evoked i n h i b i t i o n of single G-cells and enhancement of the PP population spike response. Moreover, the degree of attenuation of both the i n h i b i t i o n (61%) and population spike enhancement (80%) were s i m i l i a r to the depletion of hippocampal 5-HT. 324 ZLQ.*. 8~ ll Relationship Between Inhibi£ion Of G-cglls And Amplitu.de Of The Population Spijcej.-The p r o b a b i l i t y of i n h i b i t i o n ( c e l l s i n h i b i t e d / c e l l s tested X 100) shown in the histogram and the amplitude of a representative population spike (dashed line) are the l e f t and right ordinates, respectively. The MB was stimulated at time zero and the abscissa represents 5 msec sampling periods of neuronal (n=66) discharges., 8.0 / \ o 100 -0 ^ -c o 8 0 -15 6 0 -1c c m o 4 0 -lity 2 0 -r obabi 0 -Q_ T " 60 | i i — i — | — i — • — 1 — | • ^ 0 20 40 Time after MR Stimulation (msec) h-6.0 h-4.0 h 2 . 0 > £ Q. CO Q. O 0_ •a CL £ < L - 0 .0 "1 80 3ZG TABLE III EFFECTS OF p-CPA ON HIPPOCAMPAL 5-HT AND RESPONSES TO STIMULATION OF THE MEDIAN RAPHE pp-evoked population spike (% increase) Control 40±5 (n=15) 55 (83%) 545±29 (n=5) 66 p-CPA 8±3 (n=4) 8 (32%) 166±18 (n=4) 25 percent difference -80* -61* -69** p-CPA (400 mg/Kg, i.p .) was administered 3 days before recording experiments. 5-HT levels were assayed i n both sides of the hippocampal formation using spectrofluoremetric techniques described i n chapter 2 (*p<0.01; **p<0.001). i n h i b i t i o n 5-HT con-of G-cells centration (no. i n h i b i t e d (ng/g wet no. tested) tissue) 327 8 .J. D i s c u s s i o n E f f e c t s Of MR S t i m u l a t i o n O n S p o n t a n e o u s Discharge. Of G - c e l l s S i n g l e p u l s e s t i m u l a t i o n i n t h e r e g i o n o f the MB r e s u l t s i n s h o r t l a t e n c y (10-25 msec) i n h i b i t i o n o f t h e m a j o r i t y o f s p o n t a n e o u s l y f i r i n g G - c e l l s . . T h e d i r e c t r e l a t i o n s h i p between the l e n g t h of i n h i b i t i o n and p r o x i m i t y o f t h e s t i m u l a t i n g e l e c t r o d e t o the MB s u g g e s t s t h a t t h e i n h i b i t i o n i s mediated by t h i s p a r t i c u l a r n u c l e u s and not t h e a d j a c e n t a r e a s . I t i s not p o s s i b l e from t h e p r e s e n t data t o d i s t i n g u i s h between mono- and p o l y s y n a p t i c MR evoked i n h i b i t i o n of G - c e l l s . The o b s e r v a t i o n t h a t i n h i b i t i o n o c c u r r e d at s h o r t l a t e n c i e s and i n some c a s e s i m m e d i a t e l y a f t e r the s t i m u l u s s u p p o r t s a d i r e c t i n p u t onto G - c e l l s r a t h e r t h a n an i n d i r e c t a c t i o n i n v o l v i n g e x t r a h i p p o c a m p a l systems. W i t h i n the d e n t a t e the d i s t r i b u t i o n o f 5-HT t e r m i n a l s . i s p r i m a r i l y t o t h e r e g i o n of the d e n t a t e h i l u s . H i s t o f l u o r e s c e n c e and a u t o r a d i o g r a p h i c s t u d i e s ( H a l a r i s e t a l , 1976, see a l s o s e c t i o n 1.1.1) r e v e a l a c l e a r i n f r a g r a n u l a r band o f t e r m i n a l s which may be e i t h e r onto i n t e r n e u r o n e s o r t h e G - c e l l b o d i e s . S i n c e some o f t h e i n h i b i t e d neurones were i d e n t i f i e d as G-c e l l s by a n t i d r o m i c a l l y f i r i n g them f o l l o w i n g mossy f i b r e s t i m u l a t i o n t h e s e neurones may r e c e i v e a d i r e c t s e r o t o n e r g i c i n p u t o r i g i n a t i n g i n MR. 328 Effects Of MS Stimulation On PP;ey ok ed Responses Single pulse stimulation of the MR blocked the ac t i v a t i o n of G-cells evoked by threshold but not suprathreshold stimulation of the PP. This suggests that at higher PP i n t e n s i t i e s the a c t i v a t i o n can overcome the raphe mediated i n h i b i t i o n onto G-cells. Thus when the PP was stimulated at i n t e n s i t i e s s u f f i c i e n t to evoke a population spike, the amplitude of the population spike was not reduced by perforant path stimulation. In fact an enhancement of the population spike response was observed following MR stimulation. Since potentiation of the population spike was not accompanied by a concomitant increase i n the rate of r i s e or amplitude of the EPSP i t was not due to the action of MR on the entorhinal cortex or the region of the PP synapse on d i s t a l dendrites. Thus i t i s not necessary to postulate an increase i n transmitter release to account for potentiation of the population spike. Since conditioning stimulation of the commissural pathway (Chapter 2) and the mossy f i b r e s {Lomo, 1971b; Assaf and M i l l e r , unpublished results) potentiate the population spike without a l t e r i n g the EPSP e x t r i n s i c afferents to the dentate may a l t e r PP-dentate transmission without changing synaptic current. This point w i l l be developed further i n chapter 10., 329 BglgtionshJE Between I f i M M t i o n Of S h e l l s And Potentiation Of The Population. Spike The present data demonstrate a temporal rel a t i o n s h i p between MR evoked i n h i b i t i o n of G-cells and potentiation of the population spike response such that the onset of i n h i b i t i o n coincides with the c r i t i c a l C-T in t e r v a l s at which the population spike amplitude i s increased. Furthermore, the optimum C-T i n t e r v a l for producing maximum potentiation i s when the largest proportion of G-cells i s inhibited by MR stimulation. This r e l a t i o n s h i p between i n h i b i t i o n of G-c e l l discharge and potentiation of the population spike was also demonstrated during paired pulse stimulation of the PP i t s e l f . This suggests that i n h i b i t o r y mechanisms, i n part, may mediate the observed potentiation of the population spike without a l t e r i n g synaptic current. Sole For Serotonin: The i n h i b i t i o n of spontaneously f i r i n g G-cells and enhancement of the PP-evoked population spike observed following MR stimulation were subst a n t i a l l y reduced by p-CPA pretreatment which decreased 5-HT concentration in the hippocampal formation. Taken together with the relationship between the i n h i b i t i o n of G-cells and position of the stimulating electrode in the MR (Fig. 8-3), these data support the proposal that these 330 responses were mediated by a raphe-serotonin system. The observation that single pulse stimulation of MR enhances the amplitude of the PP-evoked population spike was recently confirmed by Winson (1978 and personal communication). He also observed that MR stimulation increases the amplitude of the population spike evoked by PP stimulation at C-T i n t e r v a l s ranging between 20 and 70 msec. However, Hinson (1978) examined the e f f e c t s of MR stimulation i n awake as well as in anaesthetized animals. Augmentation of the population spike response was observed i n slow wave sleep and in the anaesthetized state but not i n the a l e r t state (Winson, 1978). On the basis of these data, ainson (1978) suggested that the e f f e c t s of a raphe-serotonin system on neuronal transmission in the dentate gyrus may be behaviour-specific. 331 8^5 Summary In summary, single pulse stimulation of the MR resu l t s i n short latency i n h i b i t i o n of spontaneously f i r i n q G-cells and potentiation of the PP evoked population spike without a l t e r i n g the r i s e time of the EPSP. There i s a temporal relationship between the period of i n h i b i t i o n of unit a c t i v i t y and c r i t i c a l C-T in t e r v a l s for potentiation. Depletion of 5-HT in the hippocampal formation attenuates the MR-elicited respones suggesting that they are mediated by a serotonin-containing system. 332 CHAPTER 9j. THE LOCUS COERULEUS AND NEURONAL TRANSMISSION IN THE- DENTATE GJRUS I a t r g d u c t i o n C o n d i t i o n i n g s t i m u l a t i o n of t h e c o m m i s s u r a l pathway (Chapter 7) and t h e median raphe n u c l e u s (Chapter 8) i n c r e a s e t h e a m p l i t u d e of t h e p e r f o r a n t path (PP)-evoked p o p u l a t i o n s p i k e . L i k e the s e r o t o n i n i n n e r v a t i o n o f the d e n t a t e , n o r a d r e n a l i n e (NA)-c o n t a i n i n g t e r m i n a l s , which o r i g i n a t e i n t h e l o c u s c o e r u l e u s ( L C ) , are s p a r s e i n the zone of PP synapses but r e l a t i v e l y dense i n t h e d e n t a t e h i l u s . T h i s s u g g e s t s t h a t LC i s u n l i k e l y t o a f f e c t PP synapses or t h e EPSP d i r e c t l y but may a l t e r the d i s c h a r g e o f G-c e l l s at some o t h e r s i t e . The p r e s e n t c h a p t e r examines t h e e f f e c t s o f s t i m u l a t i n g LC on f i e l d p o t e n t i a l s evoked i n t h e d e n t a t e by a t e s t PP v o l l e y . The response of s i n g l e d e n t a t e c e l l s i s a l s o examined i n an attempt t o r e l a t e changes i n f i e l d p o t e n t i a l s t o a c t i v i t y of i n d i v i d u a l neurones. 333 3i2 Experimental Procedures The methods used were sim i l a r to those described i n chapters 7 and 8. In addition to a l l the electrode placements provided i n chapter 8 (PP, DG,CA3, MR), a bipolar stimulating electrode was positioned i n the locus coeruleus (AP = 1.5 mm posterior to stereotaxic ear bars; L = 1.1 mm from the midline; V = 5.5 to 6.5 mm below the surface of cerebellar cortex) of 15 normal rats and 10 rats which had previously (15-20 days) recieved a b i l a t e r a l i n j e c t i o n of 6-OHDA (4 ug in 2 u l of 0. 15M saline with 0.2 mg/ml ascorbic acid) into the dorsal NA bundle. Following the elect r o p h y s i o l o g i c a l experiments the hippocampal formation was removed and tissue samples assayed for NA as described in Chapter 2. 9f_3 Results Effects Of LC Stimulation On Dentate C e l l s The responses of 63 spontaneously f i r i n g dentate neurones were examined following single or multiple pulse stimulation of the locus coeruleus. Of the 63 c e l l s , 56 were i d e n t i f i e d as G-cells on the basis of thie r response to stimulation of PP as well as by h i s t o l o g i c a l l o c a l i z a t i o n of the recording electrode in the dentate c e l l body layer. In addition 12 of the 56 G-cells were activated antidromically by mossy f i b r e 334 2 z l L E f f e c t s Of LC Stimulation On. Dentate Neuronest Aj raster display showing the response of a spontaneously f i r i n g neurone recorded i n the G-c e l l layer following single pulse stimulation of LC (top) and MS (bottom), Response of the same neurone to single and multiple pulse (4 pulses, 300 Hz) stimulation of LC i s shown by the lower raster. BJL oscilloscope records showing a G-cell which was monosynaptically evoked by PP stimulation, antidromically evoked by mossy f i b r e (MP) stimulation. Single pulse stimulation of LC was without effect while multiple pulses resulted i n i n h i b i t i o n . pp l_Jo.l mV 5 msec • To.l mV 2 msec nimjmiiimiiiiiUttiHiij^. m V ^. LC 50 msec I Pulse t i e 4 Pulses M 0.2 mV 50 msec 336 s t i m u l a t i o n . 7 neurones c o u l d n o t be i d e n t i f i e d as G-c e l l s s i n c e t h e y were s y n a p t i c a l l y a c t i v a t e d by mossy f i b r e s t i m u l a t i o n and were r e c o r d e d below the G - c e l l l a y e r . The l a t t e r neurones were t e n t a t i v e l y i d e n t i f i e d as h i l a r neurones, p o s s i b l y i n t r i n s i c t o the d e n t a t e g y r u s . The d i s c h a r g e p a t t e r n of the m a j o r i t y of G - c e l l s (88%) was not a l t e r e d by s i n g l e p u l s e s t i m u l a t i o n of t h e LC a t i n t e n s i t i e s r a n g i n g from 0-25V ( F i g . 9-1). I n h i b i t i o n at s h o r t l a t e n c i e s ( l e s s t h a n 25 msec) was observed i n 4 (7%) u n i t s and s i n g l e s p i k e a c t i v a t i o n was observed i n 3 (5%) G - c e l l s . Only 1 of 7 non G - c e l l s was i n h i b i t e d . D u r i n g s t a b l e r e c o r d i n g s from G - c e l l s t h e s t i m u l a t i n g e l e c t r o d e was moved i n t h e d o r s a l - v e n t r a l p lane t o i n s u r e t h a t the response of the c e l l was ob s e r v e d at d i f f e r e n t s t i m u l a t i n g depths. At depths 1.0-1.5 mm below the l e v e l of the LC f a c i a l movements were evoked by t h e s t i m u l a t i o n , t h e s e movements caused a l a r g e f i e l d i m m e d i a t e l y f o l l o w i n g the s t i m u l u s a r t i f a c t and may be r e l a t e d t o a c t i v a t i o n o f the n u c l e u s o f the f i f t h c r a n i a l nerve. E l e c t r o d e s were p l a c e d i n both LC and MB i n 4 a n i m a l s a l l o w i n g a comparison between the res p o n s e s of 21 neurones f o l l o w i n g MB and LC s t i m u l a t i o n (e.g. F i g . 9-1, top) s i n g l e p u l s e s t i m u l a t i o n o f MB i n h i b i t e d t h e d i s c h a r g e o f 16 neurones whereas t h a t o f LC 337 inh i b i t e d only 2 of the neurones. The i n h i b i t i o n following MR stimulation was si m i l a r in latency (less than 25.0 msec) and duration (mode-45 msec) to that described in the previous chapter. The e f f e c t s of r e p e t i t i v e stimulation of the LC (2-5 pulses, 300 Hz) were examined on 15 G-cells recorded i n 5 animals. As shown by the lower raster i n Fi g . 9-1, stimulation of the LC with 4 pulses resulted i n r e l i a b l e i n h i b i t i o n in 8 (55%) neurones while single pulses had no ef f e c t . The onset of i n h i b i t i o n was less than 30 msec following the l a s t pulse of the t r a i n and continued for 50-100 mSec. The background discharge rate of the neurone was often (4 of the 6 neurones) higher during stimulus presentations than control periods. Slight f a c i a l movements were observed during stimulation, therefore the increased discharge rate of the neurone may be re l a t e d to the activation of sensory-motor nuclei adjacent to LC, 338 U J e c t Of LC Stimulation On PPz§.I2l£t<l JLi&l& Eg§B9M§§Sji. The effects of stimulating LC with sing l e and r e p e t i t i v e (2-5 pulses, 15V, 300 hZ.) pulses on amplitude of the population spike and r i s e time of the EPSP was examined in 6 control rats. As shown i n Fig. 9-2, r e p e t i t i v e but not single pulse stimulation of LC r e l i a b l y potentiated the population spike response i n a l l 6 rats but did not a l t e r the r i s e time of the EPSP in the same animals. The c r i t i c a l C-T i n t e r v a l s for potentiation of the population spike ranged between 20-80 msec following the l a s t pulse of the b rief t r a i n . Maximum potentiation (150-175% of control) was observed at optimal C-T i n t e r v a l s between 35 and 50 msec. Fig. 9-3 indicates that the degree of potentiation i s r elated to the number of stimulating pulses since sin g l e pulse did not r e l i a b l y r e s u l t in potentiation whereas 2-4 pulses produced r e l i a b l e potentiation. The response following a stimulus t r a i n of 5 pulses was not substantially different than that observed following 6 or more pulses, thus a 4-5 pulse stimulus t r a i n was adopted as the test stimulus for inter-animal comparisons. As shown i n Fig. 9-4, there was a direct re l a t i o n s h i p between depth of the stimulating electrode t i p and the degree of potentiation. Placements 339 0<LJ_ 2Z 2.L E f f e c t s Of LC Stimulation On Pgreygked Dentate Population EesponseSj. A i the e f f e c t s of conditioning LC stimulation (15V,5 pulses at 300 Hz.) on PP-evoked population spike and EPSP. Bz oscilloscope sweeps showing the control (lower arrow) PP-evoked population spike and potentiation of t h i s response (top arrow) by preceding stimulation of LC ( C-T = 35 mSec). Test Responses (96of Control) > O ZZ PO w * O O O O O O o —h m Tl 341 9- 3z_ Relationship Between The Number Of LC Pulses And Amplitude Of The Population Spike* Each point i s the average of 20 t r i a l s at a constant condition-test i n t e r v a l (35 msec) . Freguency of the pulse t r a i n was 300 Hz. , 342.1 FIG. 9- 4_L H i s t o l o g i c a l L o c a l i z a t i o n Of. Stimulation Electrode In LCi:,-The photomicrograph taken at the r o s t r a l l e v e l of LC shows the stimulating electrode placement (arrow). The lower schematic and corresponding p r o f i l e s indicate the relationship between proximity of the stimulating electrode to LC and enhancement of the PP-evoked population spike. Each record i s the average of 30 t r i a l s . Abbrevi atignSji. CEE cerebellum DTN dorsal tegmental nucleus LC locus coeruleus IVV fourth cerebral v e n t r i c l e 5 4 2 - 2 343 lo c a l i z e d to the locus coeruleus, e s p e c i a l l y the r o s t r a l pole, produced s i g n i f i c a n t l y more potentiation than extra LC placements. Effects Of 6-OHDA Lesjons Of-The Dorsal No|adreneraic Pathway B i l a t e r a l i n j e c t i o n of 6-OHDA (4 ug i n 2 ul saline) into the dorsal NA bundle resulted in substantial depletions (mean=91%) of hippocampal NA. As shown i n Fig. 9-5 e l e c t r i c a l stimulation of LC in 6-OHDA treated r a t s did not s i g n i f i c a n t l y potentiate the population spike response. In the same animals paired pulse stimulation of PP continued to produce a s i g n i f i c a n t potentiation indicating that the capacity for increase i n amplitude was not eliminated by 6-OHDA lesions. It i s noteworthy that the only lesioned animal in which a low degree ( 120- 125%) of potentiation was observed had the least e f f e c t i v e depletion of NA (30%). 344 ££<LL. 2Z §i Ilfects Of 6-OHDA On The Besnonse To LC Stimulation^ _A: amplitude of the test population spike plotted as a function of condition-test (C-T) in t e r v a l . The v e r t i c a l bars indicate standard errors of the mean. Bjj. oscilloscope traces of control (PP) and test (LC-PP) population spikes i n d i c a t i n g that spike amplitude i s increased following e l e c t r i c a l stimulation of LC in control (top photo) but not in 6-OHDA lesioned r a t (lower photo) . C-T (msec) LC-PP PP LC-PP 346 9 X4 D i s c u s s i o n E f f e c t s Of LC S t i m u l a t i o n On G - c e l l D i s c h a r g e In c o n t r a s t t o t h e i n h i b i t i o n o b s e r v e d f o l l o w i n g s i n g l e p u l s e s t i m u l a t i o n o f t h e MB, s i n g l e v o l l e y s d e l i v e r e d to the LC d i d not r e l i a b l y i n f l u e n c e t h e d i s c h a r g e o f t h e m a j o r i t y o f d e n t a t e c e l l s . T r a i n s of s t i m u l i were much more e f f e c t i v e r e s u l t i n g i n i n h i b i t i o n o f 55% o f t h e d e n t a t e c e l l s a t l a t e n c i e s c o n s i s t e n t l y l e s s than 30 msec f o l l o w i n g the onset o f t r a i n . However, s t i m u l a t i o n o f LC w i t h r e p e t i t i v e t r a i n s u s u a l l y r e s u l t e d i n f a c i a l movements and an i n c r e a s e i n t h e background f i r i n g of the r e c o r d e d neurone. P r e v i o u s s t u d i e s have a l s o i n d i c a t e d t h a t r e p e t i t i v e s t i m u l a t i o n of LC i s r e g u i r e d t o i n h i b i t h ippocampal ( S e g a l and Bloom, 1974) and c o r t i c a l neurones { P h i l l i s and K o s t o p o u l o s , 1976) . However t h e s e s t u d i e s do not r e p o r t whether t h e y observed f a c i a l t w i t c h e s . N e v e r t h e l e s s , t h e i r r e c o r d s ( S e g a l and Blocm, 1974b F i g . 2; P h i l l i s and K o s t o p o u l o s , 1976 F i g . 1) r e v e a l a h i g h e r background d i s c h a r g e of the neurone b e i n g r e c o r d a d . fin i n c r e a s e i n d i s c h a r g e r a t e may be r e l a t e d t o a c t i v a t i o n of a d j a c e n t sensory-motor r e g i o n s due t o c u r r e n t s p r e a d r e s u l t i n g from h i g h f r e q u e n c y s t i m u l a t i o n (Bagshaw and Evans, 1976). T h i s i n c r e a s e d r a t e o f d i s c h a r g e c o u l d o b s c u r e LC-evoked i n h i b i t i o n . The r e a s o n s t h a t s t i m u l a t i o n o f LC w i t h r e p e t i t i v e 347 s t i m u l i a t low i n t e n s i t i e s i s more e f f e c t i v e than a single strong volley may be due to the organization of the LC and i t s mode of termination upon the recorded c e l l s . Anatomical investigations have revealed that a r e l a t i v e l y small population of LC c e l l bodies (estimate range between 1400-2000) give r i s e to t h i n unmyelinated axons which bifurcate considerably to terminate in wide regions of the forebrain (Fuxe, 1965; Ongerstedt, 1971; Swanson and Hartman, 1975 ; Jones et a l , 1977 ). Perhaps r e p e t i t i v e stimulation overcomes the high pr o b a b i l i t y of conduction f a i l u r e inherent i n a large number of axon bifurcations (Hall, 1974). In the absence of acceptable reports on the discharge patterns of i d e n t i f i e d LC neurones no comparisons can be made at th i s time between optimal stimulation parameters and the spontaneous or physiological discharge pattern of LC neurones. In the present study the number of pulses was kept minimal (3-5 pulses) whereas in other reports (Segal and Bloom, 1974b) very long trains (up to 10 seconds) were used. Such differences make i t d i f f i c u l t to compare the latencies, potencies and duration of evoked responses between studies. In l i g h t of the diffuse d i s t r i b u t i o n of NA terminals i n the dentate, special care was taken to i d e n t i f y the c e l l s that were recorded.,This resulted in the incl u s i o n of only 15 c e l l s recorded in 8 rats for the purposes of data analysis. A l l 15 c e l l s were monosynaptically activated by PP stimulation, in h i b i t e d 348 by MR and were l o c a l i z e d to the v i c i n i t y of the G-cell layer. These c r i t e r i a suggest that these c e l l s were in f a c t G-cells, however, i t i s not possible to determine whether the i n h i b i t i o n observed following multiple pulse LC stimulation was mediated by a monosynaptic input d i r e c t l y onto G-cells. Effect Of LC Stimulation On PP-evoked Bespouses Conditioning stimulation of LC resulted i n an increase i n the amplitude of the PP-evoked population spike without a l t e r i n g the amplitude or r i s e time of the PP-evoked EPSP. The e f f e c t i v e C-T i n t e r v a l s ranged between 20-80 msec and maximum potentiation (150-175% of control) was observed following 3-5 conditioning pulses whereas no potentiation was observed following single pulse stimulation of LC. The observation that the degree of potentiation was related to the proximity of the stimulating electrode to LC proper further supports a s p e c i f i c action of t h i s p a rticular nucleus on PP-evoked population spike. The additional observation that 6-OHDA inj e c t i o n s which deplete hippocampal NA block the potentiation, strongly suggests involvement of a noradranergic system. Since potentiation of the population spike amplitude was not accompanied by a concomittant increase i n the rate of r i s e of the EPSP i t i s not l i k e l y to be due to the action of LC on the entorhinal 349 cortex or the region of the PP synapse on the d i s t a l dendrites of G-cells. In t h i s respect the actions of LC stimulation on PP-evoked responses may be similar to those observed following stimulation of other e x t r i n s i c afferents including the commissural input and the MB. 9.J5 Summary E l e c t r i c a l stimulation of LC caused an increase in the amplitude of the dentate poulation spike which was evoked by a test PP volley. The same conditioning stimulation did not r e s u l t i n an increase i n the rate of r i s e of the EPSP suggesting that presynaptic mechanisms could not account for enhancement of the population spike response. Injections of 6-OHDA into the dorsal NA bundle resulted in depletion of hippocampal NA and abolished the ef f e c t s of LC stimulation on the population spike response. On the basis of these data, i t was proposed that a noradrenergic system modifies neuronal transmission in the dentate gyrus. 350 CHAFER I P i GEJERAI DISCUSSIQSJ-The results of the experiments described in this thesis have demonstrated that serotonergic and noradrenergic systems originating i n MS and LC, respectively, modulate RSA and/or the population response of G-cells to stimulation of the perforant path (PP) input. Detailed discussions of the present and previous data which support t h i s conclusion were presented at the end of each chapter. However, the physiological implications of the present data and how they r e l a t e to existing concepts of the functional role of the hippocampal formation have not been discussed. Previous e l e c t r o p h y s i o l o g i c a l and behavioural studies have emphasized that RSA may r e f l e c t or predict 1) a c t i v i t y of other brain areas such as the neocortex and the septal area (Green and Arduini, 1954; Petsche et a l , 1962) 2) overt behaviour (Vanderwolf, 1 969; Ranck, 1975) and 3) inferred mental processes (Gray, 1970; Hadel and 0'Keefe, 1974) . However, the mechanisms underlying RSA and the involvement of s p e c i f i c neuronal systems in t h i s response have recieved l i t t l e attention. In contrast, hippocampal population responses have been well characterized (Gloor et a l , 1964 ; Andersen and Lomo, 1966; Lomo, 1971a) and mechanisms which a l t e r t h e i r magnitude were, and continue to be, extensively investigated (Lomo, 1971b B l i s s and Lomo, 1973; Andersen et a l , 1977; Lynch et a l , 1977; Dunwiddie and 351 Lynch, 1978). More recently, studies which examine the re l a t i o n s h i p between hippocampal population responses , behaviour (Winson and Abzuq, 1978) and RSA {Assaf and M i l l e r in preparation ) have been i n i t i a t e d . , The present thesis has 1) delineated mechanisms which could underlie the effects of pharmacologically d i s t i n c t neuronal systems on hippocampal e l e c t r i c a l a c t i v i t y and 2) indicated that e x t r i n s i c afferents to the DG such as the commissural input {Chapter 7) and monoaminergic projection (Chapters 8 and 9) a l t e r the amplitude of the PP-evoked population spike. Furthermore, the observation that a raphe serotonin system, wich increased the amplitude of the population spike, also produced hippocampal desynchronization suggests that the magnitude of the population spike response may be related to or dependent on ongoing patterns of hippocampal e l e c t r i c a l a c t i v i t y . lQ.x.1 Patterns Of Hippocampal E l e c t r i c a l A c t i v i t y Previous studies had postulated that d i s t i n c t neuronal systems, probably originating in the brainstem, mediate the c h a r a c t e r i s t i c patterns of hippocampal e l e c t r i c a l a c t i v i t y (Green and Arduini, 1954; T o r i i , 1961; Macadar et a l , 1974). However, these studies did not elucidate anatomically or pharmacologically d i s t i n c t neuronal systems that could serve as a model for studying the mechanisms mediating J 352 the d i f f e r e n t patterns. The present observation that a serotonin-containing system produced desynchronization has f a c i l i t a t e d the delineation of neuronal mechanisms within the septal area that could underlie t h i s response. The postulated mechanisms, which included i n h i b i t i o n of i r r e g u l a r l y f i r i n g I-neurones and subseguent disruption of the bursting of B-neurones, were consistent with previous anatomical (Tombol and Petsche, 1969) and electrophysiological {Petsche et a l , 1962) studies i n d i c a t i n g a heterogenous population of neurones i n the medial septal nucleus. Althouqh the present data did not eliminate the p o s s i b i l i t y that other intra-and extraseptal mechanisms can underlie the desynchronization, they provide a model which could be readily tested. The additional observations that 1) MS neurones may be d i f f e r e n t i a t e d on the basis of their discharge patterns or t h e i r hippocampal projections and 2) kainate lesions, which destroy c e l l bodies without influencing f i b e r s of passage (Assaf and M i l l e r , 1978d) eliminate RSA, f a c i l i t a t e future studies on the role of i n t r a - and extraseptal mechanisms in the generation of hippocampal e l e c t r i c a l a c i t i v t y . On the basis of previous reports (Macadar et a l , 1974), LC simulation was expected to produce BSA s p e c i f i c a l l y . The neuronal mechanisms mediating t h i s response could then be compared with the desynchronization r e u l t i n g from MR stimulation. However, the r e s u l t s of chapter 5 indicated that 6-OHDA 353 i n j e c t i o n s which depleted forebrain NA did not block RSA e l i c i t e d by stimulation i n the region of LC, The additional observation that e l e c t r o l y t i c lesions of LC did not abolish RSA suggested that examination of t h i s system may not be s u i t a b l e for delineating mechanisms which mediate RSA. These findings, which were recently confirmed by Robinson (1978) i n the f r e e l y moving r a t , do not e n t i r e l y eliminate noradrenergic involvement in the generation of RSA since: 1) stimulation i n a di f f u s e region in the v i c i n i t y of LC also i n i t i a t e s RSA, thus the e f f e c t s of 6-OHDA inject i o n s on a noradrenergic component may be masked or even compensated for by these other systems 2) NA-containing nuclei located caudal to LC project to the septal area (Moore, 1978) and may be involved i n the generation of RSA 3) multiple pathways and postulated transmitters (reviewed in section 1. 3) are implicated in the generation of RSA and none of them, with exception of the septal area, may be c r i t i c a l f o r the generation of t h i s response. Perhaps, as Gray and his colleagues (1975) have proposed, noradrenergic systems lower the threshold for a pa r t i c u l a r freguency (7.7 Hz) which i s i n i t i a t e d by the septal area. A c t i v i t y of the septal area, perhaps ref l e c t e d by the output of B-neurones, may r e s u l t from the interaction between various neuronal systems which modulate hippocampal e l e c t r i c a l a c t i v i t y i n a manner appropriate to ongoing physiological states. 354 10 «_,2 The Significance Of HifiEocamgal Population Responses, Previous studies (Gloor et a l r 1964; Lomo, 1971a), taken together with the results of chapter 6, support the interpretaion that the f i e l d potentials recorded e x t r a c e l l u l a r l y i n DG following PP stimulation r e f l e c t the direct action of the entorhinal cortex on dentate granule c e l l s . Interest i n these responses stemmed from the observations that paired-pulse stimulation of PP resulted i n an increase i n the rate of r i s e of the EPSP and amplitude of the population spike (Lomo, 1971b; Steward et a l , 1976). This potentiation has been studied either following a b r i e f period of r e p e t i t i v e conditioning stimulation (usually 1-3 seconds at 50-100 Hz) which resulted i n long-term potentiation of the e x t r a c e l l u l a r EPSP and population spike (Bliss and Gardner-Hedwin, 1973) or following a single conditioning volley which produced potentiation of the same parameters for a short (less than 1 second) period of time (Lomo, 1971b; Steward et a l , 1976). Since both short and long-term potentiation have been demonstrated i n the PP-dentate projection and there i s no evidence to indicate that d i f f e r e n t fundamental mechanisms are involved, we w i l l assume, for the purpose of the present discussion, that they share common mechanisms. I n i t i a l l y i t was proposed that auqmented transmitter release underlies the greater synaptic 355 current and subsequent increase in the number of G-c e l l s evoked by the test PP pulse (Lomo, 1971b), Previous speculations that an increased quantal release of transmitter accounts f o r post-tetanic potentiation of neuromuscular transmission (Eccles and B a l l , 1951; L i l e y , 1956) provided impetus f o r t h i s proposal. However, i n the i n i t i a l studies (Lomo, 1971b; B l i s s and Lomo, 1973) increases i n the amplitude of the PP-evoked population spike were sometimes observed i n the absence of changes in the EPSP. For t h i s reason. B l i s s and Lomo(1973) and B l i s s (personal communication) proposed that mechanisms other than augmentation of synaptic current could enhance the responsiveness of the postsynaptic c a l l s . The present thesis provides support for the l a t t e r suggestion since conditioning stimulation of Comm, MR and LC increases the amplitude of the PP-evoked population spike without a l t e r i n g the PP synaptic potential. In the same animals, conditioning pulses delivered to the PP i t s e l f potentiated both the EPSP and the population spike. These observation imply that neuronal transmission from the entorhinal cortex to DG could be modulated by eit h e r 1) presynaptic mechanisms which increase the e f f i c i e n c y of the afferent input or 2) postsynaptic mechanisms which enhance the responsiveness of the receiving c e l l or i t s effector mechanisms. Both mechanisms could account for the potentiation observed during paired-pulse stimulation 356 of the PP but only postsynaptic mechanisms could account for the increase in population spike amplitude following conditioning pulses to e x t r i n s i c afferents which did not a l t e r the EPSP. The effects of stimulating e x t r i n s i c afferents cn the PP evoked responses have not been studied i n d e t a i l previous to the present work. However, Lomo (1971b) has shown that conditioning stimulation of the mossy f i b r e system, which antidromically activates G-cells, results i n an increase in the amplitude of the PP evoked population spike without changing the rate of r i s e of EPSPs. More recently, Alvarez and Gardner-Medwin (1976) reported a s i m i l a r e f f e c t following stimulation of the medial septal area. These data, taken together with the r e s u l t s of chapters 7-9, suggest that conditioning stimulation of s i t e s other than the PP can somehow increase the number of G-cells which are evoked by a constant PP synaptic current. Therefore presynaptic mechanisms can not account for the observred potentiation. Postsynaptic mechanisms that could account for the increase in the amplitude of the population spike include 1) a c t i v a t i o n of excitatory interneurones which p a r t i a l l y depolarize the G - c e l l soma (Andersen et a l , 1967) and 2) removal of factors normally preventing the PP from evoking a given G-cell. The contribution of excitatory interneurones i s unlikely since i n the 357 present study potentiation i s observed i n the absence of any excitation. Moreover, there i s no evidence for the existence of excitatory interneurones i n the immediate v i c i n i t y of G-cells. In f a c t , the common feature of a l l the inputs presently shown to produce potentation of the population spike response, i s i n h i b i t i o n of G-cells either d i r e c t l y , as in the case of MS, or subsequent to an activation as i n the case cf the PP and commissural inputs. The relationship between i n h i b i t i o n of G-cells and potentation of the population spike was examined in most d e t a i l following stimulation of MB since t h i s particular nucleus produced a potent i n h i b i t i o n which was not preceded by a c t i v a t i o n . The duration cf i n h i b i t i o n of G-cells was related to the length and degree of potentiation. Several mechanisms could account for t h i s apparent paradox between depression of spontaneous G-cell discharge and f a c i l i t a t i o n of the PP evoked population spike. At the simplest l e v e l i n h i b i t i o n of a large number of G-cells may reset their random a c t i v i t y thereby allowing a more synchronous discharge in response to a prepotent PP volley. Since the amplitude of the population spike r e f l e c t s , in part, the synchronous discharge of a large number of G-c e l l s , i t w i l l be enhanced., This synchronizing mechanism may also be operative during rhythmical septal-hippocampal a c t i v i t y . Perhaps, as Vinogrova (1975) had suggested e a r l i e r , information a r r i v i n g via 358 the PP i s modulated by the bursting discharge pattern of septal or hippocampal neurones. lQ.i.3 BfeltliliSSl Mctivitj And Neuronal Transmission I:Q_t h e_Den tat e_ Gyrus The discussion to t h i s point has been limited to mechanisms within the dentate gyrus i t s e l f which could increase the amplitude of the population spike response. An additional approach i s to regard the population spike response as the output of the dentate via the mossy f i b r e system onto CA3. Perhaps t h i s output i s regulated in a manner appropriate to ongoing physiological states by adjusting the responsiveness of dentate G-cells. As indicated i n Fig. 10-1, the CA3 region would play a p i v o t a l role since i t receives almost the entire dentate output and has both direct connections with the dentate via the associational/commissural systems and i n d i r e c t connections through the septal area. A decrease i n mossy f i b r e output, as i n the case of G-cell i n h i b i t i o n , would i n i t i a t e mechanisms that enhance the population spike response to a given PP input. The direct feedback may be mediated by changes i n the depolarization of proximal dendrites of G-cells thereby enhancing or shunting PP synaptic current generated on the d i s t a l dendrites. The i n d i r e c t septal loop may attenuate the input by i n i t i a t i n g the bursting discharge pattern of septal neurones (McLennan and 359 fTGj. 10- 1: grpgosed E x t r i n s i c Influences On Neuronal Transmission In The Dentate Gyrus. The s o l i d l i n e s indicate that information may be transmitted from the entorhinal cortex {EC) to the dentate gyrus (DG) via the perforant path (PP). The output from DG i s via the mossy f i b r e system (MF) onto CA3 region of the hippocampus. The dotted l i n e s indicate possible feedback control of EC-DG transmission by CA3 via the associational/commissural system, the medial septal area (MS) and brainstem NA and 5-HT-containing systems. 3 £ 0 o o co co o o L _ \ I « 1 > ~ ro < (J Q n i i i i i i i Q_ O L d I em -HI c CO lO < CO 361 Miller,1976) which may then 'gate* the incoming PP input (Vinogradova, 1975 ). The additional feature offered by the i n d i r e c t septal loop would be to match CA3 output to ongoing physiological states which produce the c h a r a c t e r i s t i c discharge patterns of septal neurones (chapter 3). The recent findings of behaviourally s p e c i f i c modification of the PP evoked population spike support the above suggestions (Winson and Abzug, 1978). Moreover, preliminary experiments (Assaf and M i l l e r , 1978e) show that the amplitude of the PP evoked population spike i s dependent on the ongoing pattern of hippocampal e l e c t r i c a l a c t i v i t y and septal unit discharge. I f the PP i s stimulated during BSA or when B-neurones are bursting the amplitude of the population spike i s r e l i a b l y smaller than that evoked by the same PP volley delivered during hippocampal desynchronization. It i s int e r e s t i n g to note that Winson and Abzug (1978) report that the PP evoked population spike i s smallest when the freel y moving rat i s in an a l e r t or aroused state and l a r g s s t during slow wave sleep. Previous studies have shown that BSA occurs during a l e r t states and desynchronized or ir r e g u l a r hippocampal a c t i v i t y i s recorded during slow wave sleep (Vanderwolf, 1972; Robinson, 1978). Taken together these studies suggest that PP-dentate transmission may be enhanced during hippocampal 362 desynchronization and attenuated during RSA. It i s noteworthy that when e l e c t r i c a l a c t i v i t y of the neocortex consists of desynchronized f a s t a c t i v i t y , as i n arousal states, rhythmical slow a c t i v i t y i s prominent i n the hippocampal formation (Green and Arduini, 1954). Perhaps th i s apparent r e c i p r o c i t y i s a r e f l e c t i o n of compensatory mechanisms within the septal-hippocampal axis which maintain constancy of cortico-hippocampal-thalamic transmission. The implication of t h i s constancy and whether long-term potentiation i s an i n t e g r a l component of i t must await further experiments. Likewise, understanding the relevance of PP-dentate transmission to o v e r a l l brain function depends on knowing 1) what i s the nature of the information relayed by the PP, 2) where does the hippocampus relay t h i s information and 3) how do e x t r i n s i c inputs to the hippocampal formation such as those originating i n the septal area and monoamine-containing nuclei modify t h i s information? 363 1IIIBEJCES ADEY, W.R., MERBILLEES, N.C.R., AND SUNDERLAND, S. ( 1956), The Entorhinal Area: Behavioral, Evoked Potential And Hi s t o l o g i c a l Studies Of Its Interrelationships With Brain-stem Regions, Brain* 79 x 414-439. ALGER, B.E. AND TEYLER, T.J. ; (1976). Long-term And Short-term P l a s t i c i t y In The CA1, CA3 And Dentate Regions Of The Rat Hippocampal S l i c e . , Brain r e s i J L 1J0 A 463-480. ALGER, B.E. AND TEYLEB, T.J. (1977) . A Monosynaptic Fiber Track Studied In Vi t r o : Evidence Of A Hippocampal CA1 Associational System? Brain £€§>*. Bull, f 2f_ 355-365. ALVAREZ-LEEFMANS, F.J. AND GARDNER-MEDWIN, A.R. (1975). Influences Of The Septum On The Hippocampal Dentate Area Which Are Unaccompanied By Field Potentials. J.. P h y s i o l X J t 14P-15P. H. AND LINDSLEY, D. B. (1972). D i f f e r e n t i a t i o n Of Two Reticulohypothalamic Systems Regulating Hippocampal A c t i v i t y . Elgctrpenceph,, C li.n,, Neurophysiol^ 32 A 209-226." N.E., DAHLSTROM, A., FUXE, K., LARSSON, K., OLSON, C. AND UNGERSTEDT,U. (1966). Ascending Monoamine Neurons To The Telencephalon And Diencephalon. Acta. Physiol. Scand. x 67 x 313-326. ANDERSEN, P. (1960). Interhippocampal Impulses. I i . Apical Dendrite Activation Of CA1 Neurones. Acta physiologica scandinayia t 48:Mr 178-208 ANCHEL, ANDEN, .364 ANDERSEN, P. (1975). Organization Of Hippocampal Neurones And Their Interconnections, In: the-hippocampus, vol. ± Ed. Isaacson, R. L. And Pribram.,K.H. ,New York: Plenum Press. 155-176. ANDERS EN, P. AND LOMO, T. (1966) . Mode Of Activation Of Hippocampal Pyramidal C e l l s By Excitatory Synapses On Dendrites. Experimental brain res, f 2 X 247-260 ANDERSEN, P., BLAND, B. H. AND DUDAR, J.D. (1973a). Organization Of The Hippocampal Output. Experimental brain r e s i A Hi. 152- 168. ANDERSEN, P., BLAND, B.H., MYHRER, T. AND SCHWART2KR0IN, P.A. (1977).,, Septo-hippocampal Pathway Necessary For Dentate Theta Production. Unpublished Manuscript. ANDERS EN, P., BLISS, T.V.P. AND SKREDE, K.K. (1971). Unit Analysis Of Hippocampal Population Spikes. Expj. Brain r e s ^ 1.3^  208-221. ANDERSEN, P., BLISS, T.V.P. AND SKREDE, K.K.,, (1971b). Lamellar Organization Of Hippocampal Excitatory Pathwa ys. Experi mental brain r e s L i 13x 222-238. ANDERSEN, P., ECCLES, J.C. AND LOYNING, Y. (1964a)., Location Of Postsynaptic Inhibitory Synapses Of Hippocampal Pyramids. Journal of neurophysiology, 27 x 592-607 ANDERSEN, P., ECCLES, J. C. AND LOYNING, Y. (1964b). Pathway Of Postsynaptic I n h i b i t i o n In The Hippocampus. Journal of neurophysiology,,. 27*. 608-619., ANDERSEN, P., GROSS, G.N. , LOMO, T. , AND SVEEN, 0. (1967). P a r t i c i p a t i o n Of Inhibitory And Excitatory Interneurones In The Control Of Hippocampal C o r t i c a l Output. In M.a.b. Brazier (Ed.)., the interneurgn^ Univ. C a l i f . Press, Los Angeles, Pp. 415-465. 365 ANDERSEM, P., HOLMQVIST, B. AND VOORHOEVE, P.E. (1966b). E n t o r h i n a l A c t i v a t i o n Of Dentate G r a n u l e C e l l s , A c t a p h y s i o l o g i c a s c a n d i n a y i c a f 6 6 x 448-460. ANDERSEN, P., SO NCBERG, S. H., SVEEN, 0. AND WIGSTROM, H. (1977). S p e c i f i c L o n g - l a s t i n g P o t e n t i a t i o n Of S y n a p t i c T r a n s m i s s i o n I n Hippocampal S l i c e s . N a t u r e , l o n d . , 266, 736-737. ANDY, O.J. AND STEPHAN, H. (1974). I n : t h e s e p t a l n u c l e i . J . F. De France (ed. ). Plenum P r e s s : New York, Pp. 3-36. ANDY, O.J. AND STEPHAN, H. C. (1 964). The Septum Of The Ca t . Thomas S p r i n g f i e l d , 111. APOSTOL, G. AND CBEUTZFELDT, O.D. (1 974). C r o s s -c o r r e l a t i o n Between The A c t i v i t y Of S e p t a l U n i t s And Hippocampal EEG During A r o u s a l , b r a i n -£ § § L S ^ L 67 t - 65-75. Afi BUTHNOTT, G.8., CROW, T. J . , FUXE, K. , OLSON, L. . AND UNGERSTEDT, U. ( 1970). D e p l e t i o n Of C a t e c h o l a m i n e s I n V i v o Induced By E l e c t r i c a l S t i m u l a t i o n Of C e n t r a l Monoamine Pathways. B r a i n r e s x x 2 4 A 471-483. , ASSAF, S.Y. AND MILLER, J . J . (1977). E x c i t a t o r y A c t i o n Of The M e s o l i m b i c Dopamine System On S e p t a l Neurones. B r a i n res±x 129.. 353-36 0. ASSAF, S.Y. AND MILLER, J . J . (1978a) . The R o l e Of A Raphe S e r o t o n i n System I n The C o n t r o l Of S e p t a l O n i t A c t i v i t y And Hippocampal D e s y n c h r o n i z a t i o n . , N e u r o s c i e n c e t 3, 539-550. , ASSAF, S.Y. AND MILLER, J . J . (1978b).; Neuronal Transmission In The Dentate Gyrus: Role Of Inhibitory Mechanisms. Brain resj_ x 151x 587-5 92. 366 ASSAF, S.Y. , AND MILLER, J . J . (1978c). Inhibitory Mechanisms Modifying Neuronal Transmission In The Dentate Gyrus Of The Bat. Canada EkZ§.iolog.y.x 51JLi-- 18. ASSAF, S.Y. AND MILLER, J.J. (1978d) . A Comparison Of The E f f e c t s Of E l e c t r o l y t i c And Kainic Acid Induced Lesions Of The Septal Area On Hippocampal E l e c t r i c a l A c t i v i t y , Proc., Can, I§ l i a I A So C j _ 21 x 223. ASSAF, S.Y. AND MILLER, J.J. (1978e). Relationship Between Cortical-hippocampal Transmission And Rhythmical A c t i v i t y In The Septum., In p.reper atign. BAGSHA W, E.V, AND EVANS, M. H« Current Spread From Stimulating Within The SEain r e s l t 25 x 391-400. (1976). Measurement Of Microelectrodes When Ne rvous S ystem. EXJD,_ -BARBER, R.P., VAUGHN, J.E. AND WIMER, R. E. (1974). Genetically-associated Variations In The Dis t r i b u t i o n Of Dentate Granule C e l l Synapses Upon The Pyramidal C e l l Dendrites In Mouse Hippocampus. J x Comp.., Neurol^.x -^ 15.6^  417-434. BARTOLINI, A., WEISENTHAL, L. M. AND DOMINO, E. F. (1972). Effect Of Photic Stimulation On Acetylcholine Release From Cat Cerebral Cortex. NeuroEharmacolj.4. J J X 113-122. BAUST, «. , NIEMCZYK, H. ,. AND VIETH, J. . (1963)., The Action Of Blood Pressure On The Ascending Reticular Activating System With Special Reference To Adrenaline-induced EEG Arousal. Electroen cepji^ C l i n A Neu ropjiys i o l X x J 5 A 63-72. BENNET, T. L. , HERBERT, P.N. AND MOSS, D.E. (1 973). Hippocampal Theta A c t i v i t y And The Attention Component Of Discrimination Learning, Behavioral biology f 8 X 173-181, 367 BISCOE. T.J, AND STRAUGHAN, D.H. (1966). M i c r o -e l e c t r o p h o r e t i c S t u d i e s Of Neurones I n The Cat Hippocampus. J*. P h j [ s i o ^ j _ x 183__, 341-3 59. BLACKSTAD, T.W.,{1956). Commissural C o n n e c t i o n s Of The Hippocampal Region I n The Rat, With S p e c i a l R e f e r e n c e To T h e i r Mode Of T e r m i n a t i o n . . J*, Coilj.-JSfifl£i2l».x J f i l x 417-537. BLACKSTAD, T. W. (1958) . On The T e r m i n a t i o n Of Some A f f e r e n t s To The Hippocampus And F a s c i a Den t at a. ^  Actaj. • An at A i t 35__ 202-214. BLAND, B.H., ANDERSEN, P., AND GANES, T. (1975). Two G e n e r a t o r s Of Hippocampal Theta A c t i v i t y I n R a b b i t s . Brain. •res i > i, 9 4 x 199-218, BLISS, T. V.P. AND G ARBNER-MEDWIN, A.B. (1973). Long-l a s t i n g P o t e n t i a t i o n Of S y n a p t i c T r a n s m i s s i o n I n The Dentate Area Of The U n a n a e s t h e t i z e d R a b b i t F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t P a t h . J__ P h y ^ i o l X A 232* 357-374. BLISS, T.7.P.., AND LOMO, T. (1973). L o n g - l a s t i n g P o t e n t i a t i o n Of S y n a p t i c T r a n s m i s s i o n I n The Dentate Area Of The A n a e s t h e t i z e d R a b b i t F o l l o w i n g S t i m u l a t i o n Of The P e r f o r a n t Path. J . -P h y s i g l A (Lond.). 2 3 2 x 3 31-356. BO BILL ER, P., SEGDIN, S. , PETITJEAN, F. , SALVERT, D. , TOURET, M. AND JO0TET,M. (1976). The Raphe N u c l e i Of The C a t B r a i n Stem; A T o p o g r a p h i c a l A t l a s Of T h e i r E f f e r e n t P r o j e c t i o n s As Revealed By A u t o r a d i o g r a p h y . B r a i n . - R e s , M \ 1 3 . 449-486. BRADLEY, P.B. AND NICHOLSON, A.N. , (196 2). , The E f f e c t Of Some Drugs On Hippocampal A r o u s a l . 11 §gt.£ogSceg h. C l i n . Neuroph y s i o l . , 14* 824-834, BREMER, F. (1935). Cerveau " i s o l e " Et P h y s i o l o g i e Du Sommeil. , C.r. Soc. B i o l . . I P a r i s l * 118__1235-1241. . 368 BREMER, F. AND Cortex 109. CHATONNET, J. (1949). Acetylcholine Et Cerebral. Arch.. Int.. p hysigl. x 57 < 106-BBODAL, A. AND BOSS I, G.F. (1 955) . Ascending Fibres In Brain Stem Reticular Formation Of Cat. Arch. : Neurol.. P s y c h i a t x x 74 x 6 8-87. BRUCKE, F., PETSCHE, H. , PILLAT, B. AND DEISENHAMMER, E. (1959b). Die Beeinflussing Der "Hippocampus-arousal-Reaktion" Beim Kaninchen Durch Elektrische Reizung In Septum, Pflugers archti•• G e j i , P h y s i c l x x 269 x 319- 338. BRUCKE, F. , PETSCHE, H. , PILLAT, B. AND DIESENHAMMER, E. (1959a). Uher Veronderungen Des Hippocampus—Elektrencephalogr amines Biem Kaninchen Nach Novocainin-jektion In Die Septumragion. Arch x Exp. Path x Pharmak. ,t 237. 276-284. BRUCKE, F. , SAILER, S. AND STUMPF, C. (1958). wechselwirkungen Zwischen Physostigmin E i n e r s e i t s Und Evipan, Procain, L a r g a c t i l Und Scopolamin Undererseits Auf Die Rhinencephale Tatigkeit Des Kaninchens. Naunyn^Schmiedeberg^ archj, Exg x Path x Pharmakol7 x 2327- 433-44l7~~ BRUGGE, J.F. (1965). An Electrographic Study Of The Hippocampus And Neocortex In Unrestrained Bats Following Septal Lesions. Electrgenceph,. ; Clin..-Neurgphysiol^. 18^ 36-44. CAJ AL, R.Y (1911) Histglogie du systeme nerve us- T e i l 2A, Paris. CAJAL, R.Y (1968) S pringfieId: the structure of Charles C. Thomas. ammon's horn. CAPON, A. (1960). Analyse De L'effect D»eveil Exerce Par I1 adrenaline Et D* autres Amines Sympathico-mimetigues Sur L•electrocorticogramme Du Lapin Non Narcotise. Arch* Int xPbarmacgdyn X x J27 x 141-162. CARLSSON, A., F&ICK, B. AND HILLARP, N. A. (1962)., C e l l u l a r L o c a l i z a t i o n Of B r a i n Monoamines. A^cta E k i s i o l i . S c andij. 56 x 1-28. CELESIA, G.G. , AND JASPEB, H.H. { 1966) . A c e t y l c h o l i n e R e l e a s e d From C e r e b r a l C o r t e x I n R e l a t i o n To S t a t e Of A c t i v a t i o n . Neurology JMinneajOjtLj, J6 X-1053-1063. CHRONISTER, R. B. AND WHITE, L.E., JR. F i b r e a r c h i t e c t u r e Of The Hippocampal F o r m a t i o n ; Anatomy, P r o j e c t i o n s And S t r u c t u r a l S i g n i f i c a n c e . I n R.L., I s a a c s o n And K.H. Pribam ( E d s . ) , the fei£EO£amp_usx V o l . XL s t r u c t u r e and development^ new y o r k : plenum Pp. 41-59. CHO, N. . AND BLOOM, F.E. (1973). N o r e p i n e p h r i n e -c o n t a i n i n g Neurons: Changes I n Spontaneous D i s c h a r g e P a t t e r n s During S l e e p And Waking. S c i e n c e * 179 A 108-109. CONBAD, L.C. A., LEONARD, C. M. AN D PFAFF, D. W, (1974)., C o n n e c t i o n s Of The Median And D o r s a l Raphe N u c l e i In The R a t : .An A u t o r a d i o g r a p h i c And D e g e n e r a t i o n Study. J . C o m p t Neurol,_ x 156, 179-206. CONSOLO, S. , LADINSKY, H. , BIONCHI, S, AND GHEZZI, D. (1977). Apparent Lack Of A Dopaminergic-c h o l i n e r g i c L i n k I n The Rat Nucleus Accumbens S e p t i - t u b e r c u l u m O l f a c t o r i u m . B r a i n r e s j . x 1 3 5 x 255-263. W.M., GOTTLIEB, D. I . , HENDRICKSON, A. E. , PRICE, J.L. , AND WOOLSEY, T.A. (1972). The A u t o r a d i o g r a p h i c Demonstration Of Axonal C o n n e c t i o n s In The C e n t r a l Nervous System. B r a i n r e s i j t 3 7 x 21-51. J.T. AND HENRY, D. , (1 973). Catecholamines In F e t a l And Newborn Rat B r a i n , J o u r n a l of figlJEOchemistryji, 2 1 x 61-68. COWAN, COYLE, 370 COYLE, J.T. AND SCHWARCZ, R. (1976). Lesions Of S t r i a t a l Neurones With Kainic Acid Provides A Model For Huntington's Chorea. Nature x 263 x 244-246. CRAGG, B.G. AND HAMLYN, L.H. (1957)., Some Commissural And Septal Connections Of The Hippocampus In The Rabbit. J i P h i s i o l i . x 135 x 460-485. DAHLSTROM, A. AND FUXE, K. (1964). Evidence For The Existence Of Monoamine-containing Neurons In The Central Nervous System. I. Demonstration Of Monoamines In The C e l l Bodies Of Brain Stem Neurons. Acta p h y s i o l ^ Scand x x 6-2x~Suppl. .. 232, 1-55. DALE, H.H., FELDBERG, W. ANDVOGT, M. (1 936). Belease Of Acetylcholine At Voluntary Motor Nerve Endings. J x P h y s i o l x x 86 x 353,380. DEADWYLEB, S.A.# WEST, J.B., COTMAN, C.W. AND LYNCH, G.S. (1975). A Neurophysiological Analysis Of Commissural Projections To Dentate Gyrus Of The Rat. Jjj. Nejiro£hysiolJS_x 3 8 167-184. DINGL EDIN E, B., DODD, J. AND KELLY, J.S. (1977). I n t r a c e l l u l a r Recording Of Pyramidal Neurones In The In Vitro Hippocampal S l i c e . J x P h y s i o l X J i 269iH x 13P-15P. DOM ER, F.R. AND LONGO, V.G. (1962). Effect Of 5-hydroxytryptophan On The Cerebral E l e c t r i c a l A c t i v i t y Of The Rabbit. Areh x Int. PharmaeoXj. • 136 x 204-218. DUDAR, J.D. (1975). The Effect Of Septal Nuclei Stimulation On The Release Of Acetylcholine From The Rabbit Hippocampus. Brain r e s t _ x 83 x 123-133. DUDAR, J.D. (1977). The Role Of The Septal Nuclei In The Release Of Acetylcholine From The Rabbit Cerebral Cortex And Dorsal Hippocampus And The Effect Of Atropine. Brain r e s x x 129 x 237-246. 371 DUDEK, F.E., DEADWYLER, S.A., COTMAN, C. W. AND LYNCH, G. (1976) , I n t r a c e l l u l a r Responses From G r a n u l e C e l l L a y e r I n S l i c e s Of Rat Hippocampus. B r a i n r e s l t 39 t 3 84-3 93 . DUNWIDDIE, T. AND LYNCH, G. (1978). Long-term P o t e n t i a t i o n And D e p r e s s i o n Of S y n a p t i c Responses I n The Eat Hippocampus: L o c a l i z a t i o n a n d Freguency Dependency. J . , P h y s i o l ^ „• 276^ , 353-367. DUNWIDDIE, T. , MADISON, D. AND LYNCH, G. (1978)., S y n a p t i c T r a n s m i s s i o n I s R e g u i r e d F o r I n i t i a t i o n Of Long-term P o t e n t i a t i o n . B r a i n -resj. t 150^ 413-417. ECCLES, J.C, AND BALL, W. (1951). E f f e c t s Induced In A Monosynaptic R e f l e x P a t h By I t s A c t i v a t i o n . J*. Neurojehysiola.* J4 A- 353-376. ECCLES, J . C. (1957) . P h y s i o l o g y o f a g o g c e l l s , , John Hopkins P r e s s , B a l t i m o r e , ECCLES, J . C , KATS, B.,AND RUFFLE R, S.W. (1941) Nature Of The E n d - p l a t e P o t e n t i a l I n C u r a r i z e d Muscle. H± N e u r o p h y s i p l , 4 A 492-516. ELAZAR, Z. AND ADEY, W. R. (1967) . S p e c t r a l A n a l y s i s Of Low Freguency Components I n The E l e c t r i c a l A c t i v i t y Of The Hippocampus During L e a r n i n g . E l e c t r o en cephi. And c ! i n A N e u r o p h y s i o l j , x 23.*-225-240. ELLIOT SMITH, G. (1897). T*e r e l a t i o n o f t h e f o r n i x - t o the margin of the c e r e b r a l c o r t e x A EPSTIEN, B. AND TAUC, L. (1970). Heterosyna pt i c F a c i l i t a t i o n And P o s t t e t a n i c P o t e n t i a t i o n In A p l y s i a Nervous System. Jx P h y s i o l X J L 209^ , 1-23. 372 EULER, C. VON, AND GREEN, J.p., (1960). A c t i v i t y I n S i n g l e Hippocampal Pyramids. Acta..- P h y s i c l . S c a n d ^ 4 8 x 95-109. FALCK, B. (1 962) . O b s e r v a t i o n s On The P o s s i b i l i t i e s Of The C e l l u l a r L o c a l i z a t i o n Of Monoamines By A F l u o r e s c e n c e Method. Acta • p h y s i o l . Scand. * 56* S u p p l . 197, 1-25. FALCK, B. , HILLARP, N. A. , THIEME, G. AND TORP , A. (1962). F l u o r e s c e n c e Of Ca t e c h o l a m i n e s And R e l a t e d Compounds Condensed With Formaldehyde. J»• • Histochem. Cytoehem, M i 1 0 x 348-354. FAREL, P.B, AND THOMPSON, H.F. (1976). H a b i t u a t i o n Of A Monosynaptic Response In Frog S p i n a l Cord: E v i d e n c e F o r A P r e s y n a p t i c Mechanism. J.. N euro phys. i g l j . x 3 9 6 6 1 - 6 8 2 . FEDER, R. AND RANCK, J . B. (1973). S t u d i e s On S i n g l e Neurons I n D o r s a l Hippocampal F o r m a t i o n And Septum I n U n r e s t r a i n e d R a t s . I I . Hippocampal Slow Waves And Th e t a C e l l F i r i n g D u r i n g Bar P r e s s i n g And Other Behaviours. , E x p e r i m e n t a l -aeu£gl*_jt. HIJL 532-555. FIFKOVA, E. AND VAN HARREVELD, A. (1977). L o n g - l a s t i n g M o r p h o l o g i c a l Changes In D e n d r i t i c S p i n e s Of Den t a t e G r a n u l a r C e l l s F o l l o w i n g S t i m u l a t i o n Of The E n t o r h i n a l Area. J x Neurocyte logy,*. 6 X 211-2 30. FONNUM, F. , (1970). T o p o g r a p h i c a l And S u b c e l l u l a r L o c a l i z a t i o n Of C h o l i n e A c e t y l t r a n s f e r a s e I n Rat Hippocampal R e g i o n . J A Neurochem., 17* 1029-1037. FOX, C. A. (1940). C e r t a i n B a s a l T e l e n c e p h a l i c C e n t e r s I n The Cat. J . Comp. Ne u r o l . f 72. 1-62. FUJITA, Y. AND IWASA, H. (1976). The T r i g g e r Zone In The Hippocampal P y r a m i d a l C e l l . J x P h y s i c l ^ . S Q C a J a p a n x 38^ 108. 373 FUJITA, Y. AND SATO, T. (1 964) , I n t r a c e l l u l a r Reacords From Hippocampal C e l l s I n R a b b i t D u r i n g Theta Rhythm A c t i v i t y . J.. N§jirophysiol.tx 2 7 x 1011-1025. FOXE, K. (1965). E v i d e n c e F o r The E x i s t e n c e Of Monoamine Neurons I n The C e n t r a l Nervous System. I V . , D i s t r i b u t i o n Of Monoamine Nerve T e r m i n a l s In The C e n t r a l Nervous System. A c t a fikxsigii. S c a n d i J t 64^ S u p p l . 247, 3 9-85. FUXE, K. , H AMBERGER, B. AND HOKFELT, T. (1 968). D i s t r i b u t i o n Of N o r a d r e n a l i n e Nerve T e r m i n a l s I n C o r t i c a l Areas Of The Rat. B r a i n r e s ^ 8 X 125-131. FUXE, K. , HOKFELT, T. AND ONGERSTEDT, U. (1969). D i s t r i b u t i o n Of Monoamines I n The Mammalian C e n t r a l Nervous System By H i s t o c h e m i c a l S t u d i e s . Ins metabolism o f amines i n the b r a i n E. Hooper (Ed.j~. London: M a c M i l l a n , Pp. 10-22. GERSTEIN, G. AND KIANG, N.S (1960). An Approach To The Q u a n t i t a t i v e A n a l y s i s Of E l e c t r o p h y s i o l o g i c a l Data From S i n g l e Neurons, B i o p h y s i c a l j o u r n a l * -1* 15-28. GLOOR, P, (1955). E l e c t r o p h y s i o l o g i c a l S t u d i e s On The C o n n e c t i o n s Of The Amygdaloid N u c l e u s I n The C a t . I I . The E l e c t r o p h y s i o l o g i c a l P r o p e r t i e s Of The Amygdaloid P r o j e c t i o n S y s t e . E l e c t r o e n c e p h • , Clin.4. Neu£ophysiol i x 7* 243-264. ~ GLOOR, P. (1963). I d e n t i f i c a t i o n Of I n h i b i t o r y Neurones I n The Hippocampus. N a t u r e * 1 9 9 x 699-670. GLOOR, P., VERA, C.L. AND SPERTI, L. , (1964). E l e c t r o p h y s i o l o g i c a l S t u d i e s Of Hippocampal Neurons. I I I . Responses Of Hippocampal Neurons To R e p e t i t i v e P e r f o r a n t P a t h V o l l e y s . E l e c t r o e n c e p h a l o . g r a phy and c l i n i c a l -n e u r o p h y s i o l o g y M VJ^ 353-370. 3 7 4 GOGOLAK, G, , STUMPF, CH. , PETSCHE, H. AND STERC, J . (1968). The F i r i n g P a t t e r n Of S e p t a l Neurons And The Form Of The Hippocampal Theta Wave. Biain r e s ^ 7 X 2 01-207. GOGOLAK, G., STUMPF, CH., PETSCHE, H. . AND STERC, S. (1968). The F i r i n g P a t t e r n Of S e p t a l Neurons And The Form Of The Hippocampal T h e t a Wave. B r a i n £es,_x 7 A 201-207, GOTTLIEB, D.T. AND COWAN, W.M. (1973) . A u t o r a d i o g r a p h i c S t u d i e s Of The Commissural And I p s i l a t e r a l A s s o c i a t i o n C o n n e c t i o n s Of The Hippocampus And Dentate Gyrus Of The Rat. J. Comp... N e u r o l ^ 149,. 393-422. GRANIT, R. AMD PHILLIPS, C.E. 0 9 5 6 ) . E x c i t a t o r y And I n h i b i t o r y P r o c e s s e s A c t i v e Upon I n d i v i d u a l P u r h i n i e C e l l s Of The C e r e b e l l u m I n Cats. J,--£kysioli«. J 3 3 x 520-547. GRANTYN, A.A. (1970). An I n t r a c e l l u l a r Study Of Hippocampal Responses To R e t i c u l a r S t i m u l a t i o n . B r a i n r e s ^ 2 2 X 409-412. GRANTYN, A.A., AND GRANTYN, R. (1972) . P o s t s y n a p t i c Responses Of Hippocampal Neurons To M e s e n c e p h a l i c S t i m u l a t i o n : h y p e r p o l a r i z i n g P o t e n t i a l s . B r a i n res:xji 45* 87-100. GRASTYAN, E., KARMOS, G., 7EEECZKEY, L. AND KELLENYI, L. (1966). , The Hippocampal E l e c t r i c a l C o r r e l a t e s Of The Homeostatic R e g u l a t i o n Of M o t i v a t i o n . E l e c t r pence ph.,; And c l i n j . M e u r o p h y s i o l . K 1.1, 34-53. GRAY, J.A. (1970). Sodium A m o b a r b i t a l , The Hippocampal T h e t a Rhythm, The P a r t i a l R e i n f o r c e m e n t E x t i n c t i o n E f f e c t And The P s y c h o p h y s i o l o g i c a l Nature Of I n t r o v e r s i o n , P s y c h o l o g i c a l r e v i e w , 11M. 465-480. 3 75 GRAY, J.A. (1971), M e d i a l S e p t a l L e s i o n s , Hippocampal Theta Rhythm, And The C o n t r o l Of V i b r i s a l Movement I n The F r e e l y Moving R a t . E l e c t r pence ph. C l i n . N e u r o p h y s i o l . •*•• 30*189-197. . GRAY, J. A. , MCNAUGHTQN, N. , JAMES, D. T. AND KELLY, P.H. (1975). E f f e c t Of Minor T r a n q u i l i z e r s On Hippocampal Theta Rhythm Mimicked By D e p l e t i o n Of F o r e b r a i n N o r a d r e n a l i n e . N a t u r e , 258* 424-425. GREEN, J.D. (1964). The Hippocampus. P h y s i o l * Bey*.* 44* 561608. GREEN, J.D. AND ARDUINI, A. A. (1954).. Hippocampal E l e c t r i c a l a c t i v i t y I n A r o u s a l . J o u r n a l o f n e u r o p h y s i o l o g y . J 7 X 533-557. GREEN, J.D. AND PETSCHE, H. (1961). Hippocampal E l e c t r i c a l A c t i v i t y . I I . V i r t u a l G e n e r a t o r s . E1ectroeneeph x C l i n * N e u r o p h y s i o l x * 13* 847853. GBEEN, J.D., MAXWELL, D. S. , SCHINDLER, H. S. , &ND STUMPF, C. (1960). R a b b i t EEG ' t h e t a Rhythm': I t s A n a t o m i c a l Source And R e l a t i o n To A c t i v i t y In S i n q l e Neurons. J * ; N e u r o p h y s i o l * * 23-*- 403-420. GUILLERY, R. W. (1956). D e g e n e r a t i o n I n The P o s t -c o m m i s s u r a l F o r n i x And The Mammilary Peduncle Of The Rat. J j . A n a t ^ 91* 35 0-370. GUILLERY, B.w. , (1957) . D e g e n e r a t i o n I n The Hypothalamic Connexions Of The A l b i n o R a t . J * Anat** 9JL* 91-115. 376 HALARIS, A.E., JONES , B.E. , AND MOORE, R. Y. .. (1976) . Axonal T r a n s p o r t I n S e r o t o n i n Neurons Of The M i d b r a i n Raphe. B r a i n r e s t J t 1 0 7 * 5 5 5 - 5 7 4 . HARPER, R.M. {1971) . Frequency Changes I n Hippocampal E l e c t r i c a l A c t i v i t y During Movement And T o n i c I m m o b i l i t y . P h y s i o l o g y , and behavior,, 7 5 5 - 5 8 . HARPER, R.M. (1971). Frequency Changes I n Hippocampal E l e c t r i c a l A c t i v i t y D uring Movement And T o n i c M o b i l i t y . P h y s i o l . Behay** 7* 55- 58. HARTZELL, H.C., KDFFLER, S.W. AND YOSKIKAMI, D. (1975). P o s t - t e t a n i c P o t e n t i a t i o n : I n t e r a c t i o n Between Quanta Of A c e t y l c h o l i n e At The s k e l e t a l Neuromuscular Synapse. J t P h y s i o l * , * 251* 427-463. HATTORI, T. AND MCGEER, E.G. (1977). F i n e S t r u c t u r a l Changes In Rat S t r i a t u m A f t e r L o c a l I n j e c t i o n s Of K a i n i c A c i d . B r a i n r e s . * 1 2 9 1 7 4 - 1 8 6 . HAYAT, A. AND FELDMAN, S, (1974). E f f e c t s Of Sensory S t i m u l i On S i n g l e C e l l A c t i v i t y I n The Septum Of The Cat. Exp f .Neurol. . 40 f 298-313. HEBZ, A. AND NACIMIENTO, A.C. (1965). Uber Die f i r k u n q Von Pharmaka Auf Neurone Des Hippocampus Nach M i k r o e l e k t r o p h o r e t i s c h e r V e r a b f o l g u n g . , Naunyn-Schmiedebergl.s a r c h * Exp. P a t h * P h a r m a l e i * * 25_1* 295-314. HINES, M. (1922). S t u d i e s On The Growth And D i f f e r e n t i a t i o n Of The T e l e n c e p h a l o n I n Man. The F i s s u r e Hippocampi. J*. Comp*. Neuioi*.* 34*-73-171. HJORTH-SIM0NSEN, A. (1972). P r o j e c t i o n Of The L a t e r a l P a r t Of The E n t o r h i n a l Area To The Hippocampus And F a s c i a Den t a t a. J * Comp* N e u r o l * * 146* 219-232. 377 HJORTH-SIMONSEN, A. (1973) Of The Hippocampus Analysis.. J._ Comp^ . . Some I n t r i n s i c Connections In The Rat: An Experimental I§.urolX4. 147,,- 145-162. H JORTH-SIMONSEN, A. AND J EUNE, B. (1972). Origin And Termination Of The Hippocampal Perforant Path In The Rat Studied By Si l v e r Impregnation. J«_ Coffi. N e u r o l A x J 4 4.* 215-232. H JORTH-SI HONS EN, A. AND L AURBEBG, S. (1978). Commissural Connections Of The Dentate Area In The Bat. , J,_ Comfit Neu£ol i x J74 x 591-605. H JORTH-SI MO NSEN, A. (1972)i. Projections Of The La t e r a l Part Of The Entorhinal Area To The Hippocampus And Fascia Dentata. J& CofflEA Neurol.1,46,, 219-232. HOBNYKIEWICZ, 0. (1966). Dopamine (3-hydroxyt yramine) And Brain Function. Pharmacglj. B e v i t 18 x 925-965. JACOBOHITZ, D.M. AND PALKOVITS, M. (1974 ). Topographic Atlas Of Catecholamine And Acetylcholinesterase-containing Neurons In The Rat Brain. , I. Forebrain. J.. Coni£t N e u r o l ^ 157* 13-28. JACOBS, B. L* i WISE, W.D. AND TAYLOR, K.M. (1975). D i f f e r e n t i a l Behavioural And Neurochemical Effects Following Lesions Of The Dorsal And Median Raphe Nuclei In Rats. Brain r e s t 4 79 t 3 53-361., JASPER, H. H. AND KOYAMA, I. ( 1969) . Bate Of Belease Of Amino Acids From The Cerebral Cortex In The Cat As Affected By Brainstem And Thalamic Stimulation. Canad. J i . Physiolj. Pharmacol,*- 47,,. 889-905. JASPEB, H.H. , KHAN, E.T. AND ELLIOTT, K.A.C. (1965). Amino Acids Beleased From The Cerebral Cortex In Relation To Its State Of Activation. Science. 147^ 1448. 378 JONES, B.E. AND MOOBE, B.Y. containing Neurons Of Coeruleus In The Cat. 4 3-42. (1974). Catecholamine-The Nucleus Locus Jjt :.C211t•:\N^ 10^ i••^ JIS7J^ •• JONES, B. E. , BOBILLIEB, P., PIN, C. AND JOUVET, 8. (1973). The E f f e c t Of Lesions Of Catecholamine-containing Neurons Upon Monoamine Content Of The Brain And EEG And Behavioural Waking In The Cat. Brain res,, x 58* 157-177, JONES, B.E. , HALABIS, A. E. , MCILHANY, M. AND MOOBE, B.Y. {1977). Ascending Projections Of The Locus Coeruleus In The Rat. I, Axonal Transport In Central Noradrenaline Neurons. Brain - r e s ^ 127 x 1-21., JONES, B.E., HALABIS, A.E. # MCILHANY, M. AND MOOBE, B.Y. (1977), Ascending Projections Of The Locus Coeurleus In The Bat. L. Axonal Transport In Central Noradrenaline Neurons. Brain r e s A J t 127X JONES, B. E. , HARPER, S.T. AND HALARIS, A.E. (1977). E f f e c t s Of Locus Coeruleus Lesions Upon Cerebral Monoamine Content, Sleepwakefulness States And The Response To Amphetamine In The Cat. Brain res.*,*. J24 A 473-496. JOUVET, M. (1974). Monoaminergic Regulation Of The Sleep-waking Cycle In The Cat. In F. o. Schmitf And E.g. Worden (Eds,), the neurgscience s; t h i r d study program, Cambridge, Mass.: MIT Press, 499-508. JUNG, R. AND KORNMULLER, A.E. (1938), Eine Methodik Der Abkitung L o k a l i s i e r t e r Potential Schwankungen Aus Subcorticalen Hir ngebieten. Arch;,,, Pshychiatjj. Ne rjrenkr^nkhx * 109 1-30. KANDEL, E.B. AND SPENCEB, W.A. (1961). Electrophysiology Of Hippocampal Neurons, I I : After-potentials And Repetitive F i r i n g . J t Neurophisiolxx 2 4 X 243-2 59. 3 79 KAWAMURA, H. AND DOMINO, E.F. {1968).„Hippocampal Slow ("ar o u s a l " ) Wave A c t i v a t i o n I n The R o s t r a l M i d b r a i n T r a n s e c t e d Cat. E l e c t r o e n c e p h j , C l i n . N e u r o p j i y s i o l i x 2 5 x 471-480. KAWAMURA, H. AND DOMINO, E.F. (1968). Hippocampal Slow ( ' a r o u s a l *) Wave A c t i v a t i o n In The R o s t r a l M i d b r a i n T r a n s e c t e d Cat. E l e e t r g e n c e p j i x C l i n ^ M u r o p h y s i o l i x 2 5 , , 471-48o7" KIMBLE, D. P. (1963). The E f f e c t s Of B i l a t e r a l Hippocampal L e s i o n s I n Rats. J o u r n a l of c o m p a r a t i v e and p h y s i o l o g i c a l p.sycijology x 56 A 273-283. ~ ~~ KLEMM, W.R. (1971). EEG And M u l t i p l e - u n i t A c t i v i t y I n L i m b i c And Motor Systems During Movement And I m m o b i l i t y . P h y s i o l * . B e h a v x x 7 X 3 37-343. KLEBM, W.R. (1972b). E f f e c t s Of E l e c t r i c S t i m u l a t i o n Of B r a i n Stem R e t i c u l a r F o rmation On Hippocampal T h e t a Rhythm And Muscle A c t i v i t y I n U n a n a e s t h e t i z e d , C e r v i c a l And M i d b r a i n T r a n s e c t e d Rats. B r a i n r e s x x 4 1 x 331-344. KOBAYASHI, R.M., PALKOVITS, M., KOPIN, I . J . AND JACOBOHITZ, D.M. (1974). B i o c h e m i c a l Mapping Of N o r a d r e n e r g i c Nerves A r i s i n g From The Rat Locus C o e r u l e u s . B r a i n r e s , . x 7 7 * 269-279. KOE, B.K. AND WEISSMAN, A. (1966). F-c h l o r o p h e n y l a l a n i n e : A S p e c i f i c D e p l e t i o n Of B r a i n S e r o t o n i n . J . Pharmac,. Exp._ T h e r , x 154,.-499-516. KOELLE, G. B. ( 1965) . A n t i c h o l i n e s t e r a s e A gents. I n L.s. Goodman And A. Gilman ( E d s . ) , the-p h a r m a c o l o g i c a l b a s i s o f t h e r a p e u t i c s * - T o r o n t o : M a c M i l l a n , Pp. 441-463. KOLB, B. , AND W HIS HAW, I . Q. (1977) E f f e c t s Of B r a i n L e s i o n s And A t r o p i n e On Hippocampal And N e o c o r t i c a l EEG In The Rat. E x p x N e u r o l x x 5 6 x 1-22. 380 KONIG, F.C. AND KLIPPEL, R. A. (1963) . The r a t bra i n * B a l t i m o r e : W i l l i a m s And S i l k i n s . KONORSKI, J . , SANTIBAN EZ-H, H.G. AND BECK, J . (1 968). E l e c t r i c a l Hippocampal A c t i v i t y And Heart Hate In C l a s s i c a l And I n s t r u m e n t a l C o n d i t i o n i n g . i£ta MoioSLiae e x p e r i m e n t a l i s , 28* 169-185. KRNJEVIC, K. (1974). C h e m i c a l Nature Of S y n a p t i c T r a n s m i s s i o n In V e r t e b r a t e s . P h y s i o l * Rev.*74 A 418-540. KRNJEVIC, K. AND SILVER, A. (1965). A H i s t o c h e m i c a l Study Of C h o l i n e r g i c F i b e r s I n The C e r e b r a l C o r t e x . J . Anat, . 99, 711-759. KUHAR, M.J. (1975). C h o l i n e r g i c Neurons: S e p t a l -h ippocampal R e l a t i o n s h i p s . I n R.. I s a a c s o n And K.H. P r i b r a m ( E d s . ) , the h i p p o c a m p u s * , ^ S t r u c t u r e and development* New York: Plenum, Pp. 269-283. a. KUHAR, M.J. AND SCMMELS PACHER, H. (1972). A c e t y l c h o l i n e s t e r a s e - s t a i n i n g Synaptosomes From Rat Hippocampus: R e l a t i v e Frequency And T e n t a t i v e E s t i m a t i o n s Of I n t e r n a l C o n c e n t r a t i o n Of Free Or L a b i l e Bound A c e t y l c h o l i n e . B r a i n ESSj.* 77* 85-96. KOHAR, M.J., AGHAJANIAN, G.K. AND ROTH, R. H. (1972). T ryptophan H y d r o x y l a s e A c t i v i t y And Synaptosomal Uptake Of S e r o t o n i n I n D i s c r e t e B r a i n Regions A f t e r M i d b r a i n Raphe L e s i o n s ; C o r r e l a t i o n With S e r o t o n i n L e v e l s And H i s t o c h e m i c a l F l u o r e s c e n c e . B r a i n res*.* 44* 165-176. KUHAR, M.J., ROTH, R.H. AND AGHAJANIAN, G.K. (1972). C h o l i n e Uptake I n t o Synaptosomes From The Hippocampus: Re d u c t i o n A f t e r E l e c t r o l y t i c D e s t r u c t i o n Of The M e d i a l S e p t a l N u c l e u s , Fed, P r o c . . 3 1 . 5 1 6 . 381 LAATSCH, R.H. , AND COWAN, W, M. (19 6 6 ) . / E l e c t r o n M i c r o s c o p i c S t u d i e s Of The Dentate Gyrus Of The R a t . , I . Normal S t r u c t u r e With S p e c i a l R e f e r e n c e To S y n a p t i c O r g a n i z a t i o n . J*. Comg. N e u r o l * * 128* 359-396. LANGLEY, J.N. (1905). On The C o n t r a c t i o n Of Muscle, C h i e f l y In R e l a t i o n To P r e s c e n c e Of R e c e p t i v e S u b s t a n c e s . J . P h y s i o l . , 3 3 t 374-413. I E MOAL, M. AND CARDO, B. (1975). Rhythmic Slow Wave A c t i v i t y Recorded I n The V e n t r a l M e s e n c e p h a l i c Tegmentum I n The Rat. J o u { ' e l e c t r o e n c e p h . C l i n . N e u r o p h y s i o l . , 38, «) 139-147. LEWIS, P.R. AND SHUTE, C. C, p. ( 1967) . The C h o l i n e r g i c L i m b i c System: C h o l i n e r g i c R e t i c u l a r System And The S u b f o r n i c a l Organ And S u p r a - o p t i c C r e s t . B r a i n r e s i x 90* 521-540. LIDBRINK, P. (1974). The Effect Of Lesions Of Ascending Noradrenaline Pathways On Sleep And Sakinq In The Rat. Brain res.* 74 r 19-40. LI L E Y , A.W. (1956). The Qu a n t a l Components Of The Mammalian E n d - p l a t e P o t e n t i a l . J * I e u £ G £ h x s i o l * * J33* 571-587. LINDSLEY, D. B. ( 1960) , A t t e n t i o n , C o n s c i o u s n e s s , S l e e p And Wakefulness. I n J . F i e l d ( E d , ) , handbook o f Ekl§.i2,lo.32*. s e c * 1*. n e u r o p h y s i o l o g y . v o l . ; 3* Washington, D . c : Am. P h y s i o l . S o c , Pp. 1553-1593. LINDSLEY, D.B, AND WILSON, C.L. (1975).. B r a i n Stem-h y p o t h a l a m i c Systems I n f l u e n c i n g Hippocampal A c t i v i t y And B e h a v i o u r . , In R. L. , I s a a c s o n And K. H. Pribram ( E d s . ) , t h e hiJBfiocampus* v o l * 2:-n e u r o p h y s i o l o g y and b e h a v i o u r * New York: Plenum, Pp. 2 47-2787 382 LINDV ALL, 0. (1975). M e s e n c e p h a l i c Dopaminergic A f f e r e n t s To The L a t e r a l S e p t a l Nucleus Of The Bat. B r a i n y B e s A j L 87* 89-95. LINDV A LL, 0. AND BJOBKLOND, A. (1974). The O r g a n i z a t i o n Of The Ascending C a t e c h o l a m i n e Neuron Systems In The Bat B r a i n , As R e v e a l e d By The G l y o x y l i c A c i d F l u o r e s c e n c e Method. A c t a p h y s i o l , _ Scan& X l l LOMO, T. (1968). Nature And D i s t r i b u t i o n Of I n h i b i t i o n In A S i m p l e C o r t e x ( d e n t a t e A r e a ) . A c t a p h y s i o l . . S c a n d 1 J t 7 4 x 8A-9A. LOMO, T. , (1971a). P a t t e r n s Of A c t i v a t i o n I n A Monosynaptic C o r t i c a l Pathway: The P e r f o r a n t Path Input To The Dentate Area Of The Hippocampal F o r m a t i o n . Exp. B r a i n r e s ^ x 12*. 18-45. LOMO, T. (1971b). P o t e n t i a t i o n Of Monosynaptic EPSP's In The P e r f o r a n t P a t h - d e n t a t e G r a n u l e C e l l Synapse. Exp. B r a i n r e s . , Ylx 46-63. LONGO, V. G. (1 962) . R a b b i t . b r a i n . r e s e a r c h ^ Amsterdam: E l s e v i e r . LONGO, V.G. (1955) . A c e t y l c h o l i n e , C h o l i n e r g i c Drugs And C o r t i c a l E l e c t r i c a l A c t i v i t y . E x p e r i e n t i a * 11M. 76-78. , LOBENS, S.A. AND GULDBEBG, H.C. (1974).,, B e g i o n a l 5-h y d r o x y t r y p t a m i n e F o l l o w i n g S e l e c t i v e M i d b r a i n Raphe L e s i o n s I n The Rat. B r a i n c e s L i , 7 8 x 45-56. 383 LORENTE DE NO, R. (1 934). Studies On The Structure Of The Cerebral Cortex. I I . Continuation Of The Study Of The Amnionic System. J * Psycho1.neurol. f 46 x 113-177. LYNCH, G. AND COTMAN, C. W. (1975) . The Hippocampus As A Model For Studying Anatomical P l a s t i c i t y In The Adult Brain, In R.L. Isaacson And K. H. Pribram (Eds.) , the hijifiocamBuSi v o l * j j . structure and development* New York: Plenum, Pp. T23-154. LYNCH, G. , DEADwYLER, S. AND COTMAN, C.W., (1973a). Postlesion Axonal Growth Produces Permanent Functional Connections. Science* 180* 1364-1366. LYNCH, G. , MATTHEWS, D. A., MOSKO, S., PARKS, T. AND COTMAN, C. (1972).,, Induced Acetylcholinesterase-rich Layer In Rat Dentate Gyrus Following Entorhinal Lesions. Brain res**-4 2* 311-318. LYNCH, G., MATTHEWS, D. A. , MOSKO, S., PARKS, T. AND COTMAN, C.W. (1972). Induced Acetylcholinesterase-rich Layer In Rate Dentate Gyrus Following Entorhinal Lesions. Bra .in res** 42* 311-318. LYNCH, G. , MOSKO, S., PABKS, T. AND COTMAN, C. . (1973). Relocation And Hyperdevelopmenf Of The Dentate Gyrus Commissural System After Entorhinal Lesions In Immature Rats. Brain res*,* 50^ 17 4-178. LYNCH, G.S., DUNWIDDIE, T.V. AND GRIBKOFF, V. K. (1977). Heterosynaptic Depression: A Post-synaptic Correlate Of Long-term Potentiation. Nature* lond**, 266* 737-73 9. LYNCH, G.S., GRIBKOFF, 7. K. AND DEADWYLER, S. A. (1976) . Long Term Potentiation Is Accompanied By A Reduction In Dendritic Responsiveness To Glutamic Acid. Nature* lond** 263* 151-153. 384 MACADAR, A.W., CHALOPA, L.M. AND LINDSLEY, D.B. (1974). D i f f e r e n t i a t i o n Of B r a i n Stem L o c i Which A f f e c t Hippocampal And N e o c o r t i c a l E l e c t r i c a l A c t i v i t y . Exp. N e u r o l . t 43 x 499-5 14. MAGNI, F. AND WILLIS, W. D. (1 963 ). ,Ide n t i f i c a t i o n Of R e t i c u l a r Formation Neurons By I n t r a c e l l u l a r R e c o r d i n g . A r c h . I t a l x ; B i o l t x 101 x 681-702. ., MANTEGAZZINI, P. AND GLASSES, A. (1960). A c t i o n De La D L - 3 - 4 - d i o x y p h e n i l a l a n i n e (DOPA) E t De La Dopamine Sur L ' a c t i v i t e E l e c t r i g u e Du Chat "ce r v e a u I s o l e " . A.rch x I t a l * B i o l * * . 98* 367-374. MCGEER, E.G. AND MCGEER, P.L. (1976 ). D u p l i c a t i o n Of B i o c h e m i c a l Changes Of H u n t i n g t o n s * s c h o r e a By I n t r a s t i a t a l I n j e c t i o n s Of G l u t a m i c And K a i n i c A c i d s . N a t u r e * 263 x 517-519. MCGEER, E.G., W ADA, J . A. , TER AO, A. AND JUNG, E. (1969). Amine S y n t h e s i s I n V a r i o u s B r a i n Regions W i t h Caudate Or S e p t a l L e s i o n s . EXJG* N e u r o l x x 24 x 277-284. MCGEER, P.L., MCGEER, E.G., SINGH, V.K. AND CHASE, W.H. (1974). C h o l i n e A c e t y l t r a n s f e r a s e L o c a l i z a t i o n I n The C e n t r a l Nervous System By Immunohistochemistry. B r a i n r e s x x 81 x 373-379. MCNAUGHTON, B.L, AND EARNES, C.A. ( 1 9 7 7 ) . , P h y s i o l o g i c a l I d e n t i f i c a t i o n And A n a l y s i s Of Dent a t e G r a n u l e C e l l Responses To S t i m u l a t i o n Of The M e d i a l And L a t e r a l P e r f o r a n t Pathways I n The Ra t , J.. Coinp. N e u r o l x x J76 439-453. MCWILLIAMS, R. AND LYNCH, G. (1978). T e r m i n a l P r o l i f e r a t i o n And S y n a p t o g e n e s i s F o l l o w i n g P a r t a i l D e a f f e r e n t i a t i o n : The R e i n n e r v a t i o n Of The I n n e r M o l e c u l a r Layer Of The Dentate Gyrus F o l l o w i n g Removal Of I t s Commissural I n p u t s . J x £om£ x N e u r o l x x 180 x 581-615. 3 8 5 MCLENNAN, H. AND MILLER, J . J . (1974a). Gawa-A m i n o b u t y r i c A c i d And I n h i b i t i o n I n The S e p t a l N u c l e i Of The fiat. J . P h y s i o l . I L o n d o n l * 237 625-633. MCLENNAN, H. AND MILLER, J . J . (1974b). The Hippocampal C o n t r o l Of Ne u r o n a l D i s c h a r g e s I n The Septum Of The Rat. J . P h j ^ i ^ l . * , 23_2* 607-624. MCLENNAN, H. AND MILLER, J . J . (1 976) . F r e g u e n c y - r e l a t e d I n h i b i t o r y Mechanisms C o n t r o l l i n g R h y t h m i c a l A c t i v i t y I n The S e p t a l Area. J«_ P h y s i o l lLgndoni. i. x2 54 1 L 827-841 . M ELAMED, E. , LAHAV, M. AND ATLAS, D. (1977). B e t a -a d r e n e r g i c R e c e p t o r s I n Rat C e r e b r a l C o r t e x : H i s t o c h e m i c a l L o c a l i z a t i o n By A F l u o r e s c e n t B e t a - b l o c k e r . B r a i n r e s * * 52* 19-36. MELLGREN, S.I. AND SREBRO, B. (1973). Changes I n A c e t y l c h o l i n e s t e r a s e And The D i s t r i b u t i o n Of De g e n e r a t i n g F i b r e s I n The Hippocampal Region A f t e r S e p t a l L e s i o n s I n The Rat. B r a i n r e s . * ~ 75* 172-176. MEYNERT, T. (1872). The b r a i n of mammals..; The New Sydenham S o c i e t y , London. MILLHOUSE, 0.E. (1969). A G o l g i Study Of The Descending M e d i a l F o r e b r a i n Bundle. B r a i n res,.* 1 5 * 341-MONNIER, M. AND ROMANOWSKI, W. (1962)., L es Systemes C h c l i n o c e p t i f s C e r e b r a u x - a c t i o n s De L • a c e t y l c h o l i n e , De L a P h y s o s t i g m i n e , P i l o c a r p i n e E t De GAB A. E l e c t r oenceph. C l i n . Neuro£hysigl** J4* 486-500. MOOBE, S. E. (1977). The A c t i o n s Of Amphetamine On N e u r o t r a n s m i t t e r s : A B r i e f Review. B i o l o g i c a l a s y c h i a t r y * . 1 2 1 3 1 4 5 1 - 4 6 2 . ~*~ 386 MOORE, B.J, (1975). Monoamine Neurons I n n e r v a t i n g The Hippocampal F o r m a t i o n And Septum: O r g a n i z a t i o n And Besponse To I n j u r y . I n R.L, I s a a c s o n And K.H, P r i b r a m (Eds.) , the hippocampus* v o l * . s t r u c t u r e and development* New York: Plenum, Pp. 215-2379 MOORE, R.Y. (1978). C a t e c h o l a m i n e I n n e r v a t i o n Of The Ba s a l F o r e b r a i n . I . The S e p t a l Area. J * Comp* N e u r o l * * llj, 665-684. MOORE, R.Y. AND HALARIS, A. E. (1975). Hippocampal I n n e r v a t i o n By S e r o t o n i n Neurons Of The M i d b r a i n fiaphe I n The Ra t . J * Comp. N e u r o l , 164* 17i1-184. MOORE, B.Y.. AND HELLER, A. {1967) . Monoam i n e L e v e l s And Neu r o n a l D e g e n e r a t i o n I n The Rat B r a i n F o l l o w i n g L a t e r a l Hypothalamic L e s i o n s . J o u r n a l o f pharmacology and e x p e r i m e n t a l t h e r a p e u t i c s * 156* 12-22. MOORE, R. Y. , BJORKLUND, A. AND STENEVI, U. (1971). P l a s t i c Changes I n The A d r e n e r g i c I n n e r v a t i o n Of The Rat S e p t a l Area I n Response To D e n e r v a t i o n . B r a i n r e 3 3 * 13-35, MOREST, D. K. (1961). C o n n e c t i o n s Of The D o r s a l Tegmental Nucleus I n Rat And R a b b i t , J * Anatomy* 95* 229-246. MORUZZI, 6. AND MAGOUN, H.S. ( 1949). B r a i n s t e m R e t i c u l a r F o r m a t i o n And A c t i v a t i o n Of The EEC E l e c t r o e n c e p h * C l i n * l e u r o p ^ y s i o l * * 1 X 455473. MOSKO, S. , LYNCH, G. AND COTMAN, CM., (1973). D i s t r i b u t i o n Of The Septal Projection To The Hippocampal Formation Of The Rat. J,. Comp* Neurol** 152* 163-174. NADEL, L. AND 0* KEEFE, J . (1974). The Hippocampus I n P i e c e s And P a t c h e s : an Essay On Modes Of E x p l a n a t i o n I n P h y s i o l o g i c a l P s y c h o l o g y . , I n : essays on t h e nervous systemj. A f e s t s c h r i f t f o r P r o f e s s o r J*Z t. £onng* R. B e l l a i r s And E.G. Gray ( E d s . ) . O x f o r d : C l a r e n d o n P r e s s , Pp. 3 67-390. 387 NADLER, J.V., COTMAN, C.W, , AND LYNCH, G. S. {1973). Altered D i s t r i b u t i o n Of Choline Acetyltransferase And Acetylcholinesterase A c t i v i t i e s In The Developing Hat Dentate Gyrus Following Entorhinal Lesion, Brain r e s ^ 63* 215-230. NAFSTAD, P.H.J. (1967). An Electron Microscope Study On The Termination Of The Perforant Path Fibers In The Hippocampus And The Fascia Dentate. Z* Zgilfgrsch*.* 76* 532-542, W.J.H. (1956). An Experimental Study Of The Fornix System In The Rat. J * Come* Neurgl* x 19.4* 247-271. W.J.H, (1958). Hippocampal Projections And Related Neural Pathways To The Mid-brain In The Cat. Brain* 81* 319-340. W.J.H. AND KUYPERS, H.G.J.M. (1958). Seme Ascending Pathways In The Brain Stem Reticular Formation. In H. Jasper Et A l (Eds,), reticular, formation of the brain* Toronto: L i t t l e , Brown, Pp. 3-30. NAOTA, W. J.H. AND KUYPERS, H.G. (1958). Some Ascending Pathways In The Brainstem Reticular Formation. In: r e t i c u l a r formation of the brain H. J as per (ed.). Boston: L i t t l e , Brown And Co., Pp. 3-30. O'KEEFE, J, AND DOSTROVSKY, J. (1971). The Hippocampus As A Spa t i a l Map: preliminary Evidence From Unit A c t i v i t y In The Freely Moving Rat. Brain L^XM 34* 171-175. OLMSTEAD, C.E. , AND VILLABLANCA, J. R. (1977). Hippocampal Theta Rhythm P e r s i s t s In The Permanently Isolated Forebrain Of The Cat. Brain res.. 2, 93-100. NAUTA, NAUTA, NAUTA, .3 88 OLNEY, J.W., EHEE, V. AND HO, 0. C. (1974). K a i n i c A c i d : A P o w e r f u l N e u r o t o x i c Analogue Of Glu t a m a t e , B r a i n r e s . , 77. 307-512. PAIVA, T., LOPES DA SILVA, F . f l . , AND MOLLEVANGEH, W. (1976).., M o d u l a t i n g Systems Of Hippocampal EEG. E l e c t r o e n c e j a h . C l i n * Ne u r o p h y s i o l . . , 40* 470-480. PALETTI, C. E. AND CRESWELL, G. (1976). F o r n i x System E f f e r e n t P r o j e c t i o n s I n The S g u i r r e l Monkey: an E x p e r i m e n t a l D e g e n e r a t i o n Study. J * -Comp* N e u r o l * * J75* 101-128. PALKOVITS, H. AND JACOBWITZ, D.M.(1974). Topographic A t l a s Of Catecholamine And A c e t y l c h o l i n e s t e r a s e - c o n t a i n i n q Neurons I n Bat B r a i n . I I . H i n d b r a i n (mesencephalon, Rhombencephalon). J x Comp* N e u r o l . , 157 f 29-42. PALKOVITS, M. AND JACOBWITZ, D.M. (1 974 )., T o p o g r a p h i c A t l a s Of C a t e c h o l a m i n e And A c e t y l c h o l i n e s t e r a s e - c o n t a i n i n g Neurons I n Eat B r a i n . . H i n d b r a i n . J*,Comp. N e u r o l * * 1578 29-42. PAPEZ, J.W. (1937). A Proposed Mechanism Of Emotion. Archj. N e u r o l * P s y c h i i t j . * 38 A 725- 744. PEBKEL, D.H., GEBSTEIN, G.L, AND MOORE, G.P. (1967). N e u r o n a l Spike T r a i n s And S t o c h a s t i c P o i n t P r o c e s s e s . B i o p h y s i c a l j o u r n a l * 1, 3 91-440. PETSCHE, H. , GOGOLAK, G. „ AND VAN ZWIETEN, P. A. (1965). R h y t h m i c i t y Of S e p t a l C e l l D i s c h a r g e s At V a r i o u s L e v e l s Of B e t i c u l a r E x c i t a t i o n . E l e c t r o e n c e p h * C l i n * Neurophysio1** 12* 589-600. PETSCHE, H., GOGOLAK, G., AND VAN ZWIETEN, P. A. (1965). R h y t h m i c i t y Of S e p t a l C e l l D i s c h a r g e s At V a r i o u s L e v e l s Of R e t i c u l a r E x c i t a t i o n . E l e c t r o en ceph. c l i n , N e u r o p h y s i o l . •. 19. 25-33. 389 PETSCHE, H. , STUMPF, CH. AND GOGOLAK, A. (1962). The S i g n i f i c a n c e Of The R a b b i t s ' Septum As A Relay S t a t i o n Between M i d b r a i n And Hippocampus, I , The C o n t r o l Of Hippocampus A r o u s a l A c t i v i t y By Septum C e l l s . S i e c t r o e n c e p h * C l i n * Ne.uroEhy.siol* J4* 201-211." PHILLIS, J.W. (1968). A c e t y l c h o l i n e R e l e a s e From The C e r e b r a l C o r t e x : I t s Role I n C o r t i c a l A r o u s a l . B r a i n r e s * * 7* 378-389. PHILLIS, J . W. , AND KOSTOPODLOS, G.K. { 1977) . Act i v a t i c n Of A N o r a d r e n e r g i c Pathway From The B r a i n s t e m To Rat C e r e b r a l C o r t e x . Gen* Pharmac** 8* 207-211. PICKEL, V.M., SEGAL, M. AND BLOOM, F. E. (1974). A R a d i o a u t o g r a p h i c Study Of The E f f e r e n t Pathways Of The Nucleus Locus C o e u r l e u s . J . ; Comp. M e u r o l * * 1 5 5 * 15-42. POLC, P., AND MONNIER, M. (1970). An A c t i v a t i n g Mechanism In The P o n t o b u l b a r Raphe System Of The R a b b i t . B r a i n r e s l j t 22* 47-61 . POWELL, T.P.S. AND COWAN, W.M. (1955). An E x p e r i m e n t a l Study Of The E f f e r e n t C o n n e c t i o n s Of The Hippocampus. Brain,. 78* 115-132. RAISMAN, G. (1966). The C o n n e c t i o n s Of The Septum. B r a i n * 89* 317-348. RAISMAN, G. , COWAN, W. M. . AND POW ELL, T.P.S. (1965). The E x t r i n s i c A f f e r e n t Commissural And A s s o c i a t i o n F i b e r s Of The Hippocampus. B r a i n * 88*-963-996. RAISMAN, G., COWAN, W. M. AND POWELL, T. P. S. (1966) . The Co n n e x t i o n s Of The Septum, B r a i n * 89* 317-348. RALL, W. (1974) . D e n d r i t i c S p i n e s , S y n a p t i c Potency And Neuronal P l a s t i c i t y . I n : c e l l u l a r leGhanisis s u b s e r v i n g changes- i n - neuronal•••• a c t i v i t y ( E d i t e d By Woody, CH. D. And B r o w n T K7K77~Crow, T.J, And K n i p s e l , J.D,). B r a i n Research I n s t i t u t e , UCLA. 390 HALL, I. AND RIHZEL, J. (1973). Branch Input Resistance And Steady Attenuation For Input To One Branch Of A Dendritic Neuron Model. Biophysical i9.EEQ.al* 13* 64 8-6 88. RANCK, J.B. (1975). Behavioral Correlates And Fi r i n g Repertoires Of Neurons In The Dorsal Hippocampal Formation And Septum Of Unrestrained Rats. In tjhg hippocamgus* vol* 2 Ed, Isaacson, R.L. And Pribram, K.H..New York: Plenum Press. RANCK, J.B., JR. (1973). Studies On Single Neurons In Dorsal Hippocampal Formation And Septum In Unrestrained Rats. Part 1. Behavioural Correlates And F i r i n g Repertoires. Exp* Neji£2li.ji HIJL 462-531, RICHARDSON, J. S. AND JACOBOWITZ, D.M. (1973) . Depletion Of Brain Norepinephrine By Intraventricular Injection Of 6-hydroxydopamine: A Biochemical, Histochemical, And Behavioural Study In The Rat. Brain res** 5.8* 117-133, RINALDI, F. AND HIMWICH, H.E. ( 1955a), Alerting Responses And The Actions Of Atropine And Cholinergic Drugs. Arch* Neurol* Psyehiat** 73*-3 87-395. RINALDI, F. AND HIMWICH, H.E, {1955b). Cholinergic Mechanism Involved In Function Of Mesodiencephalic Activating System. Arch. Neurol* gsychiat** 73* 396-402. ROBINSON, T. E. ( 1978), Brainstem Influences On Hippocampal And Neocortical Slow Wave Activation During Waking Behavior And Sleep. Doctoral Thesis, University Of Western Ontario. ROBINSON, T. E. , VANDERWOLF, C. H. AND PAPPAS, B.A. (1977). Are The Dorsal Noradrenergic Bundle Projections From The Locus Coeruleus Important For Neocortical Or Hippocampal Activation? Brain res. , 1 32 . 75-98. 391 BODBIQUES, B., BOJAS-RAMIBEZ, J.A. AND DRUCKER-COLIN, R.R. (1973). Serotonin-like Actions Of Quipazine On The Central Nervous System. Eur. Is. E h a i f i a c o l i * 24*164-171. RGMMELS PACH EB, H. , GOLDBERG, A.H. AND KUHAR, M.J. (1974). Action Of Hemicholinium-3 On Cholinergic Nerve Terminals After Alteration Of Neuronal Impulse Flow. Neuropharmacolj.j, 13*-1015-1023. BOSE, A.M., HATTOBI, T. AND FIBIGER, H.C. (1976). Analysis Of The Septo-hippocampal Pathway By Light And Electron Microscopic Autoradiography. l£§JLB £§SAX 10 8 x 170-174. ROSE, J.E. (1942). The Ontogenetic Development Of The Rabbit«s Diencephalon, J * Comja* Neurol** 77* 61-129. ROUTTENBEBG, A. AND KRA MIS, R. C. (1 963 ). Hippocampal Correlates Of Adversive Midbrain Stimulation. Science, 160 1363-1365. SACHS, C. AND JONSSON, G. (1975). Ef f e c t s Of 6-hydroxydopamine On Central Noradrenaline Neurones During Ontogeny. Brain res*,* 99* 277-291. SALMOIBAGHI, G.C. AND STEFANIS, C.N. (1965)., Patterns Of Central Neurons Responses To Suspected Transmitters. Arch. I t a l * B i o l * * 103* 705-724. SCHEIBEL, H.E. ..AND SCHEIBEL, A. B. , (1953 ). Structural Substrates For Integrative Patterns In The Brain Stem Reticular Core. In H. h. Jasper Et Al. (Eds.), r e t i c u l a r formation of the brain*-New York: L i t t l e , Brown, Pp. 31-5 5. SCHUBERT, P., LEE, K., WEST, M,, DEADWYLER, S.A. AND LYNCH, G. (1976). Stimulation-dependent Release Of 3Hadenosine Derivatives From Central Axon Terminals To Target Neurones. Nature* lond**-260* 541-542. 392 SCHWARTZKROIN, P.Z. AND BESTEB, K. .(1975). L o n g - l a s t i n g F a c i l i t a t i o n Of A S y n a p t i c P o t e n t i a l F o l l o w i n g T e t a n i z a t i o n In The I n Vitr.p Hippocampal S l i c e , B r a i n r e s * * 8^* 107-1197 SEGAL, M. (1975). P h y s i o l o g i c a l And P h a r m a c o l o g i c a l E v i d e n c e F o r A S e r o t o n e r g i c P r o j e c t i o n To The Hippocampus. B r a i n r e s * * 94* 115-131. SEGAL, M. (1976)., B r a i n Stem A f f e r e n t s To The Rat M e d i a l Septum. J . P h y s i o l . , 261. 617-631. SEGAL, M.. AND BLOOM, F.E. (1974a). The A c t i o n Of N o r e p i n e p h r i n e I n The Rat Hippocampus. I , I o n t o p h o r e t i c S t u d i e s . B r a i n res*.*, 72* 79-97. SEGAL, M. , AND BLOOM, F.E. (1974b). The A c t i o n Of N o r e p i n e p h r i n e I n The Rat Hippocampus, I I , A c t i v a t i o n Of The I n p u t Pathway, B r a i n r e s * * 72* 99-114. M, . AND BLOOM, F. E. , N o r e p i n p h r i n e In The Hippocampal C e l l u l a r C o e r u l e u s S t i m u l a t i o n r e s * * 107^ 499-511. 976a). The A c t i o n Of Bat Hippocampus. I I I . Responses To Locus I n The Awake B a t . B r a i n (1 SEGAL, H. .. AND BLOOM, F.E. (1976b). The A c t i o n Of N o r e p i n e p h r i n e I n The Rat Hippocampus. IV. The E f f e c t s Of Locus C o e r u l e u s S t i m u l a t i o n On Evoked Hippocampal U n i t A c t i v i t y . B r a i n r e s * * 107* 513-525. SEGAL, M. AND LANDIS, S. (1974). A f f e r e n t s To The Hippocampus Of The Rat S t u d i e d With The Method Of R e t r o g r a d e T r a n s p o r t Of H o r s e r a d i s h P e r o x i d a s e . Bra^n r e s * * 78* 1-15. SHEPHERD, G. M, (1974) . , The. s y n a p t i c o r g a n i z a t i o n a f t h e £rain*New York: O x f o r d U n i v e r s i t y P r e s s , 393 SHERRINGTON, C.A. (1906). The Integrative Action Of The Nervous System. New Haven: Yale University Press. SHUTE, C C D . AND LEWS, P.R. (1966). The Ascending Cholinergic Reticular System: Neocortical, Olfactory And Subcortical Projections. Brain* 20* 4 97-520. SHUTE, C C D . , AND LEWIS, P.R. (1967). The Ascending Cholinergic Reticular System: Neocortical, Olfactory And Subcortical Projections. Brain £§.§*.«. 20*. 497-520. SIEGEL, A. AND TASSONI, J. P. (1971). D i f f e r e n t i a l Efferent Projections From The Ventral And Dorsal Hippocampus Of The Cat. Brain behay. Eygl^.* 4* 185-200. SIMPSON, E.A. (1952), The Efferent Fibers Of The Hippocampus In The Monkey. J * - Neurology, neurosurgery and £sychiatry*. .15* 79-92. SMITH, CM. (1972). The Release Of Acetylcholine From Rabbit Hippocampus. Br* J * Pharmacol.* 45* 172p. SMITH, CM. (1974). Acetylcholine Release From The Cholinergic Septohippocampal Pattern, L i f e -s c i j * - 20* 2159-2166. SPENCER, H.J. , GRIBKOFF, V.K. , COT MAN, C. 9. :i, AND LYNCH, G.S. (1976). GDEE Antagonism Of Iontophoretic Amino Acid Excitations In The Intact Hippocampus And In The Hippocampal S l i c e Preparation. Brain res..* 195* 471 -481. SREBRO, B. AND MELLGREN, S.I. (1974). Changes In Post-natal Development Of Acetylcholinesterase In The Hippocampal Region After Early Septal Lesions In The Rat. Brain res** 7 9* 119-131. 394 STEFANIS, C. (1964). Hippocampal Neurons: Their Responsiveness To Microelectrophoretically Administered Endogenous Amines. Pharmacologist^ 6 171., STEWARD, 0,, COTMAN, C.W. AND LYNCH, G. (1973)., Re-establishment Of El e c t r o p h y s i o l o g i c a l l y Functional Entorhinal C o r t i c a l Input To The Dentate Gyrus Deafferented By I p s i l a t e r a l Entorhinal Lesions: Innervation By The Contralateral Entorhinal Cortex. Exp.* Brain r e s t 4 18* 376-414. STEWARD, 0,, COTMAN, C. W, AND LYNCH, G. ( 1974). Growth Of A New Fibre Projection In The Brain Of Adult Rats: Re-innervation Of The Dentate Gyrus By The Contralateral Entorhinal Cortex Following I p s i l a t e r a l Entorhinal Lesions, Exp*. grain 20 x 45-66. STEWARD, 0,, WHITE, F. AND COTMAN, C.W. (1977). Potentiation Of The Excitatory Synaptic Action Of Commissural, Associational And Entorhinal Afferents To Dentate Granule Cells, Brain res** J 3 4 * 551-560. STEWARD, 0., WHITE, W.F., COTMAN, C, W. AND LYNCH, G. (1976). Potentiation Of Excitatory Synaptic Transmission In The Normal And In The Reinnervated Dentate Gyrus Of The Rat. Exp,, Brain r e s ^ 26* 423-441. STORM-MATHISEN, J. (1970). Quantitative Histochemistry Of Acetylcholinesterase In Rat Hippocampal Region Correlated To Histochemical Staining. J * Neurochem. f J7j, 739-750. STORM-MATHISEN, J. {1972) . Glutamate Decarboxylase In The Rat Hippocampal Region After Lesions Of The Afferent Fibre Systems. Evidence That The Enzyme Is Localized In I n t r i n s i c Neurones. Brain res.. 40. 215-235. 3 95 STRAUGHAN, D. H. (1975)., Neurotransmitters And The Hippocampus. In R. L. Isaacson And K. H. Pribram (Eds,)., the hipfiocampus* v o l * structure and development, New York: Plenum, P.p. 239-268. STUMPF, CH. ( 1 9 6 5 ) . Drug Action On The E l e c t r i c a l A c t i v i t y Of The Hippocampus. International review of neurobiology, 8* 77- 138. STUMPF, CH, , (1965b), The Fast Component In The E l e c t r i c a l A c t i v i t y Of The Rabbit's Hippocampus, . Electroenceph,. C l i n . IgMEaEL&ZsiaijLA J h 8 * 477-486.~ SWANSON, L.W. AND COWAN, W. M. (1975). Hippocampo-hypothalamic Connections: Origin In Subicular Cortex, Not Ammon's Horn. Science* 189* 303-304. SWANSON, L.W. AND COWAN, W. M. (1977). An Autoradiographic Study Of The Organization Of The Efferent Connections Of The Hippocampal Formation In The Rat. J * Cpmp* Neurol** 172^ 49-84. SWANSON, L.W. .. AND HARTMAN, B. K. (1 975 ) . The Central Adrenergic System. An Immunofluorescence Study Of The Location Of C e l l Bodies And Their Efferent Connections In The Rat U t i l i z i n g Dopamine-b-hydroxylase As A Marker. J t Com p. neurol. , 1.63 467-506, THOMAS, B.C. AND WILSON, V. S (1965)., Precise Localization Of Renshaw C e l l s With A New Marking Technique. Nature iLgnd . L * 22.6*. 211-213. TOMBOL, T. AND PETSCHE, H. (1969). The H i s t o l o g i c a l Organization Of The Pacemaker For The Hippocampal Theta Rhythm In The Rabbit. Brain res.. 12. 414-426. 396 TOBII, S.. (1961). Two Types Of P a t t e r n Of Hippocampal E l e c t r i c a l A c t i v i t y Induced By S t i m u l a t i o n Of Hypothalamus And S u r r o u n d i n g P a r t s Of R a b b i t ' s B r a i n . . Jap.* J * P h y s i o l * * 11* 1 47- 1 57. TORII, S. , AND WIKLEB, A. ..(1966) .. E f f e c t s Of A t r o p i n e On E l e c t r i c a l A c t i v i t y Of Hippocampus And C e r e b r a l C o r t e x In Cat. P s x c h o p h a r m a c o l A J , 9* 189-204, TRANZER, J.P. UND THOENIN, H. (1967). O l t r a m o r p h o l o g i s c k e Veranderungen Der Sympatheschen Nrvenendigungen Der K a t z e Nach Vorbehandlung M i t 5-und 6-hydroxydopamine. Naunyn-Schmiedeberg's a r c h * * 257* 343, ONGERSTEDT, U. (1971). S t e r e o t a x i c flapping Of The Monoamine Pathways I n The Bat B r a i n . Acta-p.hysiolj_ Scan d. , s u p p l . 367 f c 1-48. VAL EN STEIN, E. X, /AND NAOTA, W.J.H. (1959) . A Comparison Of The D i s t r i b u t i o n Of The F o r n i x System I n The Ra t , Guinea P i g , C a t , And Monkey. J * --Co*p*~ N e u r g l ^ 1_13* 33 7-363. VALVERDE, F. (1962). R e t i c u l a r F o rmation Of The A l b i n o Rat's B r a i n Stem, C y t o a r c h i t e c t u r e And C o r t i c o f u g a l C o n n e c t i o n s . J . Comp. > Neu r o l * * -118* 25-54. VAN DEB WOLF, C H . (1969). Hippocampal E l e c t r i c a l A c t i v i t y And V o l u n t a r y Movement I n The Rat. I l e c t r o e n c e p h * C l i n * Neuroph y s i g l * . * 2 6 x 407-4 18. VANDERWOLF, C H . (1971) . L i m b i c - d i e n c e p h a l i c Mechanisms Of V o l u n t a r y Movement. P s y c h o l * Rev** 78* 83-113. VANDERWOLF, C H . (1975). N e o c o r t i c a l And Hippocampal A c t i v a t i o n In R e l a t i o n To Be h a v i o u r : E f f e c t s Of A t r o p i n e , E s e r i n e , P h e n o t h i a z i n e s And Amphetamine. J * Comp,* P h y s i o l * , P s y c h o l * * 88* 300-323. ~ ~~ 3 9 7 VANDERWOLF, C H. , KRAMIS, R . , GILLESPIE, L. A. AND BLAND, B.H. (1975) . Hippocampal Rhythmical Slow Ac t i v i t y And Neocortical Low Voltage Fast A c t i v i t y : Relations To Behavior. In: the l i i f i v o l . - 2 R.L. Isaacson And K.H. Pribram (Eds.) . New York: Plenum, Pp. 10 1- 128. VINOGRADOV A, 0. S. { 1975) . Functional Organization Of The Limbic System In The Process Of Regisy t r a t ion Of Information: Facts And Hypothesis. In: the hippocampus.* vol. 2 Ed. Isaacson, R.L. And Pribram, K.H, New York: Plenum Press. 3-70. HANG, R.Y. AND AGHAJ AN IAN, G.K. (1977) . Antidromically I d e n t i f i e d Serotonergic Neurons In The Rat Midbrain Raphe: Evidence For C o l l a t e r a l I n h i b i t i o n . Brain res*.* 132* 186-193. WHISHAW, I.Q. AND VANDERWOLF, CH. (1973). Hippocampal EEG And Behaviour: Changes In Amplitude And Freguency Of BSA (theta Rhythm) Associated j * . Behav* vBiol*^, 8* 461-484, WILSON, C.L,, MOTTER, B.C. AND LINDSLEY, D.B. (1976). Influences Of Hypothalamic Stimulation Upon Septal And Hippocampal E l e c t r i c a l A c t i v i t y In The Cat, Brain res** 107* 55-68, WINSON, J . (1972). Interspecies Differences In The Occurrence Of Theta. Behav * B i o l * * 7* 4 7 9- 4 8 7. WINSON, J. (1974) . Patterns Of Hippocampal Theta Rhythm In The Freely Moving Rat. E l e c t r o NeurpphYSiol. , 36* 291-301. WINSON, J. (1975). The 0 Mode Of Hippocampal Function. In Robert L. Isaacson And K.H. Pribram (Eds,), the hippocampus, £21*. 2.1 neurop h ys i o l ogy and ogkayiour* New York: Pienum, Pp. 169-183. 3 9 8 WINSON, J. (1976a) Hippocampal Theta Rhythm. I. Depth P r o f i l e s In The Curarized Rat. Brain r e s * * J.03* 57-70. WINSON, J . (1976b) Hippocampal Theta Rhythm. II. , Depth P r o f i l e s In The Freely Moving Rabbit. Brain £es** JO3* 71-79. WINSON, J. (1978) . Loss Of Hippocampal Theta Rhythm Results In Spa t i a l Memory D e f i c i t In The Rat. Sci en c e*- 20 J* 160-163. WINSON, J. AND ABZUG, C. ( 1977). Gating Of Neuronal Transmission In The Hippocampus: Efficacy Of Transmission Varies With Behavioral State. Science. 196, 1223-1225. WINSON, J. AND ABZUG, C. (1978) . Neuronal Transmission Through Hippocampal Pathways Dependent On Behavior. J * Neurophysiol,.* 4.1* 716-732. WOLF, G. AND SOTIN, J . (1966). Fiber Degeneration After Lateral Hypothalamic Lesions In The Rat. J * Comp* Neurol** 127^ 137- 156. YOKOTA, T. AND FUJIMORI, B. (1964). Effects Of Brain Stem Stimulation Upon Hippocampal E l e c t r i c a l A c t i v i t y , Somatomotor Reflexes And Autonomic Functions. E l e c t r oenceph. C l i n . Neuroghysiol**-16 x 375-382. YOUNG, M. W. ( 1 9 3 6 ) . The Nucl e a r P a t t e r n And F i b e r C o n n e c t i o n s Of The N o n - c o r t i c a l C e n t e r s Of The Te l e n c e p h a l o n Of The R a b b i t (Lepuc C u n i c u l u s ) . J J - £21£a. N e u r o l * * 6 5 * 2 9 5 - 4 0 1 . ZIMMER, J. (1970)., I p s i l a t e r a l Afferents To The Commissural Zone Of The Fascia Dentata Demonstrated In Decommissurated Rats By S i l v e r Impregnation..J* ;Comp.* :Neurol** .142* 393-416. 399 ZIMMEH, J, (1974). Extended Commissural And I p s i l a t e r a l Projections In Postnatally De-entorhina ted Hippocampus And Fascia Dentata Demonstrated In Hats By S i l v e r Impregnation, Brain res** 72* 293-31 1. A. Publications BIBLIOGRAPHY S.Y. Assaf 1. Assaf, S.Y. and J . J . M i l l e r (1977a). Excitatory action of the mesolimbic dopamine system on septal neurones. Brain Research, 129: 353-360. 2. Assaf, S.Y. and J . J . M i l l e r (1978a). The r o l e of a raphe-serotonin system i n the control of septal unit a c t i v i t y and hippo-campal desynchronization. Neuroscience, 3_: 539-550. 3. Assaf, S.Y. and J . J . M i l l e r (1978b). Neuronal transmission i n the dentate gyrus: r o l e of i n h i b i t o r y mechanisms. Brain Research, 151, 587-592. 4. Assaf, S.Y. and G. Mogenson (1976). Evidence that the preoptic region i s a receptive s i t e f o r the dipsogenic e f f e c t s of angiotensin I I . Pharmacology Biochemistry and Behaviour, _5: 687-699. 5. Kucharczyk, J . , S.Y. Assaf and G.J. Mogenson (1976). D i f f e r e n t i a l e f f e c t s of b r a i n lesions on t h i r s t induced by the administration of angiotensin II to the preoptic region, subfornical organ and anterior t h i r d v e n t r i c l e . . Brain Research, 108: 327-337. 6. Mogenson, G.J., J. Kucharczyk and S.Y. Assaf (1976). Evidence for multiple receptors and neuronal pathways which subserve water intake i n i t i a t e d by angiotensin I I . In: Central actions of  angiotensin and r e l a t e d hormones. J.P. Buckley and C. F e r r a r i o (eds). Pergamon Press, New York 1976. 7. Mogenson, G.J., S.Y. Assaf, J. Kucharczyk and A.E. Abdelaal (1974). E f f e c t of ablation of the subfornical organ on water intake e l i c i t e d by systemically administered angiotensin I I . Canadian Journal of Physiology and Pharmacology, 52(6) : 1217-1220. B. Abstracts and Personal Communications 1. Assaf, S.Y. and G.J. Mogenson (1975). Evidence that angiotensin II acts on the preoptic region to e l i c i t water intake. Proc. Can. Fed. B i o l . Soc, 18(104) abstract 416. 2. Assaf, S.Y. and J . J . M i l l e r (1976). E x c i t a t i o n of l a t e r a l septal neurones following stimulation of the A-^ Q dopaminergic system. Neuroscience Abstracts, v o l . 11(1): 677. 3. Assaf, S.Y. and J . J . M i l l e r (1977b). Ascending serotenergic systems c o n t r o l l i n g rhythmical a c t i v i t y i n the septal area of the r a t . Neuroscience Abstracts v o l . I l l : 765. 4. Assaf, S.Y. and J . J . M i l l e r (1978c). Inhibitory mechanisms modi-fyi n g neuronal transmission i n the dentate gyrus of the r a t . Canada Physiology, 9(1): 18. BIBLIOGRAPHY (Cont.) S.Y. Assaf Assaf, S.Y. and J . J . M i l l e r (1978d). A comparison of the e f f e c t s of e l e c t r o l y t i c and k a i n i c acid-induced lesions of the septal area on hippocampal e l e c t r i c a l a c t i v i t y . Proc. Can. Fed. B i o l . Soc., 21: 223. 

Cite

Citation Scheme:

        

Citations by CSL (citeproc-js)

Usage Statistics

Share

Embed

Customize your widget with the following options, then copy and paste the code below into the HTML of your page to embed this item in your website.
                        
                            <div id="ubcOpenCollectionsWidgetDisplay">
                            <script id="ubcOpenCollectionsWidget"
                            src="{[{embed.src}]}"
                            data-item="{[{embed.item}]}"
                            data-collection="{[{embed.collection}]}"
                            data-metadata="{[{embed.showMetadata}]}"
                            data-width="{[{embed.width}]}"
                            data-media="{[{embed.selectedMedia}]}"
                            async >
                            </script>
                            </div>
                        
                    
IIIF logo Our image viewer uses the IIIF 2.0 standard. To load this item in other compatible viewers, use this url:
https://iiif.library.ubc.ca/presentation/dsp.831.1-0094477/manifest

Comment

Related Items