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The effects of excitatory amino acids and their antagonists on hippocampal electrophysiology Kehl, Steven James 1984

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THE E F F E C T S OF E X C I T A T O R Y AMINO A C I D S T H E I R A N T A G O N I S T S ON  HIPPOCAMPAL  AND  ELECTROPHYSIOLOGY  By Steven B.Sc,  James K e h l  The U n i v e r s i t y  of Waterloo,  1976  M . S c , The U n i v e r s i t y  of Waterloo,  1979  A T H E S I S S U B M I T T E D I N P A R T I A L F U L F I L L M E N T OF THE R E Q U I R E M E N T S DOCTOR OF  FOR THE DEGREE  OF  PHILOSOPHY in  THE F A C U L T Y OF GRADUATE S T U D I E S (Department  We  accept to  this  of Physiology)  thesis  the required  as  standard  THE U N I V E R S I T Y OF B R I T I S H May ©  Steven  conforming  COLUMBIA  1984  James K e h l , 1984  In p r e s e n t i n g  t h i s t h e s i s i n p a r t i a l f u l f i l m e n t of  requirements f o r an advanced degree a t the  the  University  o f B r i t i s h Columbia, I agree t h a t the L i b r a r y s h a l l make it  f r e e l y a v a i l a b l e f o r reference  and  study.  I  further  agree t h a t p e r m i s s i o n f o r e x t e n s i v e copying o f t h i s t h e s i s f o r s c h o l a r l y purposes may  be granted by  department o r by h i s or her  the head o f  representatives.  my  It i s  understood t h a t copying or p u b l i c a t i o n of t h i s t h e s i s f o r f i n a n c i a l gain  s h a l l not be  allowed without my  permission.  Department o f P h y s i o l o g y The U n i v e r s i t y o f B r i t i s h 1956 Main Mall Vancouver, Canada V6T 1Y3 Date  A p r i l 30,  Columbia  1984  written  i i  Abstract  The e f f e c t s o f a c i d i c amino a c i d s and t h e i r a n t a g o n i s t s on h i p p o c a m p a l e l e c t r o p h y s i o l o g y  The  role  transmission properties using  of  excitatory  and  were  the  amino  modulation  examined  conventional  acidic  in  the  of  acids  and  synaptic  electrophysiological  hippocampal  extracellular  in  slice  in  intracellular  vitro  recording  techniques. Iontophoresis potent  and  kainate  and  causing  cells  contrast  specific  to  to  and to  which (DGG)  bursts  sustained  (APV) was  of  firing  selectively  attributed  inhibited  L-aspartate; quisqualate  diethyl  in  at and  ester,  a  cell  higher  observed  acid-mediated  the  firing dose  the  also  (NMA),  unique  discharges the  in in  other  excitations,  an  D-isomer;^-D-glutamy1  i t also some  (APB)  more  DL-2-amino-5-phos-  elicited  to  was  with  NMA  NMA,  inhibited  kainate responses  L-glutamate  dicarboxylate  variously  had  by  extent.  (+)-cis-2,3-piperidine  e x c i t a t i o n s but  NMA  repetitive  blocked  to  L-glutamate  DL-2-amino-4-phosphonobutyrate  and  N-methy1-DL-aspartate  Iontophoretically-applied  phonovalerate  glycine  L-aspartate  e x c i t e d CAl neurons;  fire  the  L-glutamate, agonists  quisqualate  compounds.  effect  of  affected  complicating  and amino  secondary  actions. The  effects  of  iontophoresis  to the stratum  radiatum  in  iii  CA1  of  excitatory  evaluated  on  Schaffer  the  amino field  acids EPSP  and  evoked  collateral-commissural agonists  as  depressed  EPSPs.  Kainate-induced  rapidly  the  reversible  amplitude. EPSP  period  but  APV  transmission. of  whilst  having  -evoked in  the  properties  the  investigated. a  large  and  spike  (PS)  amplitudes  of  unchanged.  potentiated appeared  a  Schaffer  In  spontaneously also  NMA  with  basis  actions In  CAl  of  the  an  EPSP  change  CA3  fibre  of  little  or  no  antagonist fibre  quisqualate  may  The  play  type  receptors  the  on  is  and  of  the  decline  of  neuro-  processes  gyrus  v o l l e y s and  and  role  discussed.  synaptic  dentate  lower  a  interaction  i t s convulsant  region  the  synaptic  presynaptic  or  LTP.  associated  excitations and  increased  had  long-lasting increase and  the  were  receptors  kainate  and  of  collateral-commissural-  these for  the  considerably  synaptic  kainate  of  L-aspartate  d i d not  the  were  depressions an  APB  on  underlying  presynaptic  synaptic  Kainate  in  L-aspartate the  of  effect  that  processes  and  by  effective  transmission;  determine  population  were  an  was  involved  NMA  t o NMA,  DGG  be  neuronal  followed  isomers  may  and  depressions  optical  concluded  Each  t e r m p o t e n t i a t i o n (LTP)  detectable  synaptic  produced  the  no  pathways.  L-glutamate  responses  long  is  To  were  often  It  L-glutamate  toxic  The  EPSPs.  volley. receptor  and  block  as  antagonists  stimulation  e j e c t i o n whereas  abolished  could  effect  of  by  (SC)  selective  outlasted  well  some  kainate  orthodromic of  the  EPSP;  antidromic  doses  of  PSs  kainate  epileptiform  firing  following electrical stimulation.  reduced  GABA-mediated  inhibitions  and  iv  IPSPs  in  basis  of  but  CAl  the  increased  perhaps  to  for  of In  possible  coupling a  endogenous  unaffected CA3  or  folate  of  to  excitatory of  action.  of  kainate  dentate  orthodromic  of  PSs  and  underlie acids of  the could  kainate.  somata  related as  the  The  potentiating  and  are  discussed  the  kainic  whilst  antidromic  potentiated  and  suggested  perfusion  the  PSs  the  in  neurotoxicity.  gyrus  activator  depressed;  and  to  doses  is  on  non-GABAergic,  amino  higher  dendrites  which,  be  appear  depolarization  properties  the  not  actions  potentiating  and  inhibition  appears  did  its epileptogenicity CAl  an  here  between  dendritic  the  potentiation  In  CA3  Other  depressant  disinhibitory context  reported  excitability.  only  basis  in  disinhibitory action  increased  An  abolished  studies  this  mimic  and  of  acid,  a  receptor, caused  a  the  EPSP  responses  caused  folic  both  were  was  either  unchanged.  spontaneous  and 2 +  evoked  epileptiform  dependent shifts folate CAl  and  was  indicating caused  and  never  implications possible discussed.  no  of  discharges. triggered their  directly  homologies  of  by  data  origin.  change  of  depressed for  receptors  firing  paroxysmal  synaptic  measurable  these  Burst  folate for  Unlike  kainate,  inhibition  activity.  neurotoxicity kainate  Ca  depolarization  recurrent evoked  was  and  in The  and  folate  the are  V  T A B L E OF  LIST  OF T A B L E S  LIST  OF  CONTENTS  v i i  FIGURES  .  v i i i - x i i i  ACKNOWLEDGEMENTS Chapter 1. GENERAL 2.  xiv  INTRODUCTION  1  P R E P A R A T I O N AND M A I N T E N A N C E OF THE H I P P O C A M P A L S L I C E I N V I T R O AND C H A R A C T E R I Z A T I O N OF EVOKED RESPONSES Introduction General Methods I s o l a t i o n , c u t t i n g and maintenance hippocampal t i s s u e  8 .8 9  of 9  Preparation of iontophoretic electrodes . . . . S t i m u l a t i o n and r e c o r d i n g of f i e l d responses . . I n t r a c e l l u l a r recording Results C h a r a c t e r i z a t i o n of f i e l d responses CAl Dentate gyrus CA3 Paired pulse stimulation High frequency s t i m u l a t i o n Effects of e x t r a c e l l u l a r i o n concentrations on evoked a c t i v i t y i n CA3 . . ^ . . . . . . . . . Low [ C a _ ] , i n c r e a s e d [Mg ] Low [ C I ] ° . ? I n c r e a s e d [K ] I n t r a c e l l u l a r r e c o r d i n g o f CA3 r e s p o n s e s . . . . Discussion +  3.  POSTACTIVATION Introduction Methods Results Discussion  INHIBITION  I N THE  CA3 R E G I O N  . . . .  14 14 17 18 19 19 22 24 27 29 31 31 34 41 41 46 56  .  56 57 58 68  vi  4.  THE E F F E C T S OF A N T A G O N I S T S ON AMINO A C I D SCHAFFER-COLLATERAL MEDIATED EXCITATION NEURONS  AND OF C A l 75  Introduction 75 L-glutamate and L - a s p a r t a t e as neurotransmitters : neurochemical evidence 76 L-glutamate and L - a s p a r t a t e as neurot r a n s m i t t e r s t 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 . . 80 Methods 86 Results 88 E f f e c t s of e x c i t a t o r y amino a c i d a g o n i s t s and a n t a g o n i s t s o n s i n g l e u n i t a c t i v i t y . . . 88 E f f e c t s o f e x c i t a t o r y amino a c i d a g o n i s t s and a n t a g o n i s t s on s y n a p t i c a c t i v i t y 100 Discussion I l l 5.  A C T I O N S OF K A I N I C A C I D ON S Y N A P T I C THE H I P P O C A M P A L S L I C E  PROCESSES I N 125  Introduction Methods Results E x t r a c e l l u l a r l y recorded e f f e c t s of kainate on e v o k e d and s p o n t a n e o u s a c t i v i t y CAl CA3 Dentate gyrus E f f e c t s o f o t h e r amino a c i d s i n C A l Kainate's mechanism(s) of a c t i o n CAl-Extracellular analysis of kainateinduced changes o f r e c u r r e n t inhibition . . C A l - I n t r a c e l l u l a r analysis of the actions of k a i n a t e on C A l p y r a m i d a l c e l l s CA3 Dentate gyrus Discussion C o r r e l a t i o n between e l e c t r o p h y s i o l o g i c a l and neurotoxic actions of kainate Mechanism(s) of kainate's e p i l e p t o g e n i c i t y . . . 6.  F O L I C A C I D - I N D U C E D CHANGES OF NEURONAL A C T I V I T Y  125 133 134 134 134 141 147 149 151 153 155 160 167 170 175 177  . . 180  Introduction 180 Methods 183 Results 183 E f f e c t s o f f o l a t e i n C A l a n d t h e d e n t a t e g y r u s . 183 E f f e c t s i n CA3 187 A c t i o n s o f f o l a t e on r e c u r r e n t inhibition . . . 197 Discussion 200 Mechanism o f a c t i o n 202 REFERENCES  208 /  APPENDIX  I  , . 235  vii  L I S T OF Table  I  Actions  of  excitatory quisqualate,  amino  TABLES acid  responses  antagonists to  L-glutamate,  NMA,  on  kainate,  L-aspartate  ACh  Table  II  evoked  Table I I I  and ,  Changes o f t h e S c h a f f e r  tions  the  EPSP  following  collateral-commissuraliontophoretic  applica  of e x c i t a t o r y amino a c i d s  Effects  of a n t a g o n i s t s  commissural-evoked  EPSPs  92  on S c h a f f e r  101  collateral106  viii  LIST Figure  2.1  A  schematic  constant with Figure  2.2  ular Figure  2.3  flow  2.7  2.8  Some  2.9  intracell-  Changes  in  evoked  typical  ex23  region  recording  and  typical  elecextra-  responses  25  of  the  mossy  fibre-  i n CA3  fimbrial  28 and  mossy  fibre-evoked  following tetanic stimulation  actions  of low  activity  Effects  and  elec-  responses  i n t h e CA3  response  extra-  recording  s t i m u l a t i n g and  evoked  typical  elec-  20  gyrus  characteristics  evoked Figure  evoked  P o s i t i o n s of  The  for  responses  i n the dentate  responses Figure  devices  and  and  P o s i t i o n s o f s t i m u l a t i n g and  evoked •Figure  output  13  16  evoked  cellular 2.6  fluid  of e x t r a c e l l u l a r  i n the CAl region  trodes  Figure  and  slices  P o s i t i o n s o f s t i m u l a t i n g and r e c o r d i n g  tracellular 2.5  of hippocampal  cerebrospinal input  f o r the  responses  trodes  Figure  of  recording  cellular 2.4  perfusion  artificial  trodes  Figure  FIGURES  o u t l i n e of the apparatus  Arrangement the  OF  of  low  activity  [Ca  2+  ]/increased  [Mg  30 2+  ] on  i n CA3 [CI ] i n CA3  33 on  spontaneous  and 36  ix  Figure  2.10  In  the  CA3  region  amplitudes [Ca Figure  2.11  2+  2.12  of  Epileptiform to  [Mg  activity  2+  low  spike  [Cl ]  and  ]  38  spikes  synaptic  increased  population  containing  population  absence  sure  ACSF  increased  Spontaneous the  Figure  ] and  in  changes of  in  field  CA3  transmission i n CA3  in 40  following  expo-  [K ]  42  +  o Figure  2.13  Correlation evoked  between  fimbrial  extracellular  fields  and  mossy  and  fibre-  intracellular  potentials  Figure  3.1  45  Homo-  and  heterosynaptically-evoked  tion  of  fimbrial  orthodromic  inhibi-  population  spikes Figure  3.2  The uli  59  actions on  mossy  lation Figure  3.3  Chart to  of  mossy fibre  fibre and  conditioning  antidromic  test  stimpopu-  spikes  61  recordings  of  subthreshold  intracellular  mossy  fibre  and  responses fimbrial  stimuli Figure  Figure  3.4  3.5  The  63  actions  of  bicuculline  inhibitions  i n the  Changes  the  in  bicuculline  on  CA3  region  profile  of  treatment  early  and  late 65  inhibition  after 66  X  Figure  4.1  Ratemeter CAl  records  excitations  of  the  induced  effects  by  NMA,  of  PDA  kainate  on and  quisqualate Figure  4.2  Blockade APV  Figure  4.3  4.4  4.5  4.6  95  of  DL-APV  Depression  Figure  4.8  Blockade  actions  Effects  Figure  5.1  long  of  and  synaptic  Enhancement  NMA,  by  of  cell  firing  L-aspartate  DGG  99  collateral-commissural  a n d APV  antagonists  on of  acids  104  EPSPs  108  excitant-induced  o f EPSPs antagonists  term  actions  in  responses  98  e x c i t a t o r y amino  o f NMA  by  The  kainate  by  of  responses  of Schaffer  depressions  4.10  on  o f APB  antagonism  EPSPs  The  and  L-APV  acid-induced  isomers  L-glutamate  4.7  Figure  and  96  the o p t i c a l  Figure  4.9  by  neurons  evoked  Figure  responses  DGG  Differential and  Figure  amino-acid-induced  A l t e r a t i o n s o f amino by  Figure  of  Effects CAl  Figure  and  93  110 on  population  spikes  potentiation of  bath-applied  112 D-APV  on  EPSPs  plasticity  of  applied  the perfusate  synaptic either  113  excitation i n CAl  by  iontophoretically or 135  xi  Figure  Figure  5.2  5.3  Kainate-induced  changes  antidromic  potentials  Changes fusion  Figure  5.4  field  of  of kainate  Effects  of  5.5  Figure  5.6  I n CA3 and  Figure  5.7  treated Figure  Figure  5.8  5.9  5.10  Figure  5.11  after  per139  and  orthodromic  field  dentate  o f mossy  fibre-evoked  region  143  of kainate  on  spontaneous  potentials  of input/output  145  curves  i n the kainate-  gyrus  148  of EPSPs and enhancement o f  dromic  excitation  fusion  of kainate  effects  of  i n the  dentate  after  orthoper150  other  spikes  Kainate-induced inhibition  138  relations  EPSPs  by k a i n a t e  on p o p u l a t i o n Figure  i n CAl  140  the actions  Depression  The  on  i n t h e CA3  evoked  Shifts  and  spikes  Potentiation activity  orthodromic  i n the CAl region  kainate  population Figure  input/output  of  excitatory  amino  acids  i n CAl  reduction  152 of  paired  pulse  i n CAl  154  Simultaneous  intracellular  analyses  kainate's  of  and  actions  extracellular on  synaptic  processes Figure  5.12  Abolition  158 of  ically-evoked in  CAl  orthodromically IPSPs  during  and  antidrom-  kainate  treatment 159  xi i  Figure  5.13  Changes and  late  kainate Figure  Figure  Figure  5.14  5.15  5.16  of  heterosynaptic inhibitions  in stratum  blockade  inhibition  i n CA3 of  6.1  6.2  6.3  of  kainate  the Figure  Figure  6.5  6.6  gyrus  Mossy  of  curves  field  Correlation the  and  folate  and  late  and  excitation  and  gyrus  169  spikes  in  CAl  folate of  184 synaptic  activ185  EPSPs  in  CAl  and  treatment with  folate  CA3 on  between  the  den186  discharges  in  region  190  orthodromic  and  anti-  i n CA3  191  spontaneous  onset  popu-  folate  epileptiform  potentials  delayed  the  r e l a t i n g changes of  folate-treated  dromic  of  163  gyrus  fibre-evoked  of  synaptic  alterations  after  i n CA3  bicuculline  population  dentate  Effects  and  on  perfusion  spikes  of  i n h i b i t i o n s i n CA3....166  dentate  of  Input/output  tate 6.4  i n the  i n the  lation  Figure  of  Actions  ity Figure  effects  heterosynaptic  Folate-induced  early  165  the  Potentiation  pulse  iontophoresis  kainate  on  following Figure  by  kainate  inhibition  Figure  after  pyramidale  Selective  Summary  paired  of  evoked  bicuculline-treated  burst  firing  activity  slices  in 192  xi ii  Figure  6.7  The  effects  fimbrial Figure  Figure  6.8  6.9  of  stimuli  Extracellular of  tiform  activity  timing delayed  of  and  strength  burst  firing  intracellular  folate-induced  actions  recurrent  on  and  istics  The  the  of 194  character-  spontaneous  epilep-  i n CA3 folate  inhibition  196 and  kainate  of CAl neurons  on  the 199  xiv  ACKNOWLEDGEMENTS  For this  study  h i sgenerous I am i n d e b t e d  Collingridge,  described  over  t o D r . Hugh M c L e n n a n . efforts  i n Chapters  the course of To D r . Graham  made p o s s i b l e t h e  4 a n d 5, I e x p r e s s  my  thanks. Gifts  gratefully  o f compounds from D r . J . C . W a t k i n s  figures  technical  was p r o v i d e d I  external  would  reviewing  assistance i n the preparation of the  by K u r t  also like  examiner, this  Finally, Studentship  (Bristol) are  acknowledged.  Expert  for  and c o u n s e l  whose c o l l a b o r a t i v e  investigations sincere  support  Henze.  t o thank  D r . P. A n d e r s e n , t h e  a n d t h e members o f t h e a d v i s o r y  committee  thesis.  I wish  t o acknowledge  by t h e M e d i c a l  Research  t h e award o f a  C o u n c i l of Canada.  1  CHAPTER 1 GENERAL  In  the r a t the hippocampal  postero-medial  portion  comprises  than  It  more  consists  dentate Ammonis;  their  occurs of  and  transfer  dentate field  of  gyrus,  CA3  arising  granule  and  finally  3  laminae  the stratum cells;  an  laminar  from fibre  of  granulosum  projection  outer  molecular  each  bodies, are  hippocampus  which  from  fibre  to  system  2.3A).  are: a  t h e somata  layer  to the  the dentate  CAl (Fig.  gyrus  consists  cortex  collateral  containing  cell  the  circuit  the dentate  thesis  fashion.  within  i n field  The  pathways  the entorhinal  the Schaffer  (cornu  proper,  i n which  tri-synaptic  f r o m CA3 a n d t e r m i n a t i n g The  layer,  t h e mossy  i n this  efferent  information  pathway  horn  the  complex.  described  and  area.  cortex,  and t h e hippocampus  discernable  v i a the so-called  the perforant  the subicular  afferent  and  surface  o r Amnion's  of archicortex  in a clearly  The  gyrus  the  hemisphere  the entorhinal  proper  and  occupies  the cortical  investigations  i s composed  dendrites  formation cerebral  divisions:  CA1-CA3)  on t h e d e n t a t e  arranged  of  t h e hippocampus  fields  which  each  one-third  electrophysiological  of  of  of 4 major  gyrus,  focused  INTRODUCTION  central  of'dentate  consisting  of the  2 dendrites  of  ( f i e l d . CA4  these of  distributed afferents  Lorente  along from  terminating the  and  the  i n the  field  CA3  1983).  on  inner  S i m o n s e n and  fibre  from  which  entorhinal passage  moleculare Field  CA3  lateral  in  the  the  cell  cortex to regio apical  (Hjorth-Simonsen also  1961)  Schwerdtfeger neurons  and  terminate  layer  field  i s  also  (Hjorth-  arranged  neurons  aligned  basal dendrites extend  from  stratum body  1981).  as w e l l  as  the  molecular  a  forms  Jeune,  into  lucidum,  cells  narrow  (Blackstad projection  and  zone Kjaer-  from  the  synaptic contacts  d e n d r i t e s i n the and  the  stratum  1972;  afferents i n p u t s from  en  lacunosum-  Stewart, from  3  the  granule  A  inferior  into  in in  'dentate  layer  receives commissural  homotopic  from  Kjaerheim,  molecular  pyramidal  input  Gaarskjaer,  with distal  and  dendrites project  fibre  primarily  1961;  and  synaptic input  these  inferior)  apical  above  of  1977).  of  mossy  areas  outer one-third  1973;  of  dentate  somata  o r i e n s and  heim,  the  the  stratum  immediately  outputs  (regio  pyramidale  with  (Hjorth-Simonsen  receive  Cowan,  CA3  stratum  are  cells  entorhinal  and  (Blackstad  and  of  Laurberg,  with  dentata  fibres  granule  lateral  hilus  pathway  major  third  Field  terminates  dentate  The  fascia  1976).  (Gottleib  Sarvey,  The  of  and  the  Entorhinal  dendrites  medial  Stewart,  mossy  layer.  the  of  1934).  respectively  the  lamellae  No,  hilus  moleculare,  1972;  the  de  the  i n the middle  Cells both  and  approximately  stratum  Jeune,  cells;  1976).  the c o n t r a -  ipsilateral  CA3  3  neurons  (Gottlieb  fibres  of  radiatum  these  major  pyramidal  terminate tively  are  Cowan,  the  originate  mainly  in  formation  major of  thalamus,  accumbens  the  cells  strata  1978;  r a t are  cortical  the  to  strata  CAl  and  oriens  oriens  and  body,  superior)  Schaffer  and  collateral  lateral  association  the  respec-  hippocampal  nuclear  septal  areas  and  1981).  of  anterior  1980)  radiatum  et a l . ,  projections the  (regio  ( S w a n s o n et. a l . ,  Voneida  to  mammillary  and  inputs  commissural  efferent  the  in  The  1973).  i n CA3  (Swanson e t a l . , The  the  and  S w a n s o n et. a _ l . , 1 9 8 1 ) .  terminate  excitatory  cells  which  Cowan, 1973;  pathways  (Gottlieb  The  fibres  and  complex  nuclei,  (Swanson  of  nucleus  and  Cowan,  1977). Despite of  hippocampal  al.  (1982)  'the  function  worth  noting  be  that  "the  a  single  inferred  that  a  precise  definition  Indeed  S w a n s o n et_  elusive.  number  studied  the a f f e r e n t  of  general  approaches  it". input  In of  sensory  the  this  theories  of  number  of  context  the hippocampus  modality  i t s effector  and  output  i t is cannot  i t  might  is related  to  functions.  of  the  current  hippocampus  in cortical  reviews  O'Keefe  of  been  hippocampus  have  i s beyond  complexities  has  the  who  to  many c o r t i c a l It  of  1  that  confined  reasonably  function  commented  investigators  be  i t s anatomical prominence  and  scope  of  hypotheses activity. Nadel  this  thesis  regarding For that  (1979)  and  to  consider  the  role  purpose Olton  of  the  and  the the  recent  his  co-  4  workers  (1980)  general role  concensus  in  the  brain which and of  recommended. that  subserve  brain  such  as  campus  in  the  regard  to  the  of  physiological campal  pioneered  of by  for  1981). perhaps features are  the  to  alter  the  evidence in,the  interactions within  the  tissue  relatively of  the  most  extensively  ionic  hippo-  Finally,  analysis  a  connection,  the  to  Mcllwain  the  locus  with  that  composition  of  or  the  hippo-  years  the  (1966),  has  provided  simple  yet  powerful  activity (Kerkut  using and  slice  of  slices barrier  add  of and  drugs  to  both Wheal,  has  Particularly  brain  of  has  technique  hippocampal  ijn v i t r o  slow  a  recording  studied.  blood  10  ^r. v i t r o ,  neuronal  the  the  various  last  of  1979).  i_n v i v o  of  the  etiology  function  intracellular  e l i m i n a t i o n of  and the  (Delgado-Escueta,  and  incubation  of  regions  hippocampal  this  the  nuclei  state. is  learning  regarding  In  of  mammalian  diverse  involvement  a  vital  to  evaluation  and  the  a  contributed  brain  with  plays  is  doubt  and  of  Yamamoto  the  extracellular  no  However,  slices  raphe  prominence  inherent  properties  neurophysiologists method  has  information  afferents.  maintenance  some  from  there  i n humans  which  difficulty  input  behavioral  dysfunction,  epilepsy  of  there  function, particularly  additional  assume  factor  development been  may  say  formation  hypothalamus,  of  i t to  activities  receives  modulation  lobe  A  the the  cortical  hippocampus  hippocampal  cognitive  i t also  suggests  Suffice  cortical  That  coeruleus  temporal  the  abstruse  memory. the  are  been  important hippocampus the the  ability medium  5  bathing cell et  the  tissue.  bodies,  their  to  In  be  the of  determine  the  processes studies  and  are  the  each  of  the  and  major  especially  been  assessing a  than  those  its  are  pathways  amino  is in  of acids  the  to  central  be  well  due to  significance and  separately  neurochemical L-glutamate  to  and  the  inhibitory  in  the  system. of  acid  have  higher  regions of  the novel  character-  3. major  intrinsic  excitatory  implicated  L-aspartate  (GABA)  apparently  the are  also  processes  pharmacological  that  and  somewhat  Because  this  in  author's  inferior  ^-aminobutyric  circuit  studies  gyrus  the  Experiments  regio  in Chapter  established  2 of  i n h i b i t i o n which,  nervous  trisynaptic  to  to  These  potentials  dentate  not  in  transmitter  physiological  field  the  synaptic  i n Chapter  previously.  long-lasting  inhibitory  discussed  It  have  in  excitability.  CAl,  interactions  attributable  functional  inhibition  which  shown  mammalian  possible  results  and  was  "classical"  istics  CA3  well-described  potent  unexpectedly,  the  in  regions:  thesis  exploited  acids  characterization  hippocampal  heterosynaptic  revealed  other  a  neuronal  (Andersen  control.  this  been  amino  orthodromically-evoked  those  knowledge  by  of  in  of  iontophoretic  visual  have  excitatory  modulation  prefaced  antidromically  preparation  of  and  under  described  organization  pathways  recording  positioned  slice  role  lamellar  afferent  stimulating,  precisely  the  the  and  investigations  advantages  of  dendrites  a l . , 1971a) a l l o w s  electrodes  the  Furthermore,  as  the  their  and  the  acidic putative  6  transmitters. developed of  compounds  amino  on  the  stimulation results  of  of  synaptic  a  of  the  system,  in  the  of  as  are  discussed acid  recently  antagonists  spinal  neurons  stood.  With  properties by  hippocampal  the  cord  mediated  with  were  by  the  input. regard  receptor(s)  The  to  the  mediating  into  aim  the  of  amino  of  acid  excitatory reacts.  naturally  epileptogenic hippocampus  these  actions  providing the  are  an  in  the  and rn  central  neurotoxic  vivo,  not  were  however  well  insight  examined  amino Kainic  under-  into  electrophysiological  acid  intracerebral behavioral  temporal  lobe  interest  in  acid  occur  profound  electrophysiological  class  kainic  does not  kainate,  neurodegenerative  recently  one  the  in  these changes  each  major  region.  the  for  of  which  underlying  the  Since of  has  of  produced  addresses  with  injected  mechanisms  folic  number  described  CAl  amino  activation that  when  ligand  of  a  collateral-commissural  chapter  compound w h i c h  effects  many  Schaffer  excitatory  receptor,  nervous  the  been  excitations  experiments  fifth  sequellae  acid,  have  of  excitation.  The  acid  the  the  effects  excitation  these  of  the  which  acid-induced  examined  nature  Therefore  the  there  kainic  acid  attention has  been  of  kainate  can  neuropathological such is  as  Huntington's  both  scientific  identification  received which  and  diseases  epilepsy the  injections  of  receptor. in  this  shown t o  the  replicate  symptoms  of  chorea  and  and  clinical  naturally-occurring A  compound  capacity  is  which  the  cause k a i n a t e - l i k e  has  vitamin seizures  7  and a  toxicity.  Therefore  physiological  folate  is  i n the  "identity  addressed.  of  final  chapter  action"  the  between  question kainate  of and  8  CHAPTER P R E P A R A T I O N AND M A I N T E N A N C E IN  V I T R O AND  2  OF THE  HIPPOCAMPAL  C H A R A C T E R I Z A T I O N OF  EVOKED  SLICE  RESPONSES  Introduction Since ( 1966 ) vitro  i t was  the recording  has been  various  first  of  t h e CNS.  substantia  nigra,  cerebellum  and o p t i c  Perhaps  hippocampal described  in this  In  Wheal, 1980)  are  tectum  has  part  the spinal  neocortex,  studied of  from cord,  thalamus, and Wheal,  of  these,  the  the i n v e s t i g a t i o n s  the methods  of the r a t hippocampal excellent  reviews  have been p u b l i s h e d  differences  between  the  chapter  e_t a l . , 1 9 8 0 ; L y n c h ,  t o the reader.  evoked  obtained  see Kerkut  the focus  Several  procedures  major  include  ( f o r review  of this  recommended  The  Mcllwain  thesis.  described.  by o t h e r  These  been  1981; Dingledine  described  and  to tissues  extensively  and maintenance  of these  Yamamoto  olfactory-and  t h e most  the f i r s t  preparation vitro  septum,  slice,  by  of e l e c t r o p h y s i o l o g i c a l responses i n  successfully applied  regions  1981).  reported  In the main  techniques  there  reported  f o rthe  slice  i n  (Kerkut  and  1980;  Teyler,  r e c e n t l y and a r e are only here  minor  and  those  authors. aim of  this  potentials occurring  chapter  i s to characterize the  i n C A l , dentate  gyrus  and  CA3  9  preparatory  to  elicited  by  Because  they  activity  excitatory have  both  briefly.  i n v e s t i g a t i o n s of  regarding  been  i n CAl  Since the  amino well  and  acid  more  detailed  been  undertaken.  of  the  dentate  and  consideration  c u t t i n g and  Unanaesthetized were  decapitated  midline bones dura ly, ly  incision.  were  then  covering at  at  After  the  basis  of  the  the  the  removal  from  cold  pooled  facilitated the  moistened Midbrain the  right  of  the  (8-12  fluid blood  and,  was  filter  diencephalic  hemisphere  the  a has  2  latter  of  from  and  cuts  bisected  the  :  were  the  brain.  in a  beaker  artificial washed  become  tissue.  sagitally  using  ventro-medial  away  firmer,  After  placed  then  the  anterior-  95/5%)  to  and  posterior-  free  treatment  beaker,  structures  and  placed  a  occipital  Coronal  2  gm)  following  forceps  i t the  the  150-250  heavy  (0 /C0  causing  to expose  available  tissue  reflected  b r a i n was  This  handling  and  only  generation,  (Wistar,  w e r e made t o  gassed  removed  paper  rats  using  angle  by  is  hippocampal  f r o n t o - p a r i e t a l suture,  (ACSF).  evoked  electrophysiology  resected.  skull,  the  i s considered  interparietal  removed  °C)  subsequent  brain  and  the  of  was  parietal,  b r a i n was  others,  Methods  scalp  cerebello-pontine  cerebrospinal  sec  The  antagonists.  for their  CA3  albino  the  carefully  level  containing  any  and  and  information  the  maintenance male  by  gyrus  less  General Isolation,  agonists  described  substantially  nature  e l e c t r o p h y s i o l o g i c a l changes  30-60  on a  ACSF-  scalpel.  dissected aspect  of  from the  10  hippocampus  and  connections  were  beneath  the  ventricle  its  septal-anterior  cut.  was  used  to  alvear  was  surface  and  the  placed Mcllwain  its  within at  an  angle  1980).  the  cutting  using  a  the the  of  to  third  70° of  a  of  dropper,  to  adjusted  axis  f i l t e r  lifted on  of  so  the  that  the  and  the  paper  connections  with  the  of  of  cutting  a  were  spatula  cutting  stage  and  of  was  blade  (cf. Fig. 8  of  Super  mesh  a  were to  brush.  transferred, support  The  maintain  (nominal)  Blue)  soft-bristled  nylon  level  thin  tri-synaptic  hippocampus  hippocampus  (Gillette  and  the  the  thickness  right  chamber.  to  was  inserted  cavity  remaining  side  long  obtained the  incubation  Any  the  hippocampus  the  integrity  the  the  blade  were  faced  400^A,m  fine-tipped  slices  the  hippocampus  slice,  Slices  middle  into  spatula  a  chopper.  preserve  each  now  ventro-medial  tissue  To  over  uppermost.  isolated  on  margin  roll  surface  temporal-posterior  teflon-coated  ventro-medial  ventro-medial  cut  A  and  of  layer  of  cold,  using the  over  placed Teyler,  from  transferred  In  the  taken  loop  gassed  this a  manner  from ACSF 6-8  wide-mouthed  slice  bath  perfusing the  the  in  ACSF  surface  the was  of  the  minutes.  To  slices. The  entire  procedure  obtain  healthy  slices  trauma  to  hippocampus  slicing. before  the A  period  experiments A  of  normally  i t was  important  during  1-1.5  r e q u i r e d 7-9  h  was  to  minimize  mechanical  from  brain  removal allowed  the  for  and  equilibration  began.  diagrammatic  representation  of  the  perfusion  11  apparatus  and  Distilled  at  coil  bubbling water  incubation  water  maintained nichrome  the  in  34  the  °C  via a  gas  c h a m b e r was  outer  by  DC  two  (0 /C0 2  warmed  at  adjusting was  a  and  matched  the  slices  the  water 34  a  and  and  where  bath  at  volume  produce  a  submerged  in this  By  heated  atmosphere  of  the  slice  changeover  the  environs  into the  a  as  from  and  reservoir.  By  this  the  the  other  to  the  reservoir  the side  entered  warmed ACSF of  that  a  slice.  lag  time  again  coursed the  of  Furthermore,  approximately  replacement  of  bath  needle.  caused  the  was  for perfusion  ca.  to drug-containing perfusate of  bath  condition  t u b i n g , was Here  in  pump  outflow  existed  bath.  to  degassing  peristaltic  long  head  a suction  a  complete  was  circuit.  (to prevent a  polyethylene  design  required  as  outflow  slices  by  °C  reservoir  pressure  the  removed  2.1.  jacket a  heating  the  ml/min  and  the  system's  between  arrival  the  The  entered  i t was  1.0  inflow  ACSF.  around  The min  of  constant  with  t o 34  through  ca.  j a c k e t through  °C  below  of  the  to  Fig.  humidified.  drawn  height to  maintained,  to  rate  the  applied  forming  in  water  a i r s t o n e s immersed  : 95/5%)  2  f e e d l i n e s ) was  dripped  given  plexiglass  current  G a s s e d ACSF p r e w a r m e d the  is  thermistor-controlled  i t through  the  chamber  of  10  ml  medium  be  3.5  and  the  its slice  perfused  surrounding  to the  slice. The NaH P0 2  4  ionic  composition  (1.25), NaHC0  D-glucose  (10).  The  3  pH  of  ( 2 6 ) , KC1 was  the  A C S F was  (5), CaCl  maintained  at  2  (mM):  NaCl  ( 2 ) , MgS0 7.3-7.4 by  4  (124); (2)  and  constant  12  Fig. constant in  flow  vitro  planes  2.1  with  a r t i f i c i a l  I n the experiments,  microscope the  cell  more  stimulating afferent refer  field  outlet  support  9. p o l y e t h y l e n e 11.  port,  ring,  and/or  7. s l i c e  bath,  12.  water 20.  (ca.  ACSF  16. n y l o n 34°C),  jacket, 8. A C S F  providing  of  the  fashion,  the appropriate i n the figure coil, 6. n y l o n  port, 0^/Q.O^'.§b/b %,  f o r Q^/dO^'.^b/b % , 1 3 .  bath  optics  layer  electrode,  inlet  i n  a s a band o f  3. h e a t i n g  reference  (ca.  n e t , 17. s u c t i o n  19. f i b r e  reservoir.  The numbers  inlet  pre-warmed ACSF, 14. Haake w a t e r microscope,  positioned  In a similar  to activate  10. a i r s t o n e s  pump,  were  i nthedendritic  2. w a t e r  support net  t h e a i d of t h e overhead  was d i s t i n g u i s h a b l e  5. A g / A g C l  tubing,  peristaltic  on t h e nylon  electrodes  pathways.  t o : 1. t h e r m i s t o r ,  4. A C S F  t o scale  were p l a c e d  and e f f e r e n t  The  i s drawn a p p r o x i m a t e l y  t o be e x a m i n e d .  electrodes  (ACSF).  i n B and  recording  tissue,  slices  bath  and with  which  fluid  o f t h es l i c e  slices  below  layer,  translucent  hippocampal  net  from  (1.6X-4X) body  views  f o r the  o f hippocampal  cerebrospinal  i n A; t h e b a t h  illuminated  o f t h eapparatus  perfusion  o f t h ec r o s s - s e c t i o n a l  A.  were  outline  ( c a . 1 ml/min)  C areindicated in  A schematic  34°C),  needle,  15. overhead 18. d i s t i l l e d  f o rillumination of slices,  13  14  bubbling  with  changes  of  examined,  0 /C0 2  : 95/5%. . I n s t u d i e s  2  ionic  composition  osmolarity  on  where  the e f f e c t of  neuronal  was m a i n t a i n e d  activity  by a d j u s t i n g  was  t h e concen-  2+ tration mM) in  of NaCl  except:  the concentration CI  replacement  a) i n l o w Ca  o f MgSO^ w a s i n c r e a s e d experiments  substituted  f o r NaCl.  Preparation  of iontophoretic  Multibarreled on  a vertical under  visual  Barrels  were  then  distilled of  water,  the tips  drug  control  was removed  barrel  tips.  When  with  to cool  back  a Leitz by  was  18 h w a s a l l o w e d  i t was n e e d e d was f i l l e d  i n  boiling  to within  filled  with  1 cm  the test  f o rd i f f u s i o n  f o rrecording, with  drawn  micro-manipulator.  submersion  Once  were  t o a diameter of  and t h e water  by s u c t i o n .  of the electrode  and b)  propionate  f o r iontophoresis  water-filled  allowed  t o 4.7 mM  sodium  and t h e t i p broken  solutions at least  barrels'  where  medium ( 0 . 2  electrodes  electrodes  puller  5-10^m  -containing  t othe  the central  1 o r 4 M NaCl  (0.5-2.0  megohms).  Stimulation  and r e c o r d i n g  Monophasic 80  V,  were  <100 y * A ) delivered  electrode commonly stainless  (1 M  responses  s q u a r e wave p u l s e s from  a  i n some  wires  ( 0 . 1 msec d u r a t i o n ,  constant-voltage experiments  NaCl-filled;  v i aa bipolar steel  of f i e l d  through  t i pdiameter  electrode twisted  isolated  comprised  together  up t o  stimulator  a glass  micro-  1-2_/*m) b u t m o r e o f 62y*m  diameter  and i n s u l a t e d  to their  15  tips. The  Stimulus positions  Fig.  frequency of  stimulating  for  2.2  recording  output  illustrates  extra-  devices To  are  barreled  noted.  illustrated  in  tubing  resistances  input  were  silver/silver  chloride wire  figure.  used.  from  1  field  or  4  1.5 M  Singlemm  NaCl wire  microand had  connected  barrel to the preamplifier or non-inverting  a m p l i f i e r on a T e k t r o n i x The s i g n a l  was  A chlorided silver  fed to the inverting  oscilloscope.  5111 d u a l  was r e c o r d e d located  relative  input  beam to a  i n the bottom  of a  storage grounded  of the s l i c e  (Fig. 2.2). For  advanced  the  experiments  a recording  appropriate  region  evoked  the  pathway.  test  established, further often  the  elecrode of  potential  until  with  s o l u t i o n of the electrode  which  to that  and  or the central barrel of a  pulled  filled  input  or dendritic  electrode  electrodes  o f 1-5 m e g o h m s .  differential  bath  somatic  electrode  of the apparatus  responses;  i n the legend  iontophoretic  recording  capillary  intracellular  extracellular  a single glass  multi-barreled  NaCl  and  the arrangement  are identified  record  responses,  most  electrodes  where  2.3-2.5. Fig.  the  w a s 0 . 0 5 - 0 . 1 Hz e x c e p t  by  a  under v i s u a l  the s l i c e  a  Narishige  Once  increases  t h e maximum  (<0.2 Hz)  the optimum  stimulating  c o n t r o l through the  to obtain  low frequency  micromanipulator  electrode  slowly  was  advanced  obtainable;  a p o s i t i o n of the s t i m u l a t i n g electrode  tip within  surface  was  used.  were  depth  of  not  10-20^am o f t h e s l i c e  of the response  activation  recording was  field  16 A  Current Pump Main Oscilloscope Stimulus Timer Micromanipulators Ag/AgCI Reference Electrode  —<<* Computer  B  Chart Recorder  R t . meter Record a  e  Oscilloscope  Audio Monitor  Discriminator /Ratemeter Balance • Probe Output to main oscilloscope  Piezoelectric Microdrive  D C Amplifier  Amplifier Heodstage \ - A g / A g C l half-cell  Probe Driver (to stim. timer) Cycle Timer Microdrive Control  Fig. the  2.2  recording  responses.  (a)  barreled-  (left  of  to  hippocampal recording  and  RAD-II-A  bipolar  upper  dual  isolated  program PZ-500.  6.  In  addition  incorporated  devices  responses beam  Gould  delay  2200 M-701  control,  intracellular  (lower  DS 2,  (b)  to  single in  the  intra-  piezoelectric  figure)  (numbered)  3.  to  used  microprobe  DC  Burleigh  2.  Winston 4.  Angus  recorder,  10.  used  to. t h e s e , a  for  impale for  the  were:  Esterline brush  devices  electrodes  storage oscilloscope,  stimulator  5.  (B)  stimulating  electrode  The  and  discriminator/ratemeter,  Instruments and  an  output  iontophoretic-,  metal  also  analyzing 5111  and  m i c r o m a n i p u l a t o r s were  figure).  neurons.  oscilloscope,  W-P  operated  advance  window  recorder, 8.  and  Tektronix  beam  (c)  input  extracellular  experiments  microdrive  D100  (A)  of  muItibarreled  and  cellular  1.  of  Manually  position  slice  Arrangement  Electronics  Tektronix 601R  7.  Digitimer  Apple  502  dual  rectilinear II  amplifier, Inchworm  computer, 9.  Ortec  controller  17  Amplitudes from  computer.  baseline  EPSPs  t o t h e peak  PSs were  positivity  on  were  using  records  conventional  O.D.)  PE-2 v e r t i c a l  acetate  half  of the electrometer  cell  from  an  pre-stimulus  noted;  amplitudes  or the preceding  peak  Signals  silver/silver  neurons  Microelectrodes  were  pulled  puller,  and connected  solution.  C A l a n d CA3  techniques.  D o t ; 1.5 mm  a Narishige  grounded  where  the  with  recording  Haer-Omega  NaCl  directly  o r on l i n e  from  except  measured  t o t h e peak n e g a t i v i t y .  potassium  M  records  measured  amplitude  Intracellular made  filmed  were  c a l c u l a t e d from t h e b a s e l i n e  Intracellular  ick  responses  the o s c i l l o s c o p e , from  Apple  of  of evoked  just  (Freder-  before  use  with  1 M  back-filled  t o the s i l v e r / s i l v e r  (WP I n s t r u m e n t s ,  were  chloride  recorded wire  with  were  chloride  M-701) v i a a 1 respect  i n the bath  t o the  (cf.  F i g .  2.2) . The  microelectrode,  •one a r m o f a W h e a t s t o n e extracellular  fluid,  electrode  and  adjusting  the bridge  drive, cells  aided  adjustment. steady currents  state  bridge.  With  current  was  i t sresistance  the electrode was  t i s s u e and r e c o r d i n g  balance. was  slowly  Using  the electrode injected megohms) a  voltage t o -1  d e f l e c t i o n caused nA  t i p i nthe  through was  and  the  nulled  by  micro-  impalement  the negative  r e s i s t a n c e of neurons  formed  piezoelectric  advanced  by o v e r c o m p e n s a t i n g  Input  o f up  (40-60  chamber  of  capacity  was e s t i m a t e d  by t h e  by h y p e r p o l a r i z i n g  a n d c a . 200 msec  i n duration.  The  18  amplified plotted  output  on a G o u l d The  were  signal  membrane  of  either  records  or  displayed  chart  amplitudes  measured  filmed  DC  was  o s c i l l o s c o p e and  p o t e n t i a l s , EPSPs  directly  p o t e n t i a l was  an  recorder.  action  from  on  the  from  IPSPs  the o s c i l l o s c o p e ,  chart  determined  and  records.  after  The  withdrawal  from  resting from  the  cell.  Results The in  vitro  value  activities  have  function  a  1975,  normal  persistence  considerable  1977).  activity  of normal  in  criteria  were  integrity  consistency  experiments.  were vitro more either spike  t o be  clearly  unacceptable different  o r ir\ v i v o . T h e r e f o r e , of  the following  CAl or the dentate  of  i f their  from  in  1976;  of  this  correlated  features  of  neuronal  and T e y l e r ,  i s also  with  ( F r o t s c h e r e_t  established the  Generally, responses reported  slices  were  reasons.  to verify  slices  those  gyrus  the results  1981).  electrophysiological between  slice  electrophys-  preservation  histological  The  and  (Alger  vitro  and C o t t r e l l ,  of  parallelism  The  a l . , 1981; Cobbett  following  the hippocampal  i_n s i t u ,  between t h e two p r e p a r a t i o n s  apparently  judged  from  the preservation  recorded  indicated  Schwartzkroin,  the  obtained  i s d e p e n d e n t on  iological others  of data  First,  ensure  slices  i n normal as  occurring  discarded  were ACSF in  f o r one o r  the appearance i n  of a secondary  (PS) i n the o r t h o d r o m i c a l l y - e v o k e d  and  the  (2°)  response.  population This  was  19  interpreted the  ratio  as  a  failure  of the f i b r e  dendritic  with  stimulation  to produce  dentate  elicited CA3  correlated  with those  stage  their  poor  slices  ication ACSF  or  level  (baseline  activity  of  incubation  a  granular, usually  adjustment  i f i n CAl or  which  of  This  be  negativity); i n i n CA3  of  of P S s , an  was  the  often  tissue.  edematous  was  mottled  could  inhibition.  indistinct  cell also  the gas  a t any often  the case to  t o poor flow  For  lines  appearance  attributable of  of  of b u r s t s  grossly  activity.  condition  improper  o r had  intensities  activity  appearance were  was  appearance  comprised  which  this  Third,  t o peak  the  to a loss  atypical  slices  had  a  EPSP.  a n t i d r o m i c PS  Fourth,  electrical  which  surfaces  mV.  an  Second, i f  Most o f t e n  high  electrophysiological  example,  showed  15 mV  attributed  Aberrant  for  a dendritic  or evoked  w h i c h was  of  requirement  than  10  inhibition.  t h a n c a . 1:2.  t h e maximum  less  than  spontaneous effect  gyrus  was  less  a  synaptic  volley:EPSP amplitudes recorded i n the  l a y e r was g r e a t e r  correlated  the  of  in  their  humidif-  rate  or the  i n the bath.  Characterization  of f i e l d  responses  CAl The electrodes activation the  stratum  elicited  a  positions are of  of  stimulating  illustrated  Schaffer  radiatum dendritic  in  (S) and  Fig.  2.3A.  collateral-commissural  (S ) w i t h response  low  intensity  ( R ' 100-200j^m 2  recording  (R)  Orthodromic (SC)  fibres  ( F i g . 2.3C: from  the  in 5V)  soma)  20  C.  Orthodromic  Antidromic  EPSP  Population Spike  SC(S )  SC(S )  A  Fig. and  2.3  recording  stimulating the  A.  extracellular  with  fibres  with  S^)  and  Simplified cell and  (  S A  )  of  in  responses in  the  the  alveus  Dendritic  electrodes the  SC  feedback  in  stratum  schematic  receives  of  inhibition  somatic  the  stratum  pyramidal  volley field  Typical  indicated potentials  intensities explanation. stimulus  is  CAl  of  by  CAl  field  filled  (R,)  evoked  this  indicated  and by  an  a  by (S.)  CAl  (R^-'^n  The  apical  were beam"  (R ? 2  records  pyramidal dendrites  axon  of  presynaptic  (S_J See the  a  (iv).  through  records)  stimuli.  B.  activation  course  of  pyramids  potentials  collateral  antidromic  subsequent arrow.  the  below  activated  respectively.  the  cells  A  stimulation  circuitry.  EPSPs  square  orthodromic In  (R^),  from  (i)  region.  radiatum  results  Efferents  CAl  radiatum  field  ( i i ) to  via  stimulating  input;  input  ( i i i )  for  excited  and  neuronal  excitatory  of  (SC) D  interneuron  C.  stratum  pyramidale  CAl  i n "the  (S )  GABAergic  alveus.  electrodes  collateral-commissural  antidromically. recorded  Positions  electrode  Sehaffer  efferent  A  the fibre  and  somatic  and  various text  time  of  for the  21  consisting these  o f two p r e d o m i n a n t l y  represented  terminals The  potential, arising  and EPSP;  the  stratum  pyramidale  EPSP).  As  7V)  negative-going synchronous  EPSP.  Because  the  wave  efferent pyramidale  latency  PS  the only  dendrites  cells  of  (reversed  As  was  somatic  action on t h e  sometimes  the  reversed source a  apparent  i n  i n the dendrites.  the alveus  the AD-evoked  response  consisted  i n this  and a  pyramidale  through  be  2.3C:  poten-  a c t as a  coursing  would  (reversed  representing  axons  a  i n the  (Fig.  of CAl c e l l s  by  current  reached  i n the stratum  PS)  a  wave  was  the dendrites  occurring  produced  recorded  increased  (PS)  discharge  as  flow  small  amplitude  expected,  situation  was p r o d u c e d (S ) . of a  by In  short  p o s i t i v e wave ( F i g .  the p o t e n t i a l recorded  was c o m p r i s e d  of a reversed  in PS  ( n o t shown). Alvear  of  i n some  postsynaptic  (AD) a c t i v a t i o n  followed  S_).  was  spike  p o t e n t i a l recorded  stratum  2.3C,  was  positive-going  situation  sink  Antidromic exciting  for firing  i n this  current  field  a  and  e t a_l., 1978).  behaves  this  of  fibres  change  et. a _ l . , 1 9 6 6 ) w a s s u p e r i m p o s e d  positive-going the  body  first  the current  conductance  intensity  monosynaptic  (Andersen  The  excitatory  reflected  EPSP,  population  tials  for  (R-^) a s  the stimulus  the threshold  1960)  the c e l l  dendritic  Andersen  (field  dendritic Since  f o r the  volley;  wave  Andersen,  postsynaptically. source  fibre  larger  from  waves.  the d e p o l a r i z a t i o n of presynaptic  (presynaptic  second  negative  s t i m u l a t i o n , i n addition to causing  CAl pyramidal  cells,  produced  a  monosynaptic  AD  invasion  activation,  22  via  a  recurrent  interneurons ment  of  collateral  releasing  this  manner.  alternate  occasions The  difference the  An  u  c  L J  assessment  orthodromic  inhibition  conditioned  (PS -PS /PS  the  was  (C)  and  acid  msec)  then  same  inhibition  in  be  (3-6  Measure-  made  in  a PS  the  was  on  conditioning as  the  AD  percent  determinations) (U)  method  the  by  calculated  unconditioned  inhibitory  (GABA).  could  amplitude  The  of  stimulus evoking  (15-30  mean  xlOO% ).  of  inhibition  preceded  between  ( F i g . 2.3B),  ^-amino-butyric  recurrent  following  activation.  pathway  orthodromic  was  dentate  of  applied  PS  in  the  the  CA3  stimulating  and  gyrus  and  region.  Dentate  gyrus Fig.  recording medial LPP,  2.4A  indicates  electrodes.  (S )  or  c  the  respectively)  layers 2.4C, when  V).  5  recorded  As in  the  orthodromic  to  i n the  reversed An  AD  mossy  PS  response fibres  the  in  be the  perforant middle fibre  and  stratum  by  either  outer  a  and  molecular  and  EPSP  the  (MPP  EPSP ( F i g .  was  reversed  ( F i g . 2.4C,  caused  : 5 V) .  negative-going  this  recordings  elicited  and  the  of  pathway  volley  granulosum  potential  of  stimulation  region,  stimulation  dendritic  could  (MF)  CAl  stratum  somatic  i n the  i n the  presynaptic  in  Threshold appear  (Sp)  produced a  positions  intensity  lateral  respectively R :5  Low  the  was  evident  PS  as  a  ( F i g . 2.4C,  Rj.:6.2 V ) .  stimulus  backfiring  a  lucidum  of  CA3  (S  ) or  the  23 ! i  C.  Orthodromic EPSP  Antidromic  Population  MPP(S )  MPP(S )  C  Fig. gyrus. outer  2.4  stimulation pathways in  A.  medial  the  and  The  granulosum synaptic  stratum  MPP)  somatic  mossy  CA3  response  LPP  ( i ) and  a  GABAergic  volley  indicated  dendritic by  a  response  (R3)  evoked  efferent  (Sg)  pathways.  subthreshold similar  (5V) to  in  by  and  those  explanation.  filled  CA3  EPSPs  of  suprathreshold occur  in  An  the  and  of  hilus  antidromic stratum  excitatory  a  axon  recurrent  fibre  input  ; presynaptic  fibre  the  somatic  afferent  field (6.2V)  i n CAl  the  efferent  mossy  by  electrode  the  receives  (R^,  stimulation  which  an  ( i i i ) by  square)  Changes  or  evoke  and  perforant  Q  (S„)  ( i i ) .  as  (R^) evoked  (S )  recorded  MPP  dentate  middle  stimulating  and  cell  the  were  region  interneuron  terminates  Typical  A  was  v i a the  and  the  in  lateral  fibres  input  C.  for  the  granule  (iv).  text  and  The  collateral  are  c  B.  activates  in  moleculare  (S )  of  excite  (R^).  electrodes  respectively.  lucidum to  of  potentials  of  used  response.  field of  stratum  was  Positions  layers  (LPP  the  (Sg)  C  Dendritic (R,-)  Spike  (S^,  responses MPP  field  S ) D  and  between  stimulation  ( c f . F i g . 2.3C).  See  24  hilus  (S )  ( F i g . 2.4C).  E  activate axon  GABAergic  (Fig.  As  i n CAl  interneurons  this  via  AD  a  stimulation  recurrent  could  collateral  2 .4B).  CA3 The dromic avoid  positions  responses  i n CA3  electrode  CA3a  CA3  are  arising  (Rg)  in  de  the  No,  anti-  illustrated  in  from d i f f e r i n g  for  each  region  1934).  Fig.  in  To  character-  the  experiment  ortho-  2.5A.  response  corresponding  The  and  recording  the  stratum  approximately  dendritic field  to  potential  was  recorded. Though  shock  to  upon  not  the  which  S^-top  a  of  latency  always  fimbria  both of  the  the  Usually  a  shorter  latency  PS  below)  (FIM)  'Raising  msec.  (see  the  case  of  PS  the  pyramids  occasionally with  FIM  evoke  pure  at  Fig.  3.lEiv).  could  be  response  Further,  elicited  preceded  by  an  EPSP  a  which AD  PS  low  intensity and  but  PS  (up  was  never  shock  also  (Fig.  2.5C,  low  at  high  to  and  the  ( F i g . 5.14A).  15  mV)  less  than  2  recruited  a  AD  invasion trace).  possible  intensity  orthodromic  stimulus  2.5C, the  S^-lower  stimulus an  (Fig.  EPSP  increased  s t i m u l a t i o n i t was a  intensity  reversed  superimposed  i n some s l i c e s even  a  small  (0.2-0.4 msec) r e p r e s e n t i n g  CA3  AD  was  intensity  However, a  evoke  decreased  higher  below),  stimulus  reversed  PS  (see  could  negative-going  trace).  amplitude  not  evoking  ( M a s u k a w a et. a _ l . , 1 9 8 2 )  positioned  (Lorente  not  electrodes  subregions  was  pyramidale  the  in  complications  istics  of  to  (e.g.  response  intensities  was  25  C  -FIM(S ) G  Fig. showing  2.5  the  recording  in  the  A  (R^,)  the  fimbria  of  in  'were  (FIM), fibres  Field  evo.k.e.d ,  in  by  the  by  for  and  stratum  slice  stimulating  potentials  in  of  efferent  antidromic  the  hippocampal  .stimulation  afferent  and  of  electrodes  CA3.  (S„)  hilus  collateral  schematic  locations  responses  py r a m i d a l e (MF)  A.  radiatum  the  stratum  mossy  fibres  fibres  activation of  and  (S„)  of  in  Schaffer  CA1/CA2  (S  ). Pi  Dendritic  responses  representation CA3  region.  the  basal  shown  and  (v)  p o s t a c t i v a t ion activation axon  the  bifurcates  CAl  al. , in  a  the  afferent  CAl  region  1980). cell and  dendrites  C.  of  via  provide (vi)  the  layer  efferent  which  excitation  an  interneuron  at  (iv).  Schaffer  and  the  contralateral  (Rg)  pathways.  the  text  are  terminate A  the the  mainly  recurrent  either  directly  interneuron; in  The  part CA3  collateral  hippocampus field  in  inputs  commissural  following See  least  diagrammatic  circuitry  excitatory  the  I l l u s t r a t e d are  body  local  A  synaptic  i s produced  GABAergic  cells  and  B.  respectively.  feedback  possibly  to  output  ( i i i ) pathways  inhibition  of  ipsilateral  MF  mediates or  recorded.  excitatory  apical  axon  not  input,  major  ( i i ) and  collateral as  the  Two  commissural on  of  were  input  via  the  efferent input  to  (i)  to  (Swanson  et.  potentials  recorded  the  stimulation  for  explanation.  of  26  A or  CA2  stimulating (Fig.  reflecting AD  PS  the  was  usually  a c t i v a t i o n of  r e s p o n s e was  the  negative-going  MF  e_t  although  events  to  the  hilar  V)  1°  PS  upon w h i c h  a  PS  was  directly  dentate  was  successful,  with  MF  gyrus. the  stimulation  a  This  2°  electrode  through was  PS  was  i t  was  a  pure  to  as  firing  could  elicit  onset  3.7 + 0.2  was  usually  of  msec  a  large  However, CA3 in  PS  may  in  three  r e s p o n s e was the  the  hilus.  of  a  PS  of  1°  (1°)  PS,  with  up  PS for  by  a  of  i n i t i a l  stimulus  the  a  PS  a  cells  contribute  1°  a  followed  in  This  (n = 10)  elicit  The  CA3  primary  sometimes superimposed to  of  the  although  hilus.  least a portion  the  of  positioned  comprised  below).  mV  to  the  discharge  cell  2-4  difficult  activating  the  to  site  was  The  proved  an  see  referred  was  response.  It  PS  fibres.  Although  stimulation  and  latency  stimulation  CAl  AD  p o s i t i v e wave/PS  maximum a m p l i t u d e  The  of  latency  which  •  by  synchronous  recording  2°  MF  additional The  shown).  )  the  driven  1971b,  (50-70  A  upon  t r i p h a s i c waveform at  subsequently  stimulation  S  only  short  collateral  coursing  by  the  generation.  (not  evoke  evoked  a_l. ,  is  a  radiatum  obtained.  b i - or  response  closer  2.5C,  pathway  represents  (Andersen  its  to  CA3a  p o s i t i v e wave  (Fig.  never  potential  which  a  in  stratum  Schaffer  neurons were a l s o  activate  field  by  possible  i n the  evoked  A  superimposed  CA3 to  S )  followed  sometimes AD  2.5A,  electrode  to  hilar  applied to  10  mV  evoked  by  a  1-2  mV  positive  wave  ( F i g . 2. 5C,  S^,) .  CA3  stratum  response  by  granulosum  of  experiments  s i m i l a r to Increasing  where  that the  this  occurring stimulus  27  intensity and  the  Paired  in  both  A  on  the  (FIM )  potentiation (Fig.  observed.  The  intervals  was  this  analysis  PS  this  of  FIM-evoked  PS  this  MF  T  C-T  i n t e r v a l s up  to  any  MF  T  great  observed  300%)  of  the  1  PS  evoking  a  2-4  the  up  to  in  i s presented i n the  responses  which  2.6D,E; F i g .  60  in  3.2A,B).  PS  2000 (Fig.  exhibited  paired  msec  was  despite  2°  to  or  (Fig. majority neither the FIM  Thus  degrees  activity  T  and  of  (C-T)  in  some  conditioning  next  revealed  a  that at  or  chapter.  behavior  observed  20-60  inhibition  msec  an  in  the  of  the of  The  of  1°  PS  potentiation ca.  20%  of  experiments,  i n h i b i t e d nor  orthodromic  C-T  inhibition  3. 2B) . pulse  of  with  msec  at the the  potentiated  s u b s t a n t i a l changes which T  be  pharmacological  i n the  2.6B,C)  of  could  and  a f t e r the  Thus  ( F i g . 2.6D) 60  effects  later.  responses  resembled  evident  the  responses degree  at  was  to  hilus  activity.  whereas  response  the  sec  electrophysiological inhibition  msec  two  conditioning-test  FIM  2  of  various  FIM^  of  persisted  either  or  inhibition  However, of  the  2.6A).  C  20-60  longer  activity  MF-evoked  PS  of  (FIM )  have  evoked  either potentiation  the  1°  could  at  orthodromic  evoked  stimulus  to  stimulation  occurred,  slices.  amplitude  i t (Fig.  effect  An  Paired  intervals  usual  action  of  (up  3.1A,B)  an  stimulus.  PS  response  T  inhibition  2°  orthodromic  test  2°  followed  fimbrial conditioning  submaximal  either  the  Stimulation  mV  the  increased  p o s i t i v e wave w h i c h  Pulse  cases  cases  activity  were (Fig.  28  v  \ x  , 1 ImV  '  I msec  _J. 5 msec  E. (i)  L  M  V  (ii)  6 -Vi  10  14  5 msec  Fig. recorded pathway  2.6  Some  response i n the  stimulation  (4-16V)  B.  MF  by  30  a  the  which  increased. to  msec  interstimulus  (ii)  i n the  same  was  an  E.  and  traces  response  apparent C-T  Filmed  intervals, slice,  the  of  the  a t t h e 6 and  i t was in  MF  of  stimuli  the  of the  in 2°  another PS  only  ( i ) illustrate  with  1°  PS  paired  orthodromic  10 m s e c  which  conditioning  However,  MF-evoked  whereas test  the  the  onset  the amplitude  i n C.  records  f o r the  MF of  of  wave  to paired  of  interval  inhibition  inhibited  indicate  the  both  positive  latency  Potentiation  of  intensities  increase  the  of  extracellularly  activation  increasing  test  similar  the  With  decrease  i s also  (D)  stantially  there  msec.  slice  refractoriness  A.  the  of  following  response  and  response at  CA3  Overlapped  record)  separated test  1°  i t ; note  activity. (upper  in  hilus.  negative-going followed  characteristics  at  FIM PS  intervals.  was  the 6-14  shocks sub-  29  High  Frequency Fig.  Stimulation  2.7  illustrates  orthodromic  response  stimuli.  6/8  In  interval  stimulation treatment of  the  a  PS  the  63-432%  period  (LTP)  reported  Ljzfaio,  1973)  as  and  4).  In  activity  after  tetanus  potentiation) Although  the  observation the in  vivo  but  following  LTP  the  of  field AD  FIM  Using PS  or  subsequently  (Fig. 1978).  was  by  to  the of  1-2  This of  PS  stimulation  Spencer, and  FIM  by  Dudek,  i n p u t had the  shock  hilar d i d not  in  1979). no  gyrus  i n only  demonstrated  (but  see  1 of LTP  Yamamoto,  of  the  was  1972b;  and  and  see  evoked  (post  tetanic  amplitude.  an o c c a s i o n a l  vitro  a t no  time  situation (Yamamoto,  Stimuli  producing  effect (Fig.  on  the  2.7B).  LTP. of  which FIM  min  the  stimulation  8 slices  15  (Bliss  of  apparent  LTP  increase  1975;  min  exhibit  pre-  potentiation  t h e r e was  1961a) and  a  the  pre-treatment a 2°  Hz  frequency  having  Wester,  only  100  (the s h o r t e s t  term  dentate  same s t i m u l u s p a r a d i g m ,  observed  2.7B)  PS  end  long  of  high  enhancement  return  tetanic  evoked  the  1°  the for  orthodromic  evoked  the  lasted  MacVicar  response PS  the  appearance  a  the  FIM-evoked  a f t e r d i s c h a r g e s i n c o n t r a s t to the  and  see  resembles  a  transient  (Kandel  to  slices  before  g e n e r a t i o n of  1972a;  The  2  the  of  ( S c h w a r t z k r o i n and  Chapter  sec  ( F i g . 2.7A).  persisted  in  the  train  20  t e r m i n a t i o n of  occurring  CAl  in  sec  within  0 . 5 - 1 . 5 mV  and  1  potentiation  of  amplitude  recording  a  t h e r e was  of  amplitude  changes  following  slices  examined)  the  the had  MF-evoked previously  orthodromic  Yamamoto  and  input Chujo,  30  K>  >  E  • FIM  e v o k e d -00  i MF  evoked-1°  PS PS  2-  0  FIM  J  100 Hz. I sec 10  Time (min)  (i)  (Iii) •  t  FIM (134  V)  (13.4  V)  (IIV)  (IIV) ]*2mV 4 msec  (v)  (vi)  (vi)  (vii)  MF (40V)  (40V)  (40  (40  V)  FIM 00Hz. I sec  Fig.  2.7  stimulation.  Changes  A.  t=0  min  the  FIM-evoked  High  produced  unaffected  to  FIM  superimposed frequency  stimulation PS  unaffected  ( v i ) .  control  to  elicit  of  (iv)  MF  15  the min  the  a MF  MF  the  but  (v)  PS  of  was  hilar  an  not MF  of  the MF  show  15  (3  field  amplitude  had  either  amplitude  of  PS  remained  of  control  consecutive  min  stimulus  changed  at  after  potentiation  MF-evoked  high  of  that  response  was  intensity  was  comparable  ( v i i ) . no  tetanic  fimbria  1°  stimuli  pathway  (viii)  the  records  obtained  FIM the  the  Filmed  ( i i i ) the' FIM  response  later.  PS,  Traces of  In  response;  or  and  following  i n c r e a s e of  B.  ( i i ) whereas  tetanization  shown,  (i)  sweeps).  orthodromic  decreased  (OD)  decreased.  activity  stimulation  long-lasting  orthodromic  or  responses  a  MF 100 Hz;I sec  i n evoked  frequency  V)  effect  to  the  Subsequent on  immediately  the or,  FIM as  31  E f f e c t s of e x t r a c e l l u l a r i o n o n e v o k e d a c t i v i t y i n CA3 Low  [Ca  2+  ] ,  increased  q  Perfusion tions  of  Ca  were  modified  various The  ([Ca  2 +  contribution  potent  blockade  anti-  PS  p o s i t i v e wave  and  the  PS  by  a  actions  of  this  PS  in 3 slices measured,  this  modified  the  latter  (Fig. single stimuli  separated  in  in 7  allowed  be  the  assessed. response  demonstrated the  mM) the  to  orthodromic  In  i t was  same  the  slice  the  unaffected  but  substantially  p o s i t i v e wave  the on  hilus the  others  a  prolonged  caused the  was  MF-  20-50  1°  reduced  msec  was  no  the  this  PS the  (40-80  on min)  recruitment  time  an  a  to  of  response  increase  paired  revealed  the  Furthermore  AD-evoked  input  response  effect  persisted.  the  2°  were i n c o n s i s t e n t .  gradual  each  the  slices),  37+12% o f  PS  and at  with  PS  exposure  the  also  and  1° of  and  (10/10  there  MF-evoked  both  by  (4.7  Q  to  s t i m u l a t i o n was  the  ACSF  treatment  activity;  ] )  (Fig. 2.8Aiv-vi).  with  There  2 +  concentra-  transmission  2.8Ai-iii)  a maximum r e d u c t i o n  in  ([Mg  2 +  FIM-evoked  SC  the  responses  activity.  by  MF  ACSF, the  PSs  the  followed  modified  Even  2.8B). unit  to  Mg  potentials  (Fig.  which  whereas  amplitude.  additional  of  synaptic  shock  and  field  i t blocked  evoked  mM)  i n which  orthodromic  CA3  eliminated  Although  was  of  ACSF  synaptic  and  elicited  the  with  block  of  ]  (0.2  Q  conditions  latency  2°  ] )  abolition  these  Thus  2 +  components  under  2+  [Mg  slices  to of  complete  short  of  concentrations  of  homonymous substantial  32  2.8  Fig. and  Schaffer  first  A.  Control  collateral  column.  After  responses  i n CA3  altered  &  ( i v ) s t i m u l a t i o n a r e shown  the administration 2 +  medium  ( R ) t o FIM ( i )  to contain  low  20 ml  o f ca.  ]  ( 0 . 2 mM)  was  abolished  [Ca  i n the  and  of  a  increased  2 +  [Mg  ]  (4.7  same  time  whereas  PS w a s  column) of  responses. typical the  field  2°  PS  stimuli  [Ca  a  wash-out  activity. MF  [Mg  Q  and  a  the  (iii) short  ]  on  AD  2+  ] /high Q  was  1 ° PS w a s PS  i n  medium  substantial reduction  n o r m a l medium  of evoked  MF  shock  2 +  and  FIM a  evoked  20  increase  of  paired  followed Q  The  elicited  activity  ]  B.  by  FIM  ortho-  t h e 1 ° PS o f t h e  decreased  and a d d i t i o n a l PSs  inhibited  ( i i ) .  i n contrast  Concurrently,  unaffected  but  to the s i t u a t i o n  of the test  activity  (not shown).  MF  PS  [Mg  ( i v ) was  PSs o c c u r r e d ;  changes  MF  paired  the  reversed  medium.  f o r ( i ) but with AD  and  e f f e c t s were  normal  test  latency  In low [Ca response  As  multiple These  2 +  At the  unaffected  reduced  on t h e 1 ° and a s m a l l  no e f f e c t  i t ; the test  conditioning a  with  ( i i ) .  PS w a s  i t was  ( v ) . These  ] /high  amplitude.  producing  followed  2+  potential  showed  conditioning the  eliminated  after  low  following  ( i ) I n n o r m a l medium t h e f i r s t  later  dromic  c o l l a t e r a l - e v o k e d AD  t h e p o s i t i v e wave  actions  msec  t h e FIM response  the Schaffer  secondary (third  mM)  reversed  AD  PS w a s  after  again  i n normal evident.  returning  to  33  34  reduction which  of  activity  was  not  2.8Bi,ii)  (see  Low  apparent  a  discharges,  in  test  the  a l s o Yamamoto,  the  study  response  (Fig. 2.8Bii,iv)  pre-treatment  situation  (Fig.  1972b).  Since  ions  involved  reported  MF-evoked  (hilar  stimulation)  chloride  larger  ( I t o e_t  reducing  ([CI  anion  a_l. ,  by  Ogata  1962  demonstrate effects  a  of  as  an  CI  pass  on  CA3  with  has  to  this  of  this  was  and of  iono-  action  the  latter  activity  which  CI  the  increase  in  propionate.  the  inhibition  f o r the  amplitude  i n ACSF  through  treatment  component  ]  PS  epileptiform  i n the  replaced  not  basis  [CI  1°  -dependent  the  in  increase  ) was  ) this  synaptic low  ]  does  GABA-mediated  suggested  the  the  (1975)  this  phore  of  Ogata  extracellular  To  the  [Cl~] In  of  in  were  1°  PS.  response, re-examined  here. In final  the  preliminary  concentration  sodium  of  propionate.  biphasic entry  effect  into  FIM-evoked  the  on  evoked  orthodromic  PS  response,  but  2.9Av).  Some  rain  rapid FIM  recovered  increase and  i n the  MF-evoked  mM  by  of  was  1°  this  a  MF  later  amplitude field  PS  the  control  ]  was  reduced  s u b s t i t u t i o n of  large  CI  ACSF  2-4  the  2°  responses and  number  potentials  there of  PSs  (Fig.  a  of  its  of  the  PS  which  a  with had  min  reduction  and  to  NaCl  remained unaffected  levels and  low  Within  (Fig. 2.9Aii)  the  to  [CI  responses.  there  MF-evoked  suppressed  7  Perfusion  bath  3-7  experiments  of  the  (Fig.  had  been  followed i n both  a the  2.9Aiii,vi).  35  Fig. perfusion A.  2.9  with  Control  shown.  Changes  ACSF c o n t a i n i n g  responses  The  i n evoked  initial  to  FIM  effect  responses  a decreased ( i ) and  of  low  MF  i n CA3  [Cl ]  following  (7  mM).  (iv) stimulation  [Cl ]  was  to abolish  are  the  MF  PS  in  o 2  (v) and  the  ]  B.  showing  truncated). there  .  MF-evoked A  C. an  charged  number  The  increase shown  with  of  ( i i ) . here  clock-like  evoked  activity  firing  (•  J  of  evoked  comprised  of  b u r s t s of  PSs of  the  following As  i n the  amplitude slower  regularity abruptly  PSs ( v i i ) in  recovery  increased i t (peak  a prolonged  Spontaneous a  was  epileptiform  activity  and  in  potentials  evoked  (ii)  the  increase  of  treatment  1  minutes  onset  (i).  at  the  a rapid  the  a n t i d r o m i c PS  ( i v ) ceased  i ni i i ) .  in  of  2-4  collateral-evoked  effects  response  PSs  the  (i),  ( i i ) the  response  ( v i i ) but  With  Within thereafter  of  Schaffer  and  Control  was  shortly  control  In  the p e r s i s t e n c e  response.  activity  The  amplitude ]  Note  number  spontaneous  responses.  [Cl  and  ( i i i , v i ) .  appeared.  the  field  amplitude  firing  PSs.  r e c o v e r e d and  occurred  [Cl  (ii)  MF-evoked  responses the  FIM  low  (iii)  as  were  amplitudes  treatment with  low  lower r e c o r d s of  (A)  and  number  of  PSs  in  p o p u l a t i o n b u r s t s (as i n sweep  speed)  ( i i i ) but following  also  this  and  disthe  seizure-like  36  37  Coincidentally  with  onset  of  at  regular  there  also  of  discharges (Fig. 2.9Avii).  PS  the in  MF  appeared,  the  1°  PS  which  had  i t was  also  increased  with  low  a  spreading recovery  after  the  to  a reduction  AD  PS  not  the  synaptic o  was  of  administration FIM-evoked although  by For  244+19%  were  the  20-25  in  positive the  concurrently  pre-treatment  submaximal  in  activity. this  PS  MF  1°  2°  of  increases slices  in 2+  where and  AD  amplitudes  (BIC)  Chapter in mM  to  which Mg  2+  );  the  expected,  the  blocked  2.11A);  of  6).  reduce  ACSF  PS  were  additional  ; 4.7  was  this  which  increase  slices  case  altered  and  from  bicuculline  ( F i g . 2.10A,  wave  resembling  [Cl ]  As  followed  the  (cf.  this  min)  slow.  low  Ca  activity  seizure-like  of Ogata,  mM  slices  (12-20  and  since  (0.2  1°  i n two  1968)  m e d i u m was  of  effect,  SC-evoked  1983)  repeated  of  the  ( F i g . 2.9Civ)  were  mM  and  Kawai,  results  response  three  and  actions  37  of  spontaneous  amplitude  ml  Similar  peak  PS  to  J  reduced,  (n=8).  example,  monitored  only  bursts  perfusion  a_l. ,  blocked  orthodromic  both  potential  AD  spontaneous  of  inhibition  seizure-like of  e_t  the  experiments  reduced  possibility  the  was  or  to normal  of GABAergic  firing  (n=4) PS  silence  that  the  Prolonged  evoked  confirming  activity  AD  (Yamamoto  returning  suggested  At  63+11%  (Snow  epileptiform  intervals,  the  electrical  change  Therefore  ]  an  depression  of  [Cl  caused  of  Although  does  monitored,  (Fig. 2.9Ciii)  period  by  ( F i g . 2.9B).  [Cl ]  discharge by  increased  evoked  the and,  the  MF-evoked  response  increased  apparent the  PSs (ca.  i n AD  responses having 2  mV)  the  PSs. were same  showed  38  A  low CI'/Co \ high -Mg 2  :  Si  FIM evoked-OD  20  J  10  20  low Cr/Co *,high 2  Mg  30  55  60  2,  MF evoked  I " 8  CD  6^  a  if)  o 4QO 0. 2  42  J  FIM evoked-AD 10  20  30  Time (min)  Fig.  2.10  exposure  to  mM)  increased  and  altered  ACSF  abolition B. FIM  In  of  Recovery  to  CA3  the  [Mg  ] by  low  mM)  (OD)  PS  amplitudes  (37  mM)  .  A. bar)  PS  levels  of  the  , low  caused  occurred  by  i n the  FIM  following [Ca  Perfusion  evoked  stimulation  increases  pre-treatment  ]  horizontal  alternate  parallel  of  [CI  (4.7  orthodromic  slice  medium.  changes  containing 2 +  (indicated  another  revealed  normal  ACSF  In  AD  a  after  ]  (0.2  of  the  reversible  FIM hilus  and  2 +  MF  stimuli. and 1°  returning  the PSs. to  39  increases  of  212+25% and  These MF  1°  a  and  the  reduction  tion  results  and  AD of  (2)  reflected  that  16%  more, the  they  33+8%  volley  in  the  least  ter  in  increase  portion  in  the  c o n t r i b u t i o n of  the  effects  reported  supramaximal reasoned  that  condition change were CI  -  of  2+  large  ,  high  in  in  as  in  ACSF  These  the  amplitude  of  part  the  medium.  following  AD  PS. ]  It  a  this  resistance  neurons  25-30  ml  which of  of  data  indicate  that  submaximal change  MF  of  a was  under  amplitude  a  in  situation,  [CI  of  for fibre  the  a  to  basis  this  number o f  ACSF,  produces  resistance  in  low  perfusion  that  Further-  the  with  maximal  the  probably  ACSF.  pre-treatment  in  30+3%.  in  PS  ion  a t t r i b u t a b l e to  by  a t t r i b u t a b l e only  cellular  be  inhibi-  presynaptic  addressed  a  than  reported  of  the  change  2 + -containing  Mg  of  change  the  1°  have  perfused  the  increase  Following  increased  increases  are  amplitude  an  the  cells.  resistance  elicited  s t i m u l a t i o n may  excited.  /Ca  a  in  stimulus  than  of  was  slices,  any  rather  response  PS  FIM  here  MF  resistance  slices  possible  three  CA3  amplitude  olfactory cortex  In  of  -dependent  larger propionate  change the  of  (1978)  specific a  of  Keurs  with  the  suggested  a  excitation  ]  increase  a t t r i b u t a b l e t o some f a c t o r o t h e r  The  manner.  the  at  [CI  increase  that  CI  and of  (1)  synaptically-mediated  non-synaptic  substitution  r e s p e c t i v e l y ( F i g . 2.10B).  indicated: was  a  Voskyl  a  PS  212+39%  this  1° the  low  the  or  AD  extra-  resistance. -  Prolonged  perfusion  (40-120  ml)  of  low  CI  2+ /Ca  ,  high  2 + Mg  -containing  ACSF  caused  an  increase  of  single  unit  40  , .. (vi)  40msec  I t 1 I IB I 5 0 sec  Fig. decreased 2+ [Mg  ]  2.11 tCl ]  ( 4 . 7 mM)  normal  medium  shock  elicited  Orthodromic (i)  The  size  of  evoked (e.g. filmed  of  ( i ) was  of  was  of  PSs  response.  caused in  [iv]);  now  one  oscilloscope  of  PSs  ( i v ) and  frequency  of  (vi).  these  was Since  ( v i ) have  an  of the r e c o r d e r ,  (v).  onset  amplitudes  of  no v o l t a g e  reduce  i i i ; note 1  h  the  afterdischarges i s shown with  i n the chart by  the  amplitude  After  spontaneous  attenuated  of  B.  ( n o t shown).  Coincident of  (iii).  and  ( i i and  afterdischarges in  been  trains  FIM  (iv).  i n the  traces).  by  PSs  effect  followed  increase  in  intensity  amplitude  potential  followed  trace  there  further  present  medium  i n i t i a l  which  increased  activity  i n normal  f o r these  and  A higher  t h e 1 ° PS  field  containing  additional  The  wave a  the  calibrations was  and  recovered  to increase  treatment  perfusate  ( 0 . 2 mM)  ( i i ) .  antidromic  MF  to  orthodromic  eliminated  the p o s i t i v e  afterdischarges trains  an  response in  changing 2+ and [Ca ]  mM)  transmission  number  change  After  the FIM-evoked  medium  Continued and  (37  control  altered the  A.  the  calibration  low  i n the evoked  firing  of  recordings response  i s given.  41  activity evoked  by  despite 3/6  and FIM  the  slices  (Fig.  a  progressive  increase  (Fig. 2.1lAiii)  continued evoked  2.11Biv)  blockade  responses and  discharges  lasting  observed.  A l l of  returning  t o normal  Increased  [K ]  and of  were  these  to  MF  followed  sec  by  PSs  stimuli In  after-discharges  consisting  ( F i g . 2.1lBvi)  e f f e c t s were  of  transmission.  activity  20  number  (Fig. 2.1lBii)  synaptic  spontaneous f o r up  i n the  of  PS  was  also  rapidly reversible  after  medium.  +  o The cellular  e f f e c t s on  concentration  examined.  In  response number  was  and  iv-vi);  of  regular  q  slices (see  with  the  activity  [K ] +  onset  of  comprised  were  PS  of  although  reversed  the  ( F i g . 2.12 PS  and  a  (Fig. 2.12i-iii).  epileptiform  bursts  also  MF-evoked  orthodromic  evoked  extra-  were  i t increased  of  2. 1 2 v i i , v i i i ) .  rapidly  the  the  1975)  observed  of  of  5 t o 10 mM  Ogata,  FIM-evoked was  change  1°  following  the  ((Fig.  the  also  a  from  +  of PSs of  of  ([K ] )  +  a d d i t i o n a l PSs  intervals  increased  3  unaffected  of  spontaneous  activity  of K  potentiation  Coincident  PSs  appeared  These after  firing at  effects  of  returning  to  ACSF.  Intracellular Stable neurons, the  each  amplitude  recruitment  normal  CA3  FIM.  some  recording  o f CA3  intracellular of  which  A l lof these  were cells  responses  impalements identified had  were by  AD  obtained  from  excitation  15  from  similar electrophysiological  42  Fig. the  number  potential,  2.12  Raising  and a m p l i t u d e  Spontaneous  (vii)  occurred  sweep  speeds  used  were  fully  to  normal  at  to  the  to the traces  from  the  Traces  1 ° PS  within  of  15  in  increased  at  column  field  virtually  bursts  of  PSs  the  slower  [ v i i i ] ).  These  minutes  calibration  row.  ( o ) was  (apparent  records  mM  and MF-evoked  comprised  i n each  time;  5 t o 10  i n the FIM-  intervals  reversible  i n each  o  o f t h e MF  obtain  same  [K ]  activity  regular  perfusate.  approximately apply  o f PS  but the amplitude  unaffected.  effects  the  of  were  bars  returning  obtained  to  the  at  right  43  characteristics  which  (Schwartzkroin the  resting  and  spike  megohms  and  (n=ll)  studies  the  field  electrode  Shown  cellular  FIM  potential PS  of  the  the  intra-  and  7  FIM  other shock  (cf.  13  an  EPSP  wave the  a  with  the  could  short  by  The  this  latency  be  discharge  respectively with  pathway. spike  to  low  intra-  increased an  observed Higher  followed  by  a  action EPSP  correlation  was  the  cell.  the  of  )  separate  the reversed  same  AD  of  I N  these  with  a  R  33+3  In  response  onset  (  (n=15),  correlated  typical  the  response.  mV  of the impaled  of which  and  intracellular  applied  changes  t o t h e MF  onset field  threshold with  and  t h e 2 ° PS  neurons  duration  response.  for firing  was  of  pathway  In 9 of these  whose  i n the  (Fig.  i s  resistance  and  between in  each  intensity by  an  EPSP  e t a l . , 1978).  cells.  aligned  were  i n which  activated a  (+S.E.M.) f o r  -63+2  extracellular potentials  cells  The  in  field  evoked  Dodd  stimulus  pulse)  values^  respectively.  50-100^xm  F i g . 2.13A  corresponded  the  of  within  (the amplitude  hyperpolarizing  interneurons  input  were  (n=12)  of  simultaneously  stimulation  EPSP  (RMP),  potentials  recorded  in  Mean  cells  mV  intracellular  positioned  intensity  these 84+1  those  1978).  potential  of  and  response  unlike  Mathers,  membrane  heights  were  In  potential  i n the h i l u s hilar  of  reached,  superimposed  a  recorded  stimulation  elicited  these  late  same  the a c t i o n on  by  were  p a r a l l e l e d the 5  produced  the l a t e  positive  cells  where  potential positive  was wave  2.13B). In  only  4 cells  did action  potentials  coincide  with  the  44  Fig. with  2.13  FIM- and MF-evoked  simultaneous  intracellular  cellular  recording.  (-66  RMP,  mV  cellular  another  MF-evoked  wave  height) (B) At  PS  case  which  higher  nature  EPSP  elicited  correlation  cellular  intra-  t h e two  t h e PS.  82 mV  B. I n  spike  height)  the onset  of the  a t t h e peak  of the  the action  potential o f t h e MF  EPSP  whose  that of the negative-going  potential. whose  cell  potential  an i n t r a c e l l u l a r D.  onset  A  30 V  1 ° PS  was s u g g e s t e d  the action  The  I N  MF  /  stimulus  85 mV  spike  was c o r r e l a t e d a s i n  s t i m u l a t i o n (40 V)  depolarization. between  of  C. A c t i v a t i o n  of the e x t r a c e l l u l a r of  an  action  (-62 mV RMP, 40 megohms R  intensity slow  /  I N  and  was a l i g n e d w i t h  EPSP  i n this  One  showed  t h e 2° P S .  and e x t r a -  height)  an  occurred  line)  of the action potential  preceding  of  sweeps  obtained  to that of the extra-  24 megohms R  with  subthreshold  spike  coincided with  (dotted  cell  t h e peak  traces)  EPSP).  discharge  RMP,  i nt h e h i l u s  a  with a  the  1°  i n this  80 mV  (reversed  i n the extracellular  elicited  /  superimposed  coincided  i n this  I N  corresponded  intracellular  extracellular  a  (-59 mV  consecutive  discharge  the  i n record)  cell  pathway  R  wave  produced  (truncated  onset  megohms  positive  stimuli  (upper  responses  A. F I M s t i m u l a t i o n e l i c i t e d  EPSP whose o n s e t  cellular  two  36  CA3  (dotted the  line).  antidromic  by t h e a b s e n c e o f  dashed  potential  line  indicates  and t h e e x t r a -  1° P S .  j  46  1°  PS,  (Fig  and  of  these  2.13C).  abruptly evidence  In  from of  recording  a  there the  the  was  an  others  membrane  the  action  potential  p r e p o t e n t i a l was  (not  underlying  EPSP  only  potential  (Fig  obtainable  in  2.13D)  even  with  2  arose and  no  high  gain  shown).  Discussion The  recording  substantial  amount  function  a  of  waveforms  The  (R)  the  of  by  the  Law.  The  ionic  conductances  in  field  potential is  therefore  the  conductance  the  synchrony  potentials force ity;  for iv)  polarity electrode  of  and  of  the  current  and  flux  resistivity the  of  space  affected  the  of  to  the  site  on  both  of  recorded  electrophys-  an  extracellular (I)  i)  Ohm's  amplitude the  cells  of  magnitude  of a of  activated; i i )  i i i ) transmembrane  affecting  on  by  the  activation  the  membrane  extracellular  the  through  defined  The  by  increased  the  the  the  as  response;  gradients the  of  from the  number of  p o t e n t i a l i s dependent  relative  required  current  membrane.  population  through  and  with  results  neuronal  concentration  ion the  change  experimental  is  in  generation.  measured flow  a  following  magnitude)  the  provide changes  characteristics  extracellular  extracellular  can  regarding  understanding  (V)  i s generated  resistance  an  neurons  underlie their  potential  electrode  of  polarity,  events which  potentials  information  influencing  (e.g.  iological  of  However,  factors  field  population  manipulation. the  of  the  neurons  driving  permeabil-  fluid.  l o c a t i o n of (e.g.  The the  dendrites  47  or  somata)  of  consideration Rail  the active of  and Shepherd Simply  waveform  this  topic  the basis  the generation  experimental removal  must  be  recorded  by o t h e r s  t h e C A l and d e n t a t e  no  difference those  responses efferent  between  little evoked  pathways  this thesis  involved  of  changes  evoked  of  field The  following  either been  anti-  than CAl  due  that or  or  to cellular  described  was  virtually  in this  There  however,  CA3  field  afferent  and  described  by e x t r a c e l l u l a r a  report  discussion  i s ,  the  various  recording  characterization  of  necessary. maximal  gyrus. by  PS  observed  activation  be r e c o r d e d  damage,  been  potentials  no f u r t h e r  of  i n CA3,  the  with  have  of the experiments  orthodromic  dentate  various  stimulation;  field  there  here.  some  could  of  criteria  regarding  deemed  of  which  the  These  stimulation  activity  amplitude  frequency  which  Instead,  the results  and s i n c e  an a n a l y s i s  r e s p o n s e s was  components.  investigations  and s i n c e  the events  and s o m a t i c  i s provided  in  smaller  regions  the  of the e x t r a c e l l u l a r  1 9 7 5 ; D u d e k e_t a _ l . , 1 9 7 6 ) i n  information  by  of  (1971).  e t c . ) and t h e c o r r e l a t i o n  the responses  potentials  relatively  these  (e.g.  potentials.  of the e a r l i e r  these  from  of d e n d r i t i c  in  of  deduced  detailed  i n the reports  and L l i n a s  i t svarious  (Schwartzkroin,  characterization  and  of  A more  to ascertain  of e x t r a c e l l u l a r ions,  applied  found  of t h e shape  manipulations  intracellularly  be  (1968) and N i c h o l s o n  on  information  conductance.  may  i t i s not possible  underlie  the  this  ionic  this  criteria  CA3  generally  i n t h e same  Although other  was  in  slice in  might the  have cells  48  appeared  healthy  comparable For  to  and  those  example,  a  Chapter  3),  (cf.  potentiation  could  as  Dingledine amplitude  e_t  CA3  dense packing  of  short  latency  PS  It  threshold  f o r the  short  represented suggested orthodromic either  a by  most  pyramidal  fibres  its  likely cells in  reflects  spread  of  the  where  high  short  or  PS  intensity  That  also  direct current  response  by  term  a  to  a  the of  the  (see  stimulus  also  latter CA3  to  excitation  response  to  since  there  5.14A).  of  block  This  PS  invasion  of  of  efferent that  cells  were the  was  demonstrate  unlikely  activation  Kandel  neurons  altered  failure  is  CA3a.  s u p e r i m p o s e d , on  antidromic  It  Fig.  in  less  a  s t i m u l a t i o n evoked (e.g.  the  potentiation.  the the  smaller  could  that  ACSF  1981).  evoke  higher  excitation  FIM.  FIM  case  was  1978).  following  stimulus  latency  the  those  et. a l . ,  the  related  PS  to  Prince, that  the  latency  represents  the  and  term  active  (Kandel  FIM  long  the  vitro  applied  persistence ' in and  and  comparable  be  to  present long  apparent  usually  al. ,  pulse-  coursing  may  and  is particularly  latency  e_t  were  gyrus.  was  passive  suggested  longer  non-synaptic  potential  the  was  transmission  paired  therefore CA3  Dodd  a  dentate  1960)  Wong  responses  and/or  the  inhibition  i_n  1980; have  shock  wave.  a l . , 1961;  and  vivo  wich  positive  et  ir\  field  electrical  and  the  cells  ( 1980)  cells  and  impaled  Dudek,  a_l. ,  of  An  of  and  CAl  (Andersen,  shown  occurring  MacVicar  pulse  be  reported  in  GABA-mediated  paired  properties  electrophysiological activities  observed  robust  membrane  1961;  displayed  by  this the  situations  long  but  not  49  A  synaptic origin  associated  PS  was  2+  suggested  FIM-evoked  by  the  positive  blocking  wave  effects  and  of  low  of  the  2+  Ca PS  f o r the  , high  Mg  -containing  following  cellular  paired  recordings  medium  pulse  and  and  the  tetanic  confirmed  the  potentiation  stimulation.  orthodromic  Intra-  nature  of  the  mediating  the  response. The  identity  FIM-evoked  of  the  orthodromic  response  is  expected  from  contributions  would  fibres  et. aJL. , 1 9 7 3 )  are  (Mosko  slow  trains The  of  and  are  high  response  be  frequency  may  also  excitatory  a l . , 1971;  MacVicar  seems  preceding  AD  Save  an  was  f o r the  potential  which  uncontaminated reported  as  Andersen,  1960)  (Blackstad, observed seems fibres.  especially  likely  to  No,  of  septal  only  after  Nicoll,  1983).  activation 1934,  and  of  a  Lebovitz et  Wong, 1983) cases  but  where  a  observed.  be  of  a presynaptic fibre  evoked  preceding  occurring  1956).  the  Miles  characteristics  after  slices  e x p l a n a t i o n i n those  absence  could by  to  minor  depolarizations  ( C o l e and  ( L o r e n t e de  Only  activation  cholinergic  Dudek, 1980;  not  certain.  the  attributed  pathway  electrophysiological  not  i n hippocampal  be  and  pathway(s)  stimulation  unlikely  PS  since  evident  recurrent  this  afferent  in_  by AD  vivo  stimulation Accordingly  with  reflect  low the  of  FIM  volley,  orthodromic  stimuli  activity  and  that  Hamlyn,  1957;  of  commissural  orthodromic  intensities activation  of of  was  resembled  and  the  field  which  (Cragg the  the  pathway  excitation  FIM  stimulation  these  commissural  50  The stratum with  response  evoked  radiatum of  FIM  CAl  presumably  a  o r CA2  stimulation  related  by  save to  stimulating  closely for  the  a  Experiments  in  the  latency  short  reflecting coursing and  the  low  the  from  Ca  to  CA3  to  CAl.  secondary  PS  with  on  the  suggested  to  be  Mg  be  activation  dependence  longer  that  the  observed  latency  of  conduction  onset  distance.  2+ , high  PS  resembled  longer  2+  electrode in  -containing antidromic  of  Shaffer  The  blockade  the  latter  excitation  of  involved  in  ACSF  and  most  the  likely  collateral of  the  fibres  positive  treatment  afferent  indicated  wave  indicated i t s  fibres  such  FIM-evoked  as  those  orthodromic  response. The  1°  positioned an  EPSP,  Wheal  i n the  which  a  paired  of  and  presence  unaffected  tion  often and  because  cell  body  PS  layer  stimulating  interpreted  of  (Andersen  (Lanthorn the  as  consisting  and  Cotman,  proximity  ( J o h n s t o n and e_t  a_l. ,  electrode  of  1981;  the  Brown,  1971b;  of  MF  1983),  Yamamoto,  long an  q  term  Thus  the  of 1°  i n medium w h i c h refractory remained  potentiation  orthodromic  contribution  unmistakeable.  been  a  Q  and of  has  by  Ueno, 1976). A l t h o u g h i n some s l i c e s the 2+ 2 + [Ca ] / h i g h [Mg ] and t h e o c c u r r e n c e o f  low  pulse  additional  was  1983)  to the  Ogata  effects  the  hilus  superimposed  1972b;  evoked  i s negative-going  ejt a _ l . ,  termination and  response  to  component a  PS  were  consistent  i n the  non-synaptic  was  partially  with  response,  the  process  was  blocked or  wholly  blocked other synaptic excitations,  paired  virtually  pulse  inhibition  unaffected  in  [  K  +  l  r  or i  potentia-  which  caused  51  large  increases  of  component d i d not  FIM  orthodromic  appear  simply  PSs.  The  Ca  to represent  a  -independent  large  amplitude  2+ presynaptic  fibre  volley  since  in  low  -  [Ca  ] /[C1 ]  ,  Q  high  2 + [Mg ] the substantial observed  a m p l i t u d e o f t h e r e s p o n s e i n c r e a s e d up t o 4 0 0 % and r e d u c t i o n s o f t h e 1° PS o f MF^, a c t i v i t y c o u l d b e 2+ 2+ i n low [Ca ] , h i g h [Mg ] ( F i g . 2.7B). Finally,  support  for  Q  a  non-synaptic the  1°  (probably  response A  by  variable  was  AD)  was  electrical  synapse  hypothesis  is currently  and  but  Schwerdtfeger  no  evidence  Sarvey  from  CA3  exist  this  information  logical  data  that  AD  an  applied  hilar  large  1°  PS  actions of  any  and  to  variations  in  (L-APB)  the  (an  stimuli  with  there  being  activity ranged  effect  on  or and  have  Cowan  shown  the  by  refute  dentate  Yamamoto can  reported  mediated  the  of  an this  (1973) that  gyrus  a may  electrophysio(1972b)  occur  with  effects  of  a  suggest stimulus  region.  hilar  which  response  be  with  by  generation  support  and/or  and  analyses.  Gottleib  cells  by  orthodromic  the  1°  antagonist  i n invertebrates [Cull-Candy  evoked  consistent  MF to  together  CA3  in  (1983)  hilus  here of  -4-phosphonobutyrate excitations  the  presented  i n the The  to  activation  i n the  available.  and  orthodromic  intracellular  suggested  projection and  by  component  (1972b)  of  excitations  provided  non-synaptic  Yamamoto  contribution  in from  the  et  et. a l .  extents  i t s generation. a 1°  complete PS,  of  abolition  whereas  glutamate  a l . , 1976] ) on  (Yamamoto  variable  L-2-amino-  AD  of  Thus to  r  the  1983)  are  anti-  and  L-APB  had  the  responses  absence were  52  consistently underlie  unaffected.  the  (Lanthorn  apparently  and  1983)  of  a  1° PS  i n CA3;  1981)  baclofen  results  similar  explanation  contradictory  Cotman,  depressant  The  A  and  absence  action  of  had  effect  of  no  reports  baclofen  experiments  on  the  presence  (Ault  and  Nadler,  the  hilar-evoked  PSs.  assessing  the  effects  2+ stimulus gether that the  frequency  with  the MF  the  and  intracellular  p o s i t i v e wave  1°  response  neurons.  This  excitation  of  (Amaral,  1978;  either  of  However, records  of  this  al. ,  1978;  that  the  discharge  (cf.  usual of  Spencer, firing  of  the  3)  1961b; observed  CA3 or  and  Dudek, to  PS.  rapid  which  onset  of  overrides  depolarizing after Wong in  with  and low  be  Prince, 2+ [Ca ] ,  due  from  the  display  and  intra-  CA3  to  the  hilus  bursting  intracellular Prince,  and  1981).  extracellular  et_ a _ l . , 1 9 6 1 b ; D o d d et_ Misgeld  this  et  a l . , 1982)  stimulation  was  the  case  a GABA-mediated and  masks  any  p o t e n t i a l (DAP) 1981). high  of  associational  brief  orthodromic That  followed  activation  cells  1980;  to-  1983).  (Wong  the  ]  which  may  Sarvey,  [Mg  unequivocally  CA3-CA3  (see a l s o Kandel  single  to  a  from  PS  afferents  current  response  a  Chapter  influence  report  MacVicar  attributable  or  and  indicated  pathways,  spontaneously  i t i s evident  ]  synaptic  single  depolarizing  [Ca  activation  of  2+  associated  the  fibres  ACSF  the  records  S c h w e r d t f e g e r and  normal  behavior  of  the  reflect  perforant  commissural  In  and  orthodromic  fibres,  pulses  changes  also  of  on  AD  may  is  the  may  be  inhibition excitatory (Kandel  and  The A D - e v o k e d b u r s t 2+ [Mg ] may be attri-  53  butable  t o both  consequent  the abolition of synaptic  expression  of  the  DAP  and  inhibition  with  the  increased  neuronal  2+ excitability haeuser  related  and  Epileptiform subsequent (Hotson  Others an  following similar  antidromic  attributable  efferent  excitability of  may  [Cl]  have  fibres  spontaneous a  population  (Prince, 1982, evoked  of  and  activity  T  to the loss  i n the resistance  of  less  permeant  neurons  o f CA3 n e u r o n s  agents  comprised  spontaneous  (Schwartzkroin  i nthe  i s  the  following  the  burst  Prince,  firing,  population  effect  of GABA-mediated 1978)  and,  of  bursting burst,  1978; Traub  an  their  appearance  of the synchronous  (spontaneous  to the reduction and  ions;  facilitated  of  SPB)  a n d Wong,  In the present study a decrease of [ C l]  attributable  of the  membranes  an i n c r e a s e  and/or  of  1983) have  neuronal  by  1978; S c h w a r t z k r o i n and P r i n c e ,  1983).  the  o f somata.  convulsant  activity  and  was t h e p o t e n t i a t i o n  produced  activated  A c h a r a c t e r i s t i c feature with  of AD  neither  n o r t o a change  here  1980).  1981) i s t h e most  (Neumcke, 1 9 8 3 ; Ransom a n d C o n n o r s ,  invasion  treatment  Prince,  ( F i g . 2.7B).  the replacement  of  (Franken-  afterhyperpolarization  e f f e c t of low [ C l ]  was  ]  stimuli  C  reductions  increased  effects  number  AD  f o r the large  inhibition  reported  by  and  and P r i n c e ,  medium  PSs which  tissue.  elicited  Hotson  [Ca  1 9 8 0 ; Wong  unexpected  GABAergic  1957;  of  activation of a long-lasting  i n this  An AD  activity  basis  observed  of  Hodgkin,  and P r i n c e ,  probable,  to the decrease  most  caused likely  inhibition  consequently,  an  54  increase  in  excitatory As  the  efficacy  and  SPB  recruitment  (Zuckerman and G l a s e r ,  indistinguishable induced  a  of  recurrent  circuits. in v i v o  be  or  from  those  by a p e r f u s i o n activity  responses  of  alternative  o f ACSF  here  again  of  multiple  comprised  impairment  observed  synaptic  mechanisms  1968) SPBs w h i c h  i n low  i n which was  [CI ]  [ K  +  PSs.  could  ] was with  Although  inhibition  a  cannot  f o r the actions  also  increased  Q  associated  were  evoked  selective  be  precluded  of increased  [K ]  are:  +  o (i)  a  decrease  neuronal  o f membrane  excitability  reduction  of  attenuation  the  (Schwartzkroin K  o f membrane  an e f f e c t  1965;  Cooke and Q u a s t e l ,  dependent  on  Spencer,  However synaptic  did  prevent  the  Snow  that  non-synaptic  the  characteristics  observed  of in  Dudek,  study  slices  an  conductance;  +  Quastel,  investigators that  SPB  activity  excitation  1983).  (e.g. SPBs  i n some  Consistent .were  involved  burst  were  exposed,  was  and  of  SPBs  i s and  1982,  shown  Haas,  to  1982) (see  the i m p l i c a t i o n  and  different  example,  i n CA3  Wong,  in their  duration  clearly for  and  slices with  the  (Dichter  medium w h i c h  (cf. Jefferys  development  processes  these  causing  (Gage and  e t a l . , 1973; Traub  transmission  also  interval)  and  Ayala  of  synaptic  i n the present  prevent not  by s e v e r a l  synchronization  recurrent  1969b;  release  ( i i ) a  1973).  has been suggested and/or  by a K  increased  1978);  gradient  subserved  on n e u r o t r a n s m i t t e r  initiation  1983).  events  t o an  and P r i n c e ,  concentration  +  (iii)  It  potential leading  to  generation interburst from  those  increased  55  [K ]  but  indicate  retaining that  as  intact  in  the  synaptic transmission.  CAl  region  ( T a y l o r and  These  Dudek,  data 1982;  2 + Jefferys  and  processes 1981), 1981)  and  charges. normal  also  such  as  1982;  field  extracellular spontaneous Since  contribute has  and  these  medium  transmission  Yaari  electrical  electrical  produce  in  Haas,  to not  a l . , 1983)  ion  Ca  -independent  transmission (MacVicar  (ephaptic) changes  highly  events  i t is  et  interactions (Yaari  probably  Dudek,  (Jefferys,  et_ a _ l . , 1 9 8 3 )  synchronized  would  and  neuronal  also  reasonable  to  suggest  SPB  activity  in  slices  been  blocked.  be  that were  can dis-  operative they  may  synaptic  56  CHAPTER POSTACTIVATION  3  INHIBITION  I N THE  CA3  REGION  Introduction Following stimuli,  pyramidal  formation part,  et  Chan-Palay, to  a_l. ,  the  In  this  region  presented.  (GABA)  Curtis  there  inhibition  the  hippocampal  anti-  and  a_l. , 1 9 6 1 ; Kohler  an  action In  f o r an and  gyrus  and  i s attribut-  1980).  (Nicoll  1982) and t h e d e n t a t e  of  et  inhibition  i s evidence  i n  interneurons  and t h e s h u n t i n g  i n CAl  results  of  1970 ;  of a  addition  orthodrom-  Alger,  1981;  (Thalmann and  t o t h e a c t i o n s o f GABA. electrophysiological  evaluation of the i n h i b i t i o n  after  orthodromic  (Kandel  e_t a _ l . ,  i s not a t t r i b u t a b l e the  or  i s due, a t l e a s t  ( D i n g l e d i n e and Langmoen,  1982) w h i c h chapter  in  which  T h i s GABA-mediated  and N i c o l l ,  pharmacological CA3  acid  hyperpolarization  ically-activated  Ayala,  anti-  by a r e c u r r e n t p a t h w a y  GABAergic process,  Newberry  by  cells  inhibition  1964 ;  1983).  conductance this  granule  ^-aminobutyric  Andersen  to  an  to the e x c i t a t i o n  able  activation  and  exhibit  releasing  CI  their  orthodromic  and  o c c u r r i n g i n the stimulation  are  57  Methods The rats  were  where are  70  hippocampal  prepared  the  and  techniques  also  slices  obtained  maintained for  outlined.  as  intra-  An  from  29  described  and  electrode  male in  albino  Chapter  2  extracellular  recording  in  recorded  CA3a  (R,-) o  responses  evoked  (MF)  pathway  (Sg)  to  (AD)  in  In was  stimulus  hilus  alternate the  same  or  slice.  interval  The  paired  (C-T  ations)  of  stimuli  the  C  and  unconditioned applied  in  F I M , MF  the + MF  C  T  The evaluated  T  methochloride, (l-5xl0  _ 6  M,  preceded an  input  the  varied  fimbria  antidromic  mean  with  paired  difference  between  MF^  +  region  +  and  (3-6  of the  test  MF  With as  the  determin-  conditioned  Paired T  the  heterosynaptic  the  FIM ,  of  C  +  and  s t i m u l i were FIM^,,  AD  C  +  AD^.  nature  iontophoresis (pH  C  (C)  intensity  calculated  amplitudes  responses;  (0.05-0.1  20-4000 msec.  was  FIM  5-10ml).  fibre  conditioning  ,CA3  between  inhibition the  a  conditioning  combinations:  25mM  the  or  identical to  the  by  calculated.  AND  mossy  stimulation  r e s p o n s e s was  GABAergic by  test  between  between  percent  to  (T)  stimuli  T  or  S_)  a  (using  i n t e r v a l ) was  difference  the  orthodromic  different  homosynaptic  percent  T  a  2.5,  to  (2-4mV)  occasions  stimulation)  either  (PS).  experiments,  to  volleys  (Fig.  submaximal spikes  the  on  stimuli applied  the  produce  population  Hz)  by  3.5)  of  the  of  BIC  in  150  inhibitions in from mM  a  NaCl,  CA3  solution or  by  of  was the  perfusion  58  Results The FIM  effects  responses  T  conditioning at  100-1800  are  stimuli  slices  there  was  and  74%  stimuli  (MF  C  +  FIM )  (11  interval.  Activation  active  CA3 Fig.  least  that  evoked  post-firing amplitude few  neurons  least the  two  the  of  a  few  either  msec) phase  of  pathways  the  by  5  the  inhibitions  of  60%  heterosynaptic  at  (1  of  that  the  slice)  or  same  C-T  the  also  not  properties  produced  a  spontaneously  (20  inhibition.  from  late  the  small  indicates  were two  possible  to  msec)  the  or  Secondly,  at  that  i n d i c a t i o n that  inhibition  in  observations.  later of  i t was but  at  involved  activate  i n <5%  further analysis)  the  since  processes  arose  inhibition,  attributable to  first  distinct  early  was  The  i t was  or  of  of  activation  slices  early  in 3  amplitude  MF^  responses  evoke  msec,  msec)  illustrate  evoked  from  the  these  <100  homosynaptic  effect  responses  FIM^,  the  no  occurring  intervals  with  FIM^,  membrane  (which were e x c l u d e d only  either  either  shown).  3.5C  of  50  the  contrast,  activated.  of  C-T  2 cases  (300-1000  mechanistically  in  entially  PS  were  inhibition  First,  of  in  heterosynaptically,  changes  of  other  was  not  3.4A,B a n d  With  inhibition at  separated increase  inhibition (data  the  However,  In  slices)  cells  of  submaximal  A,B,C,E.  by  there  T  3.1  FIM^)  i n the  inhibition  long-lasting  Fig.  on  Thus w i t h  observed. +  stimuli  C  were a p p a r e n t .  35-75%  whereas  were  in  similar.  C  a  o r MF  c  magnitude  actions (FIM  response  T  the  m s e c was  paired  ™  F  shown  shock  msec, d i s p a r a t e  FIM  of  differ(200-300  the  slices  possible  not  both  to  even  59  A.  C.  D.  C - T Intervol (msec)  Fig.,. by  FIM  3.1  intervals. interval a  FIM  Mean  v  on  The  FIM^, a c t i v i t y  i s indicated  with  (+_S.E.M.)  number  response  C  evoked  A.  stimuli  C  FIM  of  at  slices  in brackets.  ( i ) and  t h e same  stimulus  AD^  showing  the e f f e c t s  ponses. Although  The  was  figures, labelled  traces arrow  are below  C-T  after  a  longer  C-T  f o r each  C-T  of  300 m s e c  of FIM  T  (iv-v)  a n d AD^  response  occurred  reduction MF^  the record  only. of  2  In  this 3  and  i s shown  D.  elicited records FIM  T  res-  in  (vii).  with  either  activity  at  400  subsequent  consecutive  i n d i c a t e s the time  i n msec  on  i s shown  FIM^  C,  Filmed  a t 20 m s e c  of  or  later.  responses  (i-iii) FIM  of  FIM^, a c t i v i t y ( i i )  E.  comprised  interval  and  respectively.  inhibition  activation,  observed  stimulation;  o f MF^,  unconditioned  comparable  conditioning msec  stimuli,  msec  evoked  Oscilloscope traces  intensity  by  inhibition  examined  -inhibition  as f o r A but i n h i b i t i o n  and  20  B.  Conventions MF^  percent  and  in brackets.  sweeps; type  of  60  when  FIM  or  MF  conditioning  stimuli  of  a  high  i n t e n s i t y were  applied. Fig. of  AD  3.ID  elicited  C  electrode. the  inhibition  was  occurring  with  of  or  actions  on  a  conditioning again  a  relatively  were  3.2A)  stimulation  pattern  either PS  of  paired  MF  little  of  T  the  inhibition occur,  20  than  and  these  pathways. in  the this  inhibitory  C  i n some  slices.  comparisons be  that  address  AD  C-T  (Fig.  msec  to  either  the  MF  activity  was  evident  was  at  any  C-T  interval  of  the  test  response  stimuli  the  To  when  s i m i l a r with  or  AD  input, at  400  a but  msec  only.  of  large  Although  characterized  could  FIM  inhibition  despite  at  of  stimuli  C  obtainable.  which  inhibition  of  similar  variation  made c o n c u r r e n t l y one  that  longer  o r MF  slices. C  at  duration  t o the FIM  MF  shows  However,  measured  a  separate  m s e c was  stimuli.  the  response  a  F i g . 3.1A,C  reflected  of  FIM^  through  shorter  shock  both  was  example  the  of  between of  the  i n t e r v a l s <100  have  activity  T  to  inhibition  inhibition  t h e MF  With  (Fig.  may  reduction  following  the  on  applied  conditioning  comparison  FIM  showed  C-T  a conditioning  representative  3.IE)  AD  shock  F i g 3. I D  at  T  actions  l e s s e f f e c t i v e and  the  possibility  2°  MF  differences  potency  the  FIM  of  antidromically  intervals  the  a fimbrial  of  FIM,  elicited  These  by  Comparison  following  A  illustrates  the  FIM  of  at  of  the  not  rigorously  the  2°  stimuli.  or potentiation whereas  1° PS  reductions  i t was  inhibition paired  there  (e.g.  MF  PS Thus  2°  which  investigated, resembled at  20-50  F i g . 3.2Bii)  5 0 - 1 8 0 0 m s e c C-T  PS  that msec  of  the  intervals  this  61  Fig.  3.2  A.  Paired  pulse  stimulation  to  CA3. ; I l l u s t r a t e d  i s t h e mean  of  the amplitude  t h e 1 ° PS  20-900  msec  records MF 2°  PS  was  When  potentiated  respectively.  unaffected  substantially  by  a  but  AD^  E.  response  AD  preceding in  MF^  B, C a n d E .  percent  the test  B.  and C.  interval  inhibited  at  t h e 2 ° PS  t h e MF^,  D.  of  The  produced  occurring  by  a  without  40  field  of the  and MF  preceding  400  msec  response.  ( i i ) the  1°  potential  (mean  PS was  +S.E.M.) o f MF^  ( i ) and with  The c a l i b r a t i o n b a r a p p l i e s  filmed  whereas the  response  inhibition  evoked  These  ( i ) The u n c o n d i t i o n e d FIM  activity  (ii).  a t a n y C-T  input  reduction  response  response.  conditioning  reduced.  submaximal The  t h e MF  t h e r e f r a c t o r i n e s s o f t h e 1 ° PS  to inhibition  preceded  of  the c o n d i t i o n i n g  illustrate  intervals  was  after  response  T  of  (+S.E.M.)  of  to the  stimuli. ( i i i )a records  62  2  response  had MF  similar evoked  CA3  at  last  3.2D  of  the  at  the  an 20  duration  of  somewhat  shorter  this  responses  membrane  than  subthreshold  f o r the  ca.  achieved up  to  but  was  in  the  records  of  amplitude parallel  a  fact  of  a n t i - • and  of  PS  also  of  the  interactions in  of  responses.  effect  AD  T  of  tested.  The  activity  was  11  with  CA3  cells  showed  input  at  HP  the  an  the  intensity  both  reached  phase  the  absence  response "is evident  HP  was  orthodromically  a  of  of  CA3  peak  at  membrane HP  which  and  late  a discernible  i n the  did  of  persisted for early  figure also  the  (HP)  pre-stimulus  second  the  intensity  that  action potential elicited  That  of  inhibited  (FIM)  mV  early  latter  the  orthodromic  150-200 msec and  in  on  3.1C).  -55  an  AD^  5 slices  with  FIM  a  at  occur  stimulus  1°  hyperpolarization  by  phases  Since  or  towards  of  the  the  substantially  and  than  MF  This  two  stimuli  inhibitory  impalements  F i g . 3.3C. the  input  occurring  ( F i g . 3.3A,B).  in  of  inhibition  biphasic  recovery  not  i t was  s i t u a t i o n s the  MF-evoked  as  any  i n each  interrupted  msec  could  of  discharge  a maximal amplitude  700  phases EPSP  both  msec  potential  by  the  but  s t i m u l a t i o n on  C  greater  either  followed  20  that  intracellular  In  MF  absence  potentials  neurons.  of  MF^-induced  of  EPSP  determinations  intervals  stimulation  2°  FIM^,  C).  ( c o m p a r e F i g . 3.2D  Stable  an  the  msec w h e r e a s  1 0 0 - 5 0 0 m s e c C-^T  test  inhibit  effect  shows  response  T  to  (Fig. 3.2Biii).  p o t e n t i a l ( F i g . 3.2  examined  Fig.  depressed  actions  The  AD  was  oscilloscope  shows  not  that  the  increase  in  inhibitions  of  increased. evoked  63  I  I  O  100  I  I  200  I I  300  400  500  msec  c,  v  10 20 30 40 50  1.0 mV  .MF 40msec Fig. response  3.3.  in  two  CA3  stimulation. biphasic latency the (R  I N  to  peak  of  a  20  V  increased,  for  the  an in  EPSP  the  RMP  respect i vely.  and  both a  was  IN  in  the  and  current  of  each  HP. the  the  and  input  C.  of  the  As  there  for  shown)  membrane  stimulation.  the  present  and  stimulus  was  a  non-parallel  of  each  component  of  the  HP.  this  cell  were  mV  and  33  -58  the  resistance  in  were  was  in  a  Oscilloscope  MF  HP  (B)  EPSP  values  (not  change  intensities  elicited  FIM  the  The  record.  of  and  injection  of  phases  (A)  similarities  the  by  intracellular  following  of  (RMP)  the  MF  depolarization.  amplitude R  Note  recordings  various  by  was  component  right  stimulus,  preceded  there  potential  the  of  subthreshold  (HP).  determined  to  recordings  to  case  each  following  not  increase  for  intracellular  potential After  each  was  indicated  traces  cells  membrane  which  are  were  In  chart  hyperpolarization  resting )  Moving  Values megohms  64  hippocampal  pyramidal  mediated,  the  actions  methochloride iontophoresis addition  to  intervals or  only  the  within  100yam  perfusate  with  could  been  GABA  antagonist  et  a_l. ,  1971)  the  cell  paired these  inhibition  FIM  i t s iontophoretic  amplitude  and  r e c r u i t e d a d d i t i o n a l PSs  (Schwartzkroin  and  Prince,  not  Fig.  6.6).  stimuli 1°  PS  observed  i n the  The  unchanged  results  (pre-treatment  of  unaffected  78.1+5.6%,  to  min  termination  the To  the  near  changes  in  administration  of  the were  also  C-T  facilitation BIC  for  examined  of  5-30  the  min)  or  FIM-evoked  ( F i g 3.4A,B;  employed  appear  however  of  the  the  3.5C)  responses here  (cf.  following  amplitude  with  of  MF the  indicated at  post-treatment  that measured (pre-  BIC  inhibition  60.7+7.2%,  and  levels  at  20  msec  200-300 msec  n=ll). occurred  an  3.7+8.5%,  post-treatment  respectively,  pre-treatment  approximate  BIC  experiments  whereas  inhibition of  the  increased  and  its  ( F i g . 3.4A).  inhibition  or  slice,  abolition  m e a n + S.E.M., n = l l )  of  could  by  early  Spontaneous burst  doses  PSs  BIC-treated  complete  62.7+9.1%  the  Multiple  remained  almost  with  1980).  or  by  activity.  T  (50-200nA  the  GABA-  either  at  either  PS  were  layer  Because  stimuli  be  bicuculline  experiments with of  increased  to  applied  body  were examined.  occur  a p p l i c a t i o n BIC  reported  the  of  homosynaptic  MF^-evoked  orthodromic  of  (Curtis  Following bath  have  (BIC)  the  inhibition  cells  was  values  Recovery within  of  10-25  drug a p p l i c a t i o n .  the  duration  pattern assessed  of in  the 4  of  the  GABAergic  inhibitions slices.  In  process  following  BIC  each  BIC  case  65 A  Pre - treatment  FIM  MF,  Bicuculline  FIM (20)  FIM j (300)  T  treoted (lOOnA.IOn  Recovery  _F  m V  4 msec  B.  Bicuculline  treated ( 2 . 5 x I 0 " M . 15ml) 6  4 .-msec  3.4  Fig. MF-evoked  -The  early  and  actions  late  FIM; r e s p o n s e  situations  shown  potential used or  shown  group  msec  stimulus  intensity  MF^-induced  As  responses  are  not  persistence with  explanation).  of  for A shown  of  the  burst  of a the  BIC  and late  firing  from  16  to allow PS  20  A.  MF^  a  MF^  applied  inhibition i n t h e MF^  in  during  comparison  similar of  column)  The  V  to  that  treatment  period. 13  The  field of  (third  f o r each  to  the  treatment  The  that  stimuli  pre-treatment was  on  i s representative  strength  BIC  three  column.  minute  primary  except  the  Traces  2-3  (BIQ)  activity.  elicited  later. a  FIM^  of  first  decreased  situation;  B.  correlated  was  application  changed.  The  over  inhibition  pre-treatment  the  column)  obtained  iontophoretic  i n each  t h e FIM^, r e s p o n s e  (fourth  were  in  of  i n t h e s e c o n d .column  to condition  300  b L l e u c u 1:1 i n e  inhibition  unoondifcioned is  of  and  of  FIM the the  i n the was  not  recovery  i n the p e r f u s a t e . this  response  case  was  not  (see text  for  66  Pre - treatment  2  Fig.  3.5  A,  B.  FIM )  inhibition  at  during  (*•-•) a n d  after  BIC  -  T  (A  cases, of  125  a  than  affected  at  the  200  intensity  in  BIC  of  BIC  the  of were  min; the  whereas  was  a  now  from  Note  these  field fully  later  FIM^,  early  was  potential reversible.  for  Conventions  of  in  low  as  to  23  V  MF^  (o-o),  iontophoresis 25  min). at  in  activity  inhibitions the  before  In  C-T  and The  un-  3.4A. (15  20  In  V) but  MF not  FIM^  activity  FIM  stimulus  during  the  occurred  response.  both  was  Fig.  at  of  intervals  intensity  inhibited.  +  '  inhibition  inhibition  26  ' '  the  occurred  the  decreased that  nA  situation  the  interval  only)  150  (MF  ;  i -j  intervals  inhibition  inhibition  reduced  C-T  -  C.  he t e r o s y n a p t i c  :  in.A B  control  an  msec  application. absence  of msec  produced  msec.  6  (A-A,  potentiated.  slice  stimuli 200  or  60  pu'l.s e  different  for  reduction  less  this  nA  Paired  msec  The  drug in  the  actions  67  caused  a virtual  action  of  the  msec  60  BIC  abolition  to C-T  potentiated  the  the  reduced  200  msec  which  recovery  was  increase  can  of  a  can  an  C-T  an  beyond  BIC  often  intervals,  in  i n the s l i c e  from  were  derived,  iontophoresis  of  and  revealed  an  at  neither  potassium  i n the  However,  although  i n MF  inhibition  pre-treatment  tence  of  inhibition  were s m a l l  suggesting  that  these  bursts  were  3.4B,  3. 5 C )  where  following The treatment of  does  HP  not  would  the  changes  kainic  acid  in  seem  with  i n Figure  of  calcium and  firing  nor  Nicoll,  1980)  activity  (cf.  bursts  exposed  underlie the PSs  action  activated  in  slices  to  (1)  the  to  was  BIC  comprising the  a few  i n a number o f  was  persis-  t h e u n c o n d i t i o n e d FIM  i n only  cells  cases also  and  PS, (2)  (e.g. Fig unaffected  BIC.  inhibition and  bursts  evoked  inhibition of  a  evoked  occur  late  i n the  summarized  of  since:  observed  the a d m i n i s t r a t i o n  are  not  i n comparison  any  by  in  (Alger  responses  C  this  late  caused  inhibition  F i g . 3.4A) the  cells  conductance  an  evident  HP  (e.g.  action  longer  pyramidal  e l i c i t  produce  6).  sometimes  bursts  extended  Indeed,  For example,  inhibition  apparent  never  A,B) .  the  F i g . 3.5C  d i s c h a r g e of  potentials  Chapter  at  a t 20 m s e c b u t  response.  The  which  inhibition  dramatically.  the e a r l y  inhibition  ( F i g . 3.5  inhibition  records of  BIC  the  the  interval  some c a s e s q u i t e which  reduce  of  i n CA3  compared 5.15.  which  with  a  followed  BIC  disinhibitory  68  Discussion The chapter  major  was  the  finding  occurrence  inhibition  f o l l o w i n g the  inhibition  was  blocked or  with  The also  slice  and  in  also  in  the  recorded  early to  (Knowles  HP  be  inhibition  the  conditioning of  a  Nicoll,  was  no  evidence  ity  by  a  aptic paired  MF  C  at  but  On  that  which  hippocampal  activation  experiments  of  assessing  suggested 60  the  which  a  duration  msec.  This  i t was  absence This  of  has  been  shown  possible to  any  is  intracellularly-  picrotoxin-sensitive  occasion  the  the  the  than  of  and  unaffected  appreciable  observation  GABAergic pathway  IPSP elicit PS  suggests  such  as  by  in the  occurs  in  1982). even d u r i n g  any  time  stimulus.  potentiations MF  here,  was  extracellularly-deter-  less  -dependent  in  of  duration  This  which  either  of  to  the  inputs.  resembled  treatment of  response.  Interestingly,  of  this  antagonist.  gyrus  due  in  a long-lasting  component  GABA  results  event  feedforward  ( A l g e r and  the  course  et. a T . , 1 9 8 2 ) .  early  CAl  Cl  of  afferent  early  be  The  the  of  dentate  reported a  region  inhibition  to  time  with  the  existence  an  of  the  GABA-mediated  CA3  described  p h a s e w h i c h was  f o l l o w i n g BIC  consistent  others  and  appear  inhibition  this  later  interneurons.  change  mined  into  presence  CAl  would  GABAergic  for  i n the  a  experiments  activation  BIC-sensitive early  occurs  the  and  the i n the  separable  BIC  increased  of  for a  This  which  is  were  especially with  FIM  the  a p p l i c a t i o n of  p o t e n t i a t i o n of in  contrast  apparent stimuli  to  BIC  FIM^ the  there activ-  homosyn-  occasionally (cf. Andersen,  with 1960)  69  in  either  control  consistent reflects  with a  synaptic homoalso  synaptic  but  (Misgeld  reported  to  change  of  inhibition  this  the late  inhibition would the  were  very  that  that  a  with  of  potentiation pathways  firing  a  latency  has  t o CA3  also  that  the major  that  was  increased  related  similar  fact  attributable i n  and t h e l a t e  and  indicated  t o membrane  actions  inhibition  of the  not assessed,  mechanism since  the time  underlies their  both  time  course  inhibition analyses  HP  could  that  courses  occur  that  was  evoked  both  i n the absence  events by  discharge.  sometimes  and FIM  of which  revealed  these  this  (see below).  activated  potential  was  of  F I M s t i m u l i was  currents  action  several  o f BIC on t h e  and l a t e  intracellular  of  the e f f e c t  inhibition  HP,  the  portion  i n the presence  t o other  Although  after  f o r the onset  The  biphasic  accompanying  the late  long  event.  paired  the early  and  a  inhibition  A d d i t i o n a l l y , a c t i v a t i o n o f t h e MF  inhibition  attributable ization  post-  The o c c u r r e n c e  was  be  with  similar.  Extralate  may  observed  elicited  cell  are  potentiation  inhibition  inhibition  heterosynaptic  correlated  of  data  generalized  pulse  of  long-lasting  e t a l . , 1982).  appear  and  the  a  and p e r f o r a n t  and i n d i c a t e d  antagonist  (Heyer  inputs  paired  the pattern  BIC-insensitive  GABA  drug  than  pulse  et_ a _ l . , 1 9 8 0 ) .  o f BIC i m p l i e d  the heterosynaptic  the  paired  rather  t h e MF  These  e t a l . , 1979).  late  slices  it  with  slices.  that  heterosynaptic  administration  of  change  (Creager  not  The  the  BIC-treated  the hypothesis  event  been  or  the  were  not  depolar-  Furthermore, i n the absence  70  of  any  be  activated  case  PS  i n the  also  by  by  a  with  determine the  conditioning  actions  feedforward  the  i f the of  early  late an  orthodromic  were examined.  inhibition  AD^  PS,  As  expected,  be  by  potency  to  occurring  However,  the  These  that total  data  inhibition  through  Since the ment  the  spontaneous of  a  was  MF^  there  hyperpolarizations inhibition antidromic The as  may  e x c i t a t i o n at  indicated  orthodromic  in  of  in  the  Chapter  excitation;  to  early  inhibition  comparable f i m  or  Q  in  was its  activity.  inhibition MF  was  activations.  in  effect  the  late  on  2,  AD  involve-  inhibition  at  larger  with  short  with  (20-40 of  the  the  late  blockade  intervals.  the  MF  r e s p o n s e was expected  relatively  this  intracellular  C-T  that  was  addressing  reduction  effective  unlike i t was  T  suppressed  the  Although  correlated  of  sec,  a marked  more  PS  1  experiments  relatively  1°  pathways  late  PSs.  longer  thus  activity  or  the  no  the the  recurrently,  orthodromic  FIM  up  AD  were  explain  behavior  for  on  The  which  To  weak a c t i v a t i o n o f  50-500 msec  occurred.  above).  possible  AD-evoked  implied  was  the  as  MF^,  afferent  cells  stimuli  to  could  pathway. of  also  i t be  little  was  either  process  i n t e r v a l s , at  response  with  recurrent  of  and  the  relatively  firing  paradigm  m s e c ) C-T test  of  of  as T  (see  evoked  the  after  activation  This  action  stimulus  a  a  postsynaptic  implicated. the  induced only  FIM  stimuli  C  duration  that  imply  the  that  appeared  be  contaminated  activated  than  could  on  AD  as  process  responses, would  suggested  pathway  GABAergic  strongly  shorter  response  of  again, of  an  unaffected  at  71  any  C-T  interval  2°  PS  which  PS  appeared  FIM-evoked  to  activity.  be  Evidence  for  T  the  activity  elicited  a  onset  up  to  700  by  HPs  biphasic  and  was  ties.  of  The  than  the  1°  was  MF the  CA3  in  and  region.  by  the  discussed  experiments  inhibition rat  brain  Bowery,  that  r e s u l t s of  first  with  to  and  may  least a  a  post-  reduction  of  afferent  stimulation  coinciding with The  late  BIC,  at  been  preliminary  previously. for  has  the  phase  that  test  mediated  1981)  MF^-evoked  Thus s u b t h r e s h o l d —  presynaptic  attributable  the  the  a  possible  orthodromic  However,  was  by  of  of  the  Hill  approximated  the  involvement  BIC-insensitive  confirmed  HP  i t i s not  and  the  s e c o n d HP  had  inhibition  as  which  lasted  for  msec.  the  the  response.  inhibition  reduction  implied  IPSP as  (1979) in  stimulation.  of  to  receptors  studies.  Fujita biphasic  test  depressions  measured,  inhibition  was  which  determined  a  i n the  biphasic  GABA-mediated  the  et_ a _ l . , 1 9 8 0 ,  and  intracellular  an  GABA  late  event  not  possible  in  operative  synaptic  the  process  of  same  resistant  E P S P was  on  (Bowery  portion  AD  the  pre-synaptic  also  i n the  more  considerable  PS.  here  inhibitory  reported  occur  be  AD  comment  by  the  could  Because to  despite  In HP  has  rabbit that was  the  The  HP  late  associated  existence  HP  ijn v i v o  whereas with  injection  late was  the  CA3-CA4 n e u r o n s study,  r e v e r s i b l e upon membrane,  reported  of  the an Cl  exhibited  interpreted  to  of  similar  after  fimbrial  early  component  increased or  conductance  hyperpolarization  none be  of  of a  these  proper-  dendritic  IPSP  72  although  the  gated. al. to  In  found  much  to  an  origin  inferred  a  late  but  kroin, ance  CI  be  with  the  the  more  late  (see  and  Ayala,  this et  late  al. ,  potent  1982).  appears  1978)  and  path  observation late  inhibition the  in  vitro  has  with  Evidence has  CA3 for  also  evident cells a been  and  granule as  ( O l i v e r and  to  long  in the  conductAn  an  1981;  region,  (McNaughton  1984)  with  evoked  interest a  as  (Thalmann  CA3  vivo  of  is  the  the  experiments  a by  potentiation  increase  present  and  Alger, cells  HP  of  late after  BIC.  duration  reported  (Schwartz-  responses  (1982) of  the  the  reported  i n the  particular  Ayala  for  1983).  Miller,  of  picrotoxin since  also  of  and  a  et. a _ l . ( 1 9 8 2 )  been  (Nicoll  gyrus,  Of  was  through  basis  Alger,  may  which  potassium  c o r r e l a t e d b o t h jln  t o be  very  characteristics  dentate  dentate  the  a  and  profile  and  the  Thalmann  was  exposure  potential  Fisher  stimulation.  by  HP  Knowles  suggests  long-lasting inhibition  perforant  the  In  dendrites injected  this  pyramidal  1982)  HP  and  the  Initially,  also  neurons  e a r l y IPSP and  laboratory  in  Alger,  CA3  data  occurring  CAl  pig  et  voltage-independence  in  from  investi-  Knowles  as  sensitivity  and  the  electrophysiological  both  not  d e n d r i t i c IPSP  pharmacological  Newberry  HP  communication)  similar  i n guinea  currents  soma.  recent  involved  HP  apparent  from  -dependent  personal  to  This  for  in  late  were  observations,  s e n s i t i v e than  remote  microelectrode  antagonists  Fujita's  similar  reverse.  electronically  HP,  a  GABA  with  less voltage  difficult reflect  of  agreement  ( 1982) be  effects  non-GABAergic  i n other  neurons  inhibitory such  as  the  73  turtle  mitral  cortex  (Satou  guinea  pig  distinct  cell et  To  of  (e.g.  but  which  could  number  of  First,  at  may  appear which  in is  the the  gyrus  late  occurred  suggests  that  a_l. ,  i t  to  of  also  (Fig.  afferent  be  for  1  of  with  a  see  noted  the  and  small  no  a c t i v a t i o n of  or  either  a  well  here  for  of  a  onset to  be HPs  stimulation.  A  inhibition/HP.  initiated  by  neuro-  (Nicoll  and  Alger,  that  the  EPSP  and  Ayala,  Nicoll,  late  (e.g.  non-GABAergic  Thalmann  two  postsynaptic  Newberry  preceding  of  appears  late  afferents but  the  also  latency  large  the  of  1973).  slow  events  the  a  a  for  is  presented  the  to  1981;  should  absence  dentate HP  from  existence  inhibition  account  ascribed  et_  case  This part,  may  be  However,  has  pyriform  cortex  neurons  been  subthreshold  released  Wojtowicz ).  in  The  Gershenfeld,  which  rabbit  olfactory  1980).  has  action.  mechanisms  transmitter (s ) 1980;  by  evidence  least  the  the  invertebrate  review  follow  these  in  1981),  in  al.,  inhibition  long-lasting  attributable,  1982  see  aJ. ,  and  e_t  IPSPs  conclude,  BlC-insensitive  e_t  a l . , 1982)  (Constanti  types  documented  (Mori  HP  Fig.  late  HP  1982).  preceding  could 3.3C) in  the  That  the  depolarization  voltage-dependent  2 + Ca  conductance  influx  (Ransom  probably from  not  or  an  e_t  a_l.,  involved.  inhibitory  co-activated  However,  were  the  this  1975;  pump  Padjen  Second,  the  neurotransmitters  pathways  with  electrogenic  the  with  the  case  differential  major  i t would  inhibitory  initiated and  Smith,  inhibition released excitatory be  a  could  to  is  arise  extrinsic  inputs  of  sodium  1983)  from  difficult  actions  by  to  CA3.  reconcile anti-  and  74  orthodromic  responses  fimbria.  Third,  the  activation  of  anatomical  evidence  al. ,  late MF^ this  in a  and  inhibitory  by  stimuli  inhibition  applied could  interneurons  for  the  feedforward was  elicited  because event.  AD  possible pathway i n the  to  the  reflect  the  which  ( e . g . H a n d e l m a n n e_t a T . , 1 9 8 1 ;  Furthermore,  inhibition response  late  non-GABAergic  1982).  interneurons  evoked  i s suggested  stimulation  of  is  M o r r i s o n et_  involvement  absence  there  of  these  since a PS  only weakly  the  i n the  activated  75  CHAPTER  THE E F F E C T S OF A N T A G O N I S T S  4  ON AMINO A C I D - AND S C H A F F E R  C O L L A T E R A L - C O M M I S S U R A L M E D I A T E D E X C I T A T I O N OF C A l NEURONS  Introduction The in  first  the modulation  Hayashi  (1954)  L-glutamate and  colleagues  excited  the  cortex.  neurons  i n the iontophoretic  studies  shown  property  by of  of L-glutamate confirmed  of Curtis  by t h e s e  that  virtually  a l l types  compounds  (Johnson,  1972).  and S c h w a r t z ,  1967) and n e r v e  that  an  receptors  was  regarding  their  al.  acids  some  and h i s  ( C u r t i s e t a_l. , 1 9 5 9 ; 1960) a n d t h e i n v e s t i g a t i o n s  inferred  opinion  was  amino  provided  convulsant  spinal  1963) a r e n o t d e p o l a r i z e d  mitter  was  An a c t i o n  Phillis,  (Graham  foracidic  excitability  described  to excite  have  (Krnjevic  neuronal  i n the cerebral  later  others  of  who  L-aspartate  years  of  i n d i c a t i o n of a role  et. a l . , function  These  data  Because  fibres  by  ( K r n j e v i c and  together  acid  synaptic  information  nervous  1967) advanced  the p o s s i b i l i t y  f o r L-glutamate  and L-aspartate  was o r i g i n a l l y  Krnjevic  with  with  d i s t r i b u t i o n s i n the central  to the contrary  neuroglia  L-glutamate  i n t e r a c t i o n o f t h e amino  likely.  of neurons a r e  expressed  of a  system trans-  although  an  b y C u r t i s e_t_  (1960). In  t h e two decades  which  have  elapsed  since  these  76  original amino  investigations  acid-mediated  principle  types  neurochemical neuronal  of  concentrations, has  recording elicited vis  a  by  adduced more  general  brief  role  data  recent  of  or  of  (1981)  of and  employed  of  specific  use  such  as  tissue  The  second  postsynaptic  synaptic of  the  amino the  amino  changes analogues  activation evidence  acids  hippocampus.  Watkins  and  and  been For  a  synaptic  Evans  Foster  of from  has  acid-mediated  Fagg  two  has  i t s structural  from  of  electrophysiological  the  review  derived  involved  amino a c i d s .  for acidic  reviews  McLennan  of  existence  One  indices  accompanying  consideration  the  the  have  the  application  L-glutamate  changes  on  release  determine  following  focuses  (1981),  the  with  presynaptic  and  to  transmitter  excitation Puil  uptake  involved  The  confirm  excitation  coupled  assay  exogenous  neurons. a  to  those  to  experimental approaches.  techniques  vis  which  synaptic  analyses  lesions  approach  attempts  (1981),  (1983)  are  recommended.  L - g l u t a m a t e and L - a s p a r t a t e as Neurochemical evidence Although trations  of  laminae  Okada,  1979 ) a is  glutamatergic L-glutamate following  i s no  L-glutamate  somatic  compounds  there  of  or  and  removal  the  drop  following  aspartatergic not  difference  the  the  the  in  the  afferent  contralateral  (Nitsch  levels  destruction fibres. declined  concen-  dendritic  dentate gyrus  L-aspartate levels of  in  L - a s p a r t a t e i n the  significant  evident  but  clear  and  C A l , CA3  neurotransmitters:  of  and and  these  of  presumed  For  example,  significantly  hippocampus  and  the  77  consequent see  also  fascia the Di  In the  Nadler  dentata  Lauro  et  of  amino  1981).  of or  metabolic  activity  and  of  is  of  the  time  now  course  the  these  as  any  of  of  this  of  L-glutamate  In  gliosis  of  acids.  value  reliable  the  also  metabolic  amino  function. as  see  studies  between  discounted  such  of  but  concentrations  studies  to  from  uptake  for  that than  uptake  the  may  role  be  synaptic  system  appears  to  acids  such  the  markers addition,  alteration  of  complicate  the  type  (cf.  occur  Curtis  Watkins  the  the with  was  the  effects  of  the  the  enzymatic  perisynaptic  with  of  A the  high  However, actions  of  catabolism,  is  cells  and/or  affinity  (K^  concentrations whereas  effects  of  (1960)  acids  levo-isomers.  on  cleft.  e_t a l .  amino  termination  excitation,  terminate as  from  and  depending by  by  excitatory  dextro-  involved  normal  of  recovery  synaptic  during  amino  of  lesion  1981;  distinguish pools  of  a  of  1981;  content  after  disadvantage  i n d u c t i o n of  in  applied  rather  attributable  ^AM)  Smith,  objections expressed  apparent  amino"acids,  diffusion  to  lesions  data  iontophoretically it  L-glutamate  s y n t h e t i c enzymes  neurotransmitter  similar  major  (Nitsch,  1981).  One a  and  neurotransmission of  interpretation  the  et. a l . ( 1 9 7 8 )  their  effects  fibres  decreased  hippocampal  secondary  for  The  inability  acidergic  Evans,  was  neurotransmitter  measurement L-aspartate  and  (Nadler  studies, Nadler  or  and  1978);  input  i s an  presumptive their  commissural  synaptosomes  a_l. ,  levels  of  et_ a _ l . ,  perforant path  tissue and  degeneration  a  low  higher  iontophoretic  by <  50  present affinity  levels  of  applications  78  (Watkins  et. aJL.,  dependent  (Storm-Mathisen  in  glia  excitations glutamate there  Lodge  uptake, processes  terminals extent  1977;  caused  no  occur  and  e_t  by  analogues  is  neuronal  (Henn  1980).  such  and  mechanism  1979)  1974 )  compounds  presynaptic to a  smaller of  the  D-glutamate  and  the  as  due  and  sodium-  Terminations  N-methyl  is  These  primarily in  Iversen,  aJL. ,  kainate  uptake  et. a l . ,  to  aspartate  for  which  diffusion  away  from  receptors.  By  examining  following  the  selective  glutamatergic  or  terminals  may  that  decline  any  properties  changes  in  l e s i o n i n g of  aspartatergic  be  presynaptic  the  determined.  uptake  an  of  those  i s assumed  uptake  i s a t t r i b u t a b l e to  boutons  rather  than  glia  or  secondary  postsynaptic  tissue  studies  the  loss  changes  neurons.  the  afferent  i n these  of  of  a  a f f e r e n t pathway,  nature  It  in  in  Selective  of the  uptake  3 of  [ H]  for  the  dentate and  the  labelled  L-glutamate  synaptic  terminals  g y r u s , the mossy commissural  (Storm-Mathisen, al. ,  1978).  hippocampal uptake  in  1977,  efferent the  Because  accumulated possible  in  by  the  uptake  D-  L-aspartate  the  1981;  perforant  pathway  Schaffer  section  1978;  L-aspartate  same o r s i m i l a r to  to  the  the dentate  to  CA3  afferents  to  CAl  the  the  uptake  reported  input  Nadler  fornix,  produced  and  Woxen-Opsahl,  studies  path  been  et_ a l . , 1 9 7 8 ;  of  also  septum  and  from  has  collateral  Sandoval  pathway,  lateral and  of  fibre  Bilateral  (Storm-Mathisen 1980).  and  and  a  a  Walaas and  major  reduction  mammillary and  d i s t i n g u i s h between  of  bodies Fonnum,  L-glutamate  processes,  et  are  i t i s not glutama-  79  tergic  or  aspartatergic  Additional excitation  in  pathways.  support  the  for  amino  hippocampus  has  acid-mediated  been  the  synaptic  demonstration  of  2+ Ca  -dependent  Bradford,  stimulus-evoked release  1978).  distinguishes  A  ( f o r r e v i e w see Cox and 2 + f o r Ca in this process  requirement  i t from  glial  release  (but  see  Roberts,  1974).  2 + Ca  -dependent  slices  of  release  dentate  increased  t  electrical  K  l  +  o  had  previously  reason  endogenous  hippcampal  attributed  to  (Nadler  a l . , 1976;  et  aspartate  failure  CAl  to  studies  fibre  the  role  terminals  from  amino  of  (Nadler  and  by was  perforant that  the  in  the  of  but  For  release  of  isolated  commissurotomized  also  CAl  rats  was  synaptic  boutons  pre-loaded  D-[ H]-  not  3  metabolized)  by  collateral-commissural  been  reported  Malthe-Sorenssen,  -dependent  amino  the  +  Schaffer  has  with  Release  K -evoked  Release  Ca  1976)  suggesting  acid-using  S k r e d e and 2+  of  i n which  L-aspartate  of  demonstrate  et_ a _ l . ,  in  presynaptic terminals.  accumulated  v it r o  depolarization  lesioned,  1978).  e_t a j _ . , 1 9 7 9 ;  controversial mossy  is  i_n  slices  from  slices  stimulation  to  Sorenssen  and  of  been demonstrated  ejt a _ l . , 1 9 7 7 ) .  reduction  loss  (which  electrical fibres  a  for  mainly  L-glutamate  of  (Nadler  been  the  has  following  (White  decreased  originated  same  region  gyrus  stimulation  L-glutamate the  L-glutamate  veratridine  r  0  significantly pathway  of  et. a l . ,  1981).  release  acid-mediated 1978;  (Malthe-  has  A  made  transmission at but  see  uptake  above.) Together  these  biochemical  data  support  a  role  for  80  L-glutamate  in  perforant and  path-mediated  dentate  gyrus  L-glutamate  Schaffer  collateral-commissural input  L - g l u t a m a t e and L - a s p a r t a t e as E l e c t r o p h y s i o l o g i c a l evidence The acids over  number  producing the  past  agonists These  Quisqualis amino a c i d .  Each  with  an  that  occurring charged In  amino of  of  their  cationic  may  Curtis  and  their groups  actions to  ships  have  supported  the  there  i s evidence  observation to  of  L-glutamate,  substantiated interneurons  by and  a  has  the  the  (NMA),  a  been  itself.  isolated  from  the  seeds  of  synthetic  a molecular structure  anionic  groups  proposed by  a  single  of  similar  of  a  by  McLennan  and  Renshaw  class  to  between  than  one  of  postsynaptic relationattachment,  receptor  patterns and  type.  with  thalamic acid  L-glutamate  acidic  attachment  ejt a l . ( 1 9 6 8 )  response to  the  activity  DL-homocysteic  cells  of  three-point  sensitivity  differing  that  three-point  structure  concept  differential NMA  have  L-glutamate  from  amino  considerably  interest  acid  a v a i l a b l e f o r more  suggested a  grown  although the separations  Watkins  studies  first  and  synthetic  differ.  Although  was  than  quisqualic  receptor.  This  has  these analogues  groups  charged  to CAl.  N-methyl-aspartate  of  produce  the  anthelminthic  i n L-glutamate  1960  acids  a c i d , an  and  arrangement  in  particular  potency  simplex;  indicus  L-aspartate  neurotransmitters:  Of  greater  include: kainic Digenea  the  and/or  excitation  decades.  a  in  n a t u r a l l y - o c c u r r i n g and  neuronal  two  with  seaweed  these  of  transmission  the  neurons  and  later  of  spinal  L-aspartate  81  (Duggan,  1974) and k a i n a t e  That the  more  than  depolarizing  the  existence  membrane  and  Thus  decrease  low and h i g h  has  now  doses  (Flatman  (MacDonald  and  resting increase  +  and/or  a 1.,  potassium  o f membrane  current,  conductance  a  as  an  current-clamp  "increase  analysis  to  or decrease  and,  Constanti at  least  resistance (MacDonald have  been  is et  of  (cf.  and L-glutamate  1981;  et al. ,  mechanistically 1982).  attributed  to a  t o produce a  of  to  a an  tetrodotoxincarried  by  ( F l a t m a n e_t this  was with Both  related  effects  (Lambert  et. a l . ,  and W o j t o w i c z ,  neurons  Decreases  pyramids  1983b).  dose  similar  cells  resistance"  resistance  1980; MacDonald  the cat  reducing  Paradoxically  similar  spinal  than  conductance  Dingledine,  have  CAl  current  membrane  membrane  i n cultured  a_l. ,  2 +  1983a,b).  L-aspartate increase  inward Ca  of  effect  spinal  appears  dose  increase  same  cells  by a c t i v a t i n g  voltage-dependent  Dingledine,  NMA  are  an  This  Rather  and  motorneurons  hippocampal  1983a,b).  kainate  feline  in  implies example,  NMA  cultured  and  voltage-dependent  1983 ;  measured  occurring  1980)  by  o f NMA.  a _ l . , 1983),  1982; D i n g l e d i n e ,  insensitive Na  e_t  by  reported  in  respectively as  For  decreased  produced  underlie  also  types.  resistance  Wojtowicz,  outward  receptor  1974).  may  excitants  e_t a l . ( 1 9 7 9 )  reported  neocortex  mechanism  acid  changes  membrane  with  (Hablitz,  the  Engberg  of  been  o f amino  i s consistently  whereas  dependent.  (McCulloch, e t a l . ,  conductance  multiple  resistance  quisqualate  one  actions of  a n d NMA  the  1982);  increase  that  membrane  for  of NMA  resistance  voltage-independent increase  of  82  conductances dependent the  to  amino  cases  of  Na  ,  acid  NMA  K  and  uptake  and  Ca  (Puil,  (Engberg  kainate  et  which  1981)  and  a_l. , 1979)  are  not  ion-  except  actively  in  trans-  ported. Therefore, common  action  to  same  the  the  to  although depolarize  ionic  presence  ionophores  the  of or,  neurons,  conductance several  acidic  This  i s not  indicates  linked  to  activation  of  several  b l o c k a d e o f a m i n o a c i d - m e d i a t e d e x c i t a t i o n may  as  an  i s usually  events  at  inhibition  inferred,  the  or  ionophore.  of to  An  evident  agonist-receptor a  reduction  example  of  of  the  tied  different  differential  to  I t i s also  a  either  by  either  receptor.  have  conductances  due  single  the  acids  effect  types  alternatively, a  this  events.  receptor  amino  that  a be  interactions receptor-linked  latter  situation  2 + is  the  selective  NMA-induced  excitations  Another transmitters that as  an  that  similarity response CAl  ment in  that  observed  connection,  in  criterion  is  of  and  in  the  studies  cells.  this  identification action"  of  activation  Langmoen  which  of  the  (1983)  have  f o r the  (for  and  a  discussion  In  for this a  L-glutamate evoked  complete  practical  effects  reported  collateral-commissural assuming  requires  pathway  transmitter.  potential  Even  of  neuro-  t h e same p o s t s y n a p t i c  theoretical kind  e_t a _ l . , 1 9 8 0 )  1983b).  putative  reversal  rigorous of  the  i s the  Schaffer  hippocampal the  elicit  and  the  (Ault  " i d e n t i t y of  following  Hablitz  Mg  f o r the  of  compound  by  (Dingledine,  exogenous a g o n i s t  are  which  blockade  EPSP  f u l f i l l -  prerequisites  see  Brown  and  83  Johnston,  1983)  these  L-glutamate  as  example  glycine  may  with  act  lying  via  the  the  the  employed.  This  ly  a  and  on  amino  the  of so  McLennan,  receptors  mediating  The  type  is  first  that  receptors  1981)  identified  and  as  a l .,  potent 1982);  Watkins,  for  as  for  compounds  same  under-  be  1981), NMA  D-(-)  receptor  the  1982);  the  of  the  (PDA) and  the  (Davies,  the  of  excitatory  i t s most  potent  (see  antagonized  types  amino and  by  of  acids.  selective I  compounds  for which  (D-(-)-o( -aminosuberate and  Lodge,  6J -phosphonate  certain  (APV)  which  known  agonist  dipeptide  (Watkins,  Appendix  (McLennan  Evans  on  neurotransmitter  three  of  be  antagon-  antagonists  distinguished  (DAA)  NMA  also  subsequent-  cord  antagonists  partial  and  spinal  (NMDA)  isomer  may  to  deduced.  dicarboxylates  1979);  approach  action  these  (+)-2-amino-5-phosphonovalerate  dicarboxylate Liu,  the  firing,  of  in  by  is  D - ( - ) - <?(, - a m i n o a d i p a t e  such et  and  monoamino  cell  responses  structures)  Davies  role  c h a r a c t e r i z a t i o n of  may  have  the  of  nature  studies  chemical  and  distinct  pharmacological  a  the  N-methy1-D-aspartate  the  a  actions  agonist,  include  a  since,  activate  identity  the  Pharmacological  first  to  acid-induced  postsynaptic  1981a,b;  structurally  study,  requires  excitation  the  confirm  transmitter  receptors  of  comparison  synaptic  GABA,  present  demonstration  effects  and  cannot  conductance(s). in  ist  endogenous  different  ionic As  experiments  et  is  compounds  (Davies, one  (Davies  of and  1979;  Francis the  most  Watkins,  (+)-cis-2,3-piperidine al. ,  1981;  McLennan  Y-D-glutamylglycine  and (DGG)  84  (Davies  and  entially  a f f e c t e d by L - g l u t a m a t e  and  Watkins,  (Honore  diethylester interacts pylene  side  second  itself  (GDEE).  chain  agonists  actions  by  as  1981).  receptor  and  by  APV  (Watkins,  responses  synaptically  or  by  interest, the  receptors. determine  Despite synaptic  investigation  with  the  by  DGG  as  greater  of  blockers  and  Watkins,  reduction  1979).  L-glutamate  or  profile  kainate  information type  may  either  L-aspartate  of  and  of  Therefore,  induced  NMA,  their  NMA  excitation  is a  mixed  the  inter-  quisqualate then  be  of e x c i t a t o r y  used amino  responses.  i t spotential  of  charged  f o r example,  to  a  Lodge,  of  informative  and  3  conductances  cord,  (Davies  and  pharmacological  function  isopro-  blockade  DGG  produces  a p p l i c a t i o n of  on s y n a p t i c  of  sensitive  the c o n t r i b u t i o n of each  receptor  an  behave  on membrane  degrees  more  (McLennan  antagonists  This  actions  antagonism  more  kainate  antagonized  L-aspartate  1981b) and  GDEE  of  as the u s u a l  In the s p i n a l are  L-glutamate  which  the presence  acid  PDA.  the varying  Conversely,  of  their  by  with  are  acid  propionic  blocked  receptor  and  excitations  differential  action  by  be  I) as w e l l  1981) and  prefer-  a n d by q u i s q u a l i c  excitations  antagonists.  L-glutamate  acid  (Appendix  i s implied  L-aspartate  of  third  L-glutamate  above)  while  A  can  in requiring  and W a t k i n s , That  such  1981),  Kainate-induced  (Davies  (see  e_t a l . ,  i s unique  groups.  to  The  -amino-5-methyl-3-hydroxy-4-isoxazole-  (AMPA)  is  1981).  use  i n the analysis  dysfunction, pharmacology  there and  has  of been  physiology  cortical l i t t l e of  the  85  receptors higher  mediating  CNS.  In  reported  to  pathways  such  and  the  hippocampus  affect as  amino  the  the  commissural  GDEE,  White  et. al. ,  antagonists (Baudry  and  purpose CAl  in  of  on  by  and  applied  or  the  in  the  have  Schaffer  been  collateral  have  acids the  of  the  achieve  Evans,  and  receptor  collateral-commissural  (SC)  the  effects The  firing  determine  developed  i n the  This  spinal  of the  range  cord  information  was  type(s) mediating  evoked  as  1981).  to  recently  described  antagonists.  analysis  to  1976;  selective  to characterize  of  been  and  (APB)  (Segal,  sufficiently  required  amino  excitations  which  that  (HA-966)  Watkins  acidic  selective  to  Schaffer  doses  1981;  the  these  substances  potent  not  t h e p r e s e n t s t u d y was  neurons  effects  are  high  Lynch,  responses  DL-2-amino-4-phosphonobutyrate  1979)  the  acid  compounds  l-hydroxy-3-aminopyrolid-2-one  1978;  of  excitatory  activation  of  as  then the CAl  cells. The exhibited  phenomenon by  stimulation 1976;  changes not  that  SC  to the  of  (Cull-Candy  the  Wester,  by  Cotman  raises  Alger  whether  function.  LTP  although  (LTP)  high  Although  clear  i s  frequency  and  Teyler,  i t has this  been  reflects  Furthermore, i t the  observation  DL-2-amino-4-phosphonobutyrate et  a_l. , 1981)  invertebrate  et. a _ l . , 1 9 7 6 )  brief  1975;  1978).  triggers  1978;  potentiation  following  postsynaptic  prevented f  term  i t i s not  what  be  e_t a _ l . at  Lynch,  synapse  p r e - and/or  i t could  antagonist  projection  and  understood  (Dunwiddie  long  ( S c h w a r t z k r o i n and  Dunwiddie  localized  is  the  of  the  an  L-glutamate  neuromuscular possibility  that  junction LTP  may  86  be  produced  acid. of  by  a  Therefore  antagonists  postsynaptic some  action  experiments  to affect  of  also  an  excitatory  addressed  the  amino  ability  LTP.  Methods Techniques in  vitro  hippocampal  For CAl  a  4  visual  advanced neuron  0.1-3  integrated  Agonists  stratum currents  of 2  as  of  sensitive  chart  record  applied the  equal  currents  reduce,  agonist.  ejecting  Finally,  of  a  placed  to  the  of  dendritic  DC  level  frequency. from  dendrites  in  layer;  i n the f o l l o w i n g of  rates the  the  Submaximal  the  manner: timed but  of  r u n was  the  as by  Then,  the antagonist  effects  recovery  The  obtained  excitants.  a  backing  automatically  were  a  potentials,  firing  body  and  f o r a l l compounds.  continued,  50-100%,  was  adjustable  recorded.  firing  single  iontophoretically  comprised  excitants by  an  the c e l l  examined  were  of  amplitude  Action  apical  +20-30nA  excitants  a p p l i c a t i o n of to  agonist. with  2.  (R-^, F i g . 2 . 3 )  i n response  onto  were  activity  potential  gated  were  cycles  possible  adjustment  applied  a  generally  control of  or  100-200j*.m f r o m  antagonists  applications  serial  were  the  the  i n Chapter  microelectrode  action  microelectrode  were  least  nearly  kHz  maintenance of  pyramidale  excitatory  to obtain  radiatum,  actions  an  and a n t a g o n i s t s  7-barreled  at  of  of  glass  i n stratum the  and  been d e s c r i b e d  recording  spontaneously  iontophoresis  and  have  NaCl-filled  maximize  firing  at  M  control  to  filtered  slice  extracellular  cells  under  f o r the preparation  as was  most  performed  87  (e.g.  F i g . 4.1). The  pH  8.0),  8.0  solutions  L-aspartate,  in  NaCl),  following  160  mM  (500  NaCl),  mM,  pH  acetylcholine  (NMA)  NaCl),  in  160  amylglycine N HC1 mM,  mM  8.0  (D-APB)  ]  in  6  N  pH  7.0  mM  pH  NaCl),  7.5-8.0  i n 160  mM  (GDEE)  ( 2 0 0 mM, The  NaCl,  pH  [oC]  ( 5 0 0 mM,  antagonists  in  on  the  SC-evoked  mM  i n 160  mM  pH  8.0  4.0),  i n 160  ([<tf]  in 6  D  (L-APV)  mM  (20  N mM,  X-D-glut-  NaCl,  dicarboxylate  mM  [<*]  in 6  Q  (PDA)  (100  mM  NaCl,  D  in 6 N pH  HC1  (L-APB) +20.8°),  3.5-4.0),  in 6  N  HC1  (20  mM,  pH  L-glutamate  D-(-)-<-aminoadipate  8.0).  synaptic electrode  cell  body  submaximal by  a  barrels  t h e same a s  the  positioned layer,  reversed  2.  effects  the  Briefly,  of  central  agonists barrel  of  CAl in and an  i n s t r a t u m r a d i a t u m 100-300  (1*2'  Usually PS.  and m e a s u r i n g  i n Chapter  excitation  EPSP.  contaminated were  i n 160  +24.2°)/  i n 100  160  experiments analyzing  iontophoretic j>\m f r o m  pH  techniques for evoking, recording  the  pH  D-(-)-2amino-4-phosphonobutyrate  r e s p o n s e s have been d e s c r i b e d of  mM,  8.0  (D-APV)  HC1  mM,  mM,  ( 2 0 mM,  (+)-cis-2,3-piperidine  mM,  diethylester  most  pH  (500  (5-10  8.0  (500  L-(+)-2-amino-4-phosphonobutyrate  7.5-8.0  field  [  ( 5 0 - 1 0 0 mM,  i n 100  (20  -23.5°),  NaCl,  (DGG)  -25.2°),  pH  (DAA)  mM,  (ACh) (APV)  pH  L-(+)-2-amino-5-phosphonovalerate  0  8  ( 5 mM,  D-(-)-2-amino-5-phosphonovalerate  HC1-24.2 ), pH  kainate  (20-50  chloride  (+)-2-amino-5-phosphonovalerate  L-glutamate  8.0),  quisqualate  N-methyl-DL-aspartate  NaCl),  were used:  F  ^9* this  Solutions  those used  2.3)  recorded  potential filling  in single  unit  was the  the not outer  experiments  88  with  the f o l l o w i n g changes: quisqualate  pH 8 ) , k a i n a t e mM  i n 1 0 0 mM  of  10-120  until  a  NaCl,  constant  measured  was  before  (2-tailed  unpaired  mimicked are  repeated  In  studies  frequency  applied a  during  recovery  The  of  o f t h e EPSP  the a d m i n i s t r a t i o n of the (p  from  <  0.05)  the drug  slice  controls.  from  cases  by  few e x p e r i m e n t s  the  application reproducible  and were  Results  never  presented  on LTP o f P S s o r EPSPs  s t i m u l a t i o n ( 1 0 0 Hz f o r 1 s e c ) a n t a g o n i s t s  i n most  an  a period of 1 minute  t h e same  effects  (100  f o r periods  action  i f the size over  NaCl,  administered  E f f e c t s were h i g h l y  current  of t h e i r  applied usually  different  using  appropriate  +S.E.M.  only  2 for details)  t-tests).  were  i n 100 mM  pH 8 ) a n d NMA  achieved.  real  and a f t e r  means  high  in  were  by  was  significantly  amplitude  tests  antagonists  measurements)  was  NaCl,  E x c i t a n t s were  effect  (see Chapter  antagonist  i n 1 0 0 mM  8).  considered  consecutive  when  pH  sec whereas  antagonist  (6  ( u p t o 50 mM  ( 5 0 mM  iontophoresis  by  were  t o the d e n d r i t e s , but  the D-(-)-isomer  o f APV  was  perfused  -5 (10-20  ml  adjusted  of  5x10  M) .  The  intensity  o f SC  activation  t o p r o d u c e a s u b m a x i m a l PS o f 0 . 5 - 4 . 0 mV  negative  deflection  positivity  from  the  baseline  or  was  and t h e peak  preceding  peak  was m e a s u r e d . Results  Effects  o f e x c i t a n t s on s i n g l e The  recording  unidentified i n this  study  unit  activity  C A l n e u r o n s e x a m i n e d by were  sensitive  extracellular  to dendritic  applica-  89  tion  of  the e x c i t a t o r y amino  L-aspartate agonists  and L - g l u t a m a t e .  i t was  iontophoretic of  action  possible  drastically  reduced  the  effects  the  ejected  compounds  fulfilled  a  large  potentials could  the  gate;  iably;  and  3)  that that  number  elicited  by  where  the rate  equally On  potency applied few  of  rates  were  action onset  were  was  no  or  equal  equivalent  sec).  on  that  examined,  of  were:  only  1)  that  by a d j u s t m e n t  tests  achieved or  of  apprec-  recoveries  to  for excitation  quisqualate,  t o onset level  a l l of  of f i r i n g  and  which  declined  of the e j e c t i n g  current.  difference  i n the  as  ejecting  responses  Only  or  away.  i n the stratum  the other  the  implied  d i d not change  L-aspartate  currents  of  to diffusion  were  a stable  obvious  firing  attained.  the termination  of kainate (2-10  height  (< 2 s e c ) l a t e n c y  to the a p i c a l dendrites  nearly  cells  L-glutamate  L-glutamate  exceptions  of  easily  rapidly attained  there  of  produced The  a short  rapidly with  average  due  i n pharmacological  L-aspartate,  showed  not  the  abolished  This  suitably isolated  the spike  criteria  which  dendrite  response.  where  movement  f o r s e l e c t i o n which  be  80-110% o f c o n t r o l f i r i n g These  nA e l i c i t e d  the  and  three  dendritic site  vertical  from  local  the c r i t e r i a  2)  a  quisqualate,  the l a t t e r  t o t h e s o m a 100-200y*.m  action DC  where  away  were  kainate,  o f 0-50  the neuronal  obtained  Although 129  to locate  ejection currents  t i p 2 5 - 5 0 j^m  NMA,  Especially with  p o t e n t i a l s , and  electrode  acids  the  excitants  when  radiatum.  With  two  (see a l s o  hand  had  occasionally  apparent  a  could  amino Segal,  longer a  acids 1981). latency  submaximal  90  plateau  of  firing  continuous kainate  slow  rise  situation  The  usual  Yamamoto  and  was  Sawada,  i n bursts  which  high  frequency  (>35  spikes  rapidly  recruit  because  and of  pattern  The  aspartate,  often  there  firing  from  was  rate  a  during  Cells applied  of onset  initially by  gated.  current  bursts  was  firing  tendency cells  stopped  to the rapidly  at a the  NMA  be  to  well-  particularly  i t was with  to  of  of  to  addition,  inactivation,  also  the c e l l  the amplitude  target  of  (2-10 s e c ) ( c f .  t o cause  The  In  case  which  continuous  which  i n height.  agonists potent since  the problem could  currents  than  substance,  latency  required  i n the  often  NMA.  diffiIn  as t h e  most  response  inactivating  high  train.  order  these  difficult  this  during  ejecting  however,  rank  to  spike  more  and w i t h  ejecting  ACh  more  e q u i l i b r i u m response  specific  clearly  active  an  spike  more  succeeded  Hz)  the r a p i d  shifted  precise  were  neurons  t h e NMA  frequency  barrels  and  neuronal  1 9 8 2 ) was  carefully  to obtain  instances  even  declined  adjacent  isolated  mined  the  had a long  fire  were  in  response  characteristically  cult  attained,  iontophoresis. The  NMA.  be  used  three  NMA, than  kainate  and q u i s q u a l a t e a l l  either  L-glutamate  transport  numbers  of achieving  were  stable  n o t be a s s i g n e d .  On  agonists  appeared  t o be  not  firing  i n the 20-100  L-  deterrates  the basis  and the c o n c e n t r a t i o n s  or  a  of the  electrode times  more  t h e n a t u r a l l y - o c c u r r i n g compounds. could with  also  high  be  excited with  currents  ACh.  ( 8 0 - 1 0 0 nA)  The  had a  response latency  to 5-10  91  times  longer  achieving firing  the  was  that  same  persisted  current al.,  than  plateau  for  effects  on  NMA,  kainate,  are  summarized  the  of  in  preferential Higher  PDA  three  elicited agonists  was,  20  In  after  L-aspartate  addition,  the  ACh  Straughan,  enhanced  excitatory APV  partial PDA  current  NMA or  the  not  of  cell  ejecting  1966;  Dodd  APV  to PDA  CI  was  CAl  et  could  neurons  et.  of  with  an  the  of  NMA of  and  ACh  ejecting  time,  PDA (Fig.  induced  by a l l  This  antagonism  accompanying  excitatory  (Fig.  blocked  manifested  4.1A). to  a  be  That  large  mimicked  The by  by the  extent  its activity  E v a n s et_ a _ l . , 1 9 8 1 ) . not  by  administration  responses  is consistent with  could  induced  response  l o w e r d o s e s was  be  and  low  periods  a l . , 1981). by  L-APB  L-glutamate  periods  quisqualate  (Davies,  other  of  longer  Evans  D-APB,  applied  short  depression  shown)  selectively two  and/or  antagonists  c a r r i e d by the  for  which even at  agonist other  Of APV  effect  When  complicated  response  (data  I.  (Davies,  DGG,  L-aspartate,  antagonism  potent  ( F i g . 4.IB)  DAA,  hippocampal  nA)  doses  unfortunately,  effect  of  to  Table  exhibited  and  of  excitations  APV,  quisqualate,  (up  4.1A).  PDA,  e x c i t a t i o n of  currents  of  and  amino a c i d - i n d u c e d  GDEE  with  rate.  seconds  (Biscoe  doses  1981).  The  an  with  firing  20-80  terminated  A n t a g o n i s m of  of  observed  as  a  effects a  100  nA  ( F i g . 4.1A). amino  acid  antagonized the  more  antagonists  tested,  NMA-induced  excitations;  potent  as  has  been  only  reported  DAA and by  92  Antagonist  Agonist (mean  1 S.E.  of  NMA  Kainate  512.2  (n-13)  +38.318.8  (n-10)  -89.  512.6  (n-23)  -49.515.5  (n-=28)  D-APB  -68.  013.6  (n-20)  -43.018.0  (n«r5)  DAA  -58.  917.2  (n=16)  +38.5H7.6  (n = 7 ) *  +59.418.7  PDA* *  -52.  718.1  (n=7)  -  9.9H1.7  (n=7)*  +14.7114.4  GDEE  -11.  3U4.7  (n=8)*  +  9.512.9  L-APB  + 53. 9 H 5 . 3  (n-=9)  +14.319.5  APV  -53.  DGG  -61. 8111.2  D-APB  -29.  L-APB  + 17. 4 H 3 . 4  NT  -  excitatory  not  with  -12.512.6  (n=7) Kn=8)  +  (n=5)*  period.  I.  different  Actions  changes  6.214.2  (n=30)*  -  7.515.5  (n-19)*  3.H7.3  (p  <0.05)  currents  (n=8)*  (n-9)  +  1.4H.8  (n=5)*  +  2.914.0  (n-3)*  +  2.813.7  (n=4)»  (n=5)*  < 20  (n=7)» (n-15)  ACh  (n-9)*  from  (n=16)  NT  zero, nA ( s e e  text).  o f amino  t o NMA,  acid  kainate,  percent  quisqualate,  (mean  +S.E.M.)  b y some of  test  depression drugs  unidentified  ont h e L-gluta-  i s indicated  i n parentheses.  compared  to that  one-tailed  paired  rates  were  neurons  of  poten-  during  of by  of  cells  firing  rate  the antagonist  was u s e d  significant.  or  evoked  T h e number  The average  occurring t-test  (-)  i n the frequency  CAl  applications of excitants.  infiring  antagonists  and ACh.  discharges  A  -  +66.9*31.8  (n-10)  + 30.4110.4  iontophoretic  (+) c a u s e d  was  (n-11)  +14.015.3  (n-4)*  -14.2H0.7  the end of a 2 - 4 min a p p l i c a t i o n  tested  .  +28.019.0  tested.  L-aspartate  ionophoretic  (n=9)  -  L-Glutamate  (n-10)  817.0  responses  The  at  3i5.3  significantly  Table  tested  Quisqualate  -93.  ••applied  spike  rate)  DGG  •not  tiation  firing  APV  L- • A s p a r t a t e  mate,  % change  being  the control  t o determine i f  93  A.  CI J O O n A , 2 m m  A  40-  I  A  PDA. 20nA,2min i  f  1  A  0  J  SJ  m  S  52  Kam.35nA  m  m  11  11  \ 1 S3  S3  C M N M A . 12 nA  Quis.lZr.A  2 0 sec  PDA. 100nA. 2min  40n  0  J  KSJ KSJ N M A , 1 5 nA  Fig. on  4.1  two  minute  from  a  carried  to  by  bar  time  lower  In  CAl  each  .  of  PDA  the  identifies  the  elapsed  since  and  ejection  bars the  (20  showing  PDA  indicate legend  (100  by  following  of  below  identifies  a  obtained  was  itself  nA  current  100  the  upper  current  ejection.  duration  and  antagonized  records  antagonist the  B,  and  i t sejecting  PDA  following  nA)  a  of  kainate  reduced  agonists  antagonist,  the s t a r t  the e f f e c t s  In r e c o r d  not a f f e c t e d the  20~sec  t o NMA,  were  nA).  three  and  currents.  I Ouis.l2.5nA  responses  neuron,  this  horizontal  their  neuronal  In  administration and  records  E x c i t a t i o n s were CI  |  ( A ) r e s p o n s e s t o NMA  different  excitatory.  solid  CAl  a p p l i c a t i o n of  responses  the  Ratemeter  hippocampal  quisqualate.  ^Kain.30nA  and The  of  agonist  the  agonists  94  others  (Davies, APV  Francis  effect  of  4.2.  Potentiation  r e s p o n s e s was  on  the  et  NMA  of  aJL. ,  response  the  an e f f e c t  1981). is  kainate-  The  preferential  illustrated  and  in Figure  quisqualate-induced  n o t e d w i t h b o t h APV  ( F i g . 4.2)  and  DAA  mixture  and  (Table I ) . Experiments the  resolved  same  cells  implying  (98%)  Thus  the  effect  21.8+4.9%  current  DL-APV p r o d u c e d induced  responses  response  could  currents  of  The  to  qualate  and  of  equal  to  DL-APV  the  of  least  with  be  and  also  of  affected  ( F i g . 4.2).  currents  nA)  rather  Of  the  hippocampal by  DGG  than  1982).  the  used  response  to  with apply  or q u i s q u a l a t e of  attributed  to  the  NMA  ejection the  small  isomer.  d e p r e s s i o n of kainate  three CAl  15-50  NMA  ejected  nA)  although  the  its  McLennan,  (40-100  When a p p l i e d  and  D-(-)-  those  marked  the  the  L-APV  antagonized  extent.  with  the a c t i v e  a  APV  antagonism  high  may  produced  have some a c t i o n (75-100  of  on NMA-  That  on  effective,  quisqualate  while  half  DGG  response  was  decreased  the  effect  ( F i g . 4.3A)  lesser  compared  mixture  increased  observed  response  were  a _ l . , 1981;  o f t h e L-APV sample by  a  was  e_t  significant  dipeptide  NMA-induced  examined,  be  t h e DL  quisqualate reside  ( F i g . 4.3A).  L-APV  contamination  albeit  no  APV DL  (mean + S.E.M. , n = 4)  intensities  of  antagonist properties  ejection  94.3+3.9%  of  the  Francis  iontophoretic by  only  upon  ( c f . Davies,  response by  that  both  potentiating  the e f f e c t s  L-(+)-isomer  showed  that  isomer  i n which  the  responses,  specific  agonists  neurons  to  a  with nA  high large used  quis-  dose  did  ejecting to  obtain  95  SDGG.20nA.2mm  ssso N M A  Fig. APV  and  antagonized of  Blockade  DGG.  quisqualate  order  4.2  -  8 5nA  APV and NMA,  2-APV.  vzzn Komate 3 0 n A  of  reduced  amino  kainate  sensitivity.  and  Quisqualate !2 5 n A  acid-induced  responses  kainate-induced  20nA.2min  to  NMA  excitations and  excitations  quisqualate  by  potentiated while  responses  in  DGG that  96  A  D.L-APV, 2.5nA.lmin  L - A P V . 1.3 nA. 2 mm,  L-APV. 50nA,2mm  20-  m  a  a  fi  a  ma  I Ouis. 7.5 nA  ma  a  E3 N M A . 3 n A  2 0 sec  D.L-APV.I5nA. 2 mm.  B  40-,  n  ,ja  ftp  0  I.L-glutamate. 2 0 n A  Fig.  4.3  hippocampal  CAl  racemic to  NMA  mixture and  reproduced DL-APV  Effects neurons of  APV  only  with  a to  L-asparitate.-22.5nA  S3 N M A . l5r,A  of  DL-APV  and  to excitatory decreased  quisqualate,  selectively  L-aspartate.  •  'a  L-APV  amino  and  blocked  higher  doses  excitations  responses of  acids.  increased  respectively.  much  on  2 0 sec  These of  In  the  (A)  responses  actions  L-APV.  induced  the  by  In NMA  were (B) and  97  the  data  of Table  I , DGG  by 8 0 . 7 + 4 . 9 %  responses The  two  90% p u r i t y  4 . 4 A ) .  D-APB  while In  contrast,  each  of  itself  antagonized  three  reported  actions did  were  excluded of  responses paralleled In antagonize induced  and  indicates A  dose  from  I.  Table  pre-treatment to  a l l 3  the s p i n a l t h e NMA  (Hicks Davies, that  et_ a _ l . ,  inhibition  rate), an  as  was  inconsistent  agonists.  GDEE  7  other  cells  GDEE  tended  which  was  which  recovery to  block  sometimes  height.  tend  centres  also  to  L-glutamate  1 9 7 8 ; Lodge  the  was  1982).  incomplete  effect  and h i g h e r  trend  had  was  L-APB  et. a _ l . ,  GDEE  and  occurs  NMA  compounds  block  which  L-aspartate-  responses  relatively  et a l . , 1 9 7 8 ; CollingLodge,  also  ( F i g . 4 . 3 B ) o r DGG of  shown),  of  1 9 7 9 ; McLennan  t h e same  not  I because  leaving  occasion of  with  receptor  the action of  on  the s p e c i f i c  i n spike  cord  unaffected.  effect  However,  agonists,  kainate  any o f t h e e x c i t a t i o n s o f the  firing  by a d e c r e a s e  o f e i t h e r APV  complete  antagonize  i n Table  excitations,  unaffected ridge  ejecting by  and  potentiated  Francis  approx-  actions( F i g .  t o NMA  (data  (Evans,  on e x c i t a t i o n s i n d u c e d  reported  (<80%  D-APB  1 0 0 nA  to  not s i g n i f i c a n t l y  cells  by  to  relatively  excitatory  i n the s p i n a l cord up  different  remained  quisqualate  resolved  ( F i g .4 . 4 A ) and  This  blockade  Currents  were  o f APB  agonists  and  (n=6) respectively.  the response  response  excitatory. to  which  strikingly  the L-(+)-isomer  the  sensitive  o f APB  had  the quisqualate  the kainate  ( n = 6 ) and 2 8 . 3 + 8 . 6 %  isomers  imately  decreased  1979).  Table  I  i n the hippocampus.  which  response  produced also  almost  markedly  98  D - A P B . 5 0 n A . 2 mm  40-,  /M  I  O  0  0  L - A P B . 5 0 n A . 2 mm  40-i  0  S  J  2 0 sec  0uis'6nA  E3  Kainate O nA  D - A P B . 5 0 n A . 2 mm 40-|  •  a  •  a  .a  L - A P B . 5 0 n A . 2 mm.  40-,  0  J  o 20sec  Fig. responses amino  of  by  firing  CAl  NMA,  a  of  neurons  •  the to  was  With  optical  and  both  observed  •  a L-osportate 2 0 n A  an  75nA  isomers  kainate  cells  a NMA  S3  iontophoresis  L-aspartate  D-APB. rate  •  o  L-glutamate 2 0 n A  Effects  two  acids.  antagonized control  4.4  •  of  of  APB  excitatory  responses  increase  i n the presence  of  on  were  from L-APB.  the  99  A  DGG. 3 n A . 2 min 40i  L• L a  L •  a  S3  N M A  -  a  a  a  2nA  a  an  • a  L-aspartate. 3nA  20 sec  DGG. 15 nA  B  DGG. 5 0 nA  L-aspartate. 3 nA  Fig.  4.5  responses  of  Record  illustrates  A  aspartate reduced effective  CAl  The  neurons  response or  by  abolished  against  differential to  NMA,  blockade DGG. the  In  antagonism  by  DGG  of  the  L - a s p a r t a t e and  L-glutamate.  of  not  (B)  the  NMA  higher  L-aspartate  L-glutamate-induced  but  doses  of  response  excitations.  the  DGG were  L-  which less  100  depressed  the L-aspartate  change  of the L-glutamate  doses  of  APV  or  DGG  effectively  to  appreciably  changing  Liu,  reaction.  (Fig.  antagonize  DGG  L-glutamatemore  receptors  4.5A)  i t was  t h e NMA-induced  the L-aspartate  was  applied  sensitive with  or  with  often  no  lower  possible  response  reaction  effect  and  at a  dose  without  (McLennan  DGG  ( F i g . 4.5B).  and  seen  also  with  ized  I , F i g .4.4B).  significant  effect  L-glutamate  (Table firing  application currents  Effects  The  both  the l a t t e r  observation and  to that thus  responses  i s  quisqualate  induced  observed t h e NMA were  L-( + ) - i s o m e r  on e x c i t a t i o n s  o f APB  with and  antagonh a d no  by L - a s p a r t a t e  and  I , F i g . 4 . 4B ) . of CAl c e l l s  o f ACh was  ejecting  o f NMA  D-APB;  but not the L-glutamate,  The  This  similar  L-aspartate, (Table  reduced  b y DGG.  qualitatively  was  which  excitations,  the s e n s i t i v i t i e s  t o antagonism  An  produced  by t h e i o n t o p h o r e t i c  not s i g n i f i c a n t l y  DGG, A P V a n d D-APB  a f f e c t e d by  100 nA  (Table I ) .  o f e x c i t a n t s on EPSPs The  recording recorded the  little  Interestingly,  and L - a s p a r t a t e - i n d u c e d  consistent  APV  but caused  1982). When  were  response  least  currents  excitants, site  applied  i n stratum  EPSPs  (Table  potent needed  radiatum,  II).  as judged and  the  iontophoretically depressed  L-Glutamate  of  the  extracellularly  and L - a s p a r t a t e  by t h e i n t e n s i t i e s magnitude  at  were  of the ejecting  the  effects,  and  101  % Excitant  EPSPs  ( m e a n +S.E.M.  depressed/  slices  EPSPs t e s t e d  NMA Kainate Quisqualate L-aspartate  24/24 22/22 15/16 5/6  L-glutamate  2/4  Table sural-evoked excitatory The not  I I .  Changes  EPSP  10.0+5.8 of t h e Schaffer  o f t h e EPSP different  the depression  Quisqualate was  tested)  collateral-commis-  iontophoretic  applications  induced  and k a i n a t e  of  acids.  significantly  there  a l l  77.4+3.6* 65.8+4.8** 4 5.4+5.8 31.7+8.7  following  % depressions  than **  reduction  no  was  induced more  one a n o t h e r  by t h e o t h e r  potent  significant  L-aspartate-induced  from  b y NMA  than  b u t were  excitants  L-glutamate  greater  (p<0.01).  (p<0.01) b u t  d i f f e r e n c e between L-glutamate-  effects.  were  and  102  reductions  in  EPSPs  terminating  the  potent  but  which  usually fully s t i l l  long-lasting slices  depression  This  was  of  kainate  and  the  for  periods  up  potentiation  of  complete  of  was  slices  i n 1-3  t h e EPSP  to  24  min  t h e EPSP  Depression detectable  change  of  with  higher  size  the  fibre  of  kainate  and  always  small,  volley  on  preceded These be  was  were  13  with  recovery  more  o f t h e EPSP  in  was  not observed,  took  1.5-2 was  In  18  f o l l o w e d by  an  ( F i g . 4.6A,B).  6  whereas  any  full  were  often  apparent  antagonized  sometimes  completely  t h a t o f t h e EPSP  the  from  without fibre  Recovery  of  (Figs.  4.6B,  the  volley, i n the With  volley the  fibre  was  fibre volley  4.7A).  the f i e l d  selectivity  any  observed.  abolished  o f t h e e x c i t a n t s on and  where  recovery  r e d u c t i o n of the f i b r e  NMA  levels  min.  of the presynaptic  occasions.  effects  but i n  slices  doses of the e x c i t a n t s reductions volley  was  causing  ( F i g . 4.6A)  In  9  (4-20 m i n ) .  rapid.  a n d was  in 1 slice  More  usually  observed  to kainate  ( F i g .4.6B).  t h e EPSP  quisqualate  several  ( F i g 4.6A).  observation  min  was  depressions  d i d not return to control  i n the size  however,  min  much  after  Quisqualate  other  potency  9 min  NMA-induced d e p r e s s i o n  could  period  recovery  potentiation lasted  always  i n the  similar  EPSP  was  but  min  similar  recovery  (6-165%) i n the amplitude  remaining  1-2  i t s effects  Full  min  of  the  the  the  w i t h i n 1-3  3-25  recovery  increase  qualitatively  ( F i g . 4.6A).  NMA  reversed  recovered  throughout  Although  slices  elicited  was  within  incomplete  fully  iontophoretic application.  more  potent  were  potentials  resembled  that  103  Fig. EPSP.  A.  (Quis)  and  The  peak  4.6  record  the effects  kainate  (Kain)  were  amplitude  site  were  records  letters  Note  that  followed  (a-e).  recovery  by  greater  at  taken  o f a PS  presynaptic EPSP d u r i n g  Note fibre  was  volley,  and  and  the  by t h e b a r s of  5  superimposed  reduced  t h e a d m i n i s t r a t i o n o f NMA ( g ) .  t h e EPSP.  with  produced  recovery  size  the v i r t u a l  by t h e  recovery  associated  following  the  above t h e  depression  NMA  to  values at  indicated  the slow  slice  slice.  i n response  iontophoretically  In this the  Hz,  NMA-induced  depression  also  quisqualate  e x c i t a n t s depressed  the  field  t h e same  at the times  A l lthree  ( d ) . B.  on  comprised  p o t e n t i a t i o n o f t h e EPSP  depression.  0.1  indicated  responses  from  o f NMA,  measured  p o t e n t i a t i o n (b) and t h a t  kainate-induced  generation  was  applied  were  the SC-evoked  compared  EPSP  f o r the periods  Representative  small  each  delivered  Drugs  oscilloscope  the  of  stimuli  plotted. recording  o f e x c i t a n t s on  In this  successive  graph.  Effect  but  was from the  a much  from the broadened  abolition  of the  104  (b)  (a)  NMA 5 0 nA  Kain  (c)  (d)  Quis ICOnA  (e)  _j2mV 10 msec  105  observed  when  excitation  the  antagonists  of s i n g l e u n i t s .  were  T h u s APV c o m p l e t e l y  NMA-induced  reduction  the  p o t e n t i a t i o n o f t h e EPSP  ensuing  (Fig.  4.7B).  either slices;  either  (2/3  slices).  reduced slightly  by  APV,  unaffected DGG  or  slices,  (3/6 s l i c e s )  F i g . 4.7B)  were  i n t h e EPSP and f i b r e  I n t h e same  unaffected  also  quisqualate  kainate  abolished kainate  prevented  the  as w e l l as  or  NMA  were  i n duration  (3/6  slightly  responses  similarly potentiated  (3/3 s l i c e s ) ,  (2/3  responses  examined  responses  responses  responses  quisqualate  the  volley,  or increased  and  against  i n a l l 11 s l i c e s  (1/3 s l i c e s )  abolished  reduced  examined  slices)  (3/4  slices;  and Fig.  4.8B).  Effects  of antagonists The  tered on  evoked  abolish  EPSPs  expelled  o f APB  By  contrast  pathway results  when of  acid  antagonists,  i n stratum  ( F i g . 4. 7 B ) . a l l similarly  applied  analyses  been  expected  t o reduce  well  as those  induced  with  with  DAA,  b y NMA  little  doses  o r no  as  synaptic  unit  on  APV, to  effect optical  antagonists  antagonist  of  the basis  of  firing,  and q u i s q u a l a t e  ( F i g . 4.8B).  actions  needed  GDEE a n d t h e  which,  adminis-  I I I .  those  ineffective  single  kainate  of  had  DGG w a s a n e f f e c t i v e  had  i n Table  i n excess  depressions,  also  radiatum,  are summarized  i n doses  were  pathway.  the  site  the NMA-induced  t h e SC  this  amino  which  t h e EPSP a m p l i t u d e  isomers of  excitatory  at the recording  although  on  on EPSPs  would  have  responses  I n 14 t e s t s u s i n g  as 9  106  EPSPs d e p r e s s e d / EPSPs t e s t e d  Antagonist  % reduction 21.3+2 8.2+2  DGG 21/23 L-APB 4/9 D-APB  3/6  6.4 + 3,  DAA  2/4  4.2+2,  APV  10/26  3.3+1,  GDEE  0.1+1.9  0/4 Table  I I I .  Effects  collateral-commissural-evoked Column  2 gives  significantly post-drug  t h e number  (p<0.05)  of  antagonists  on  Schaffer  EPSPs. of s l i c e s  depressed  administration,  and  i n which  compared the total  t h e EPSP  t o both number  was  p r e - and of  slices  tested. The  third  calculated DGG  was  compounds. potencies  column  presents  f o ra l l s l i c e s significantly There were of the other  the  mean  depression  tested. (p<0.05)  more  no s i g n i f i c a n t 5  +S.E.M%  substances.  potent  than  differences  the  other  between t h e  107  Fig.  4.7  the  somatic  are  illustrated.  superimposed phoresis of  fibre and  lerate  on  response pletely gonist  are  PS.  and  B.  the  NMA  was  abolished  composed dendrites  activity  first  SC-evoked by  evoked  by  of  EPSP  and  the  on  depression  as  three  consecutive  d i d not change  there  the amplitude  ionto-  was  a  reduction  The  presynaptic by  the  the depressions (Quis).  The  EPSP  by  NMA  depression. of the  EPSP.  of  this  graph i s  f o r F i g . 4.6.  induced  quisqualate-induced  and  DL-2-amino-5-phosphonova-  quisqualate  constructed  2  Following  (t=l min). of  (R )  stimulation  (t=2 m i n ) ; f o l l o w e d  Effect and  EPSP SC  n e g a t i v i t y i s upwards.  recovered the  volley-dendritic  i n CAl  to the a p i c a l  induced  potentiated  (R^  postsynaptic  the  continuous  fibre  Records  sweeps;  volley  finally  The  potentials  o f NMA  p r e - and  A.  APV  and  com-  slightly  The  anta-  108  NMA.lOOnA.lmtn.fO  t'-2mm  t»tmin  l'2mm  i«5mm  2 - Ammo - 5 - Phosphonovalerole 100 n A  I«15 mm  109  slices,  where  13.0-30.7%,  this  the amplitude  simultaneously significantly depression of  dipeptide  administration  other  slices,  above)  weakly these  substances  at reducing  and  were  tested  was  was n o t  t h e DGG-induced  within  min a f t e r  the start  3-6  min and  terminating the  which  and  response.  21.4 + 5.1%  F i g . 4.8B).  L-«(.-aminoadipate  t o be  These were  depression)  However,  (Hall  complicates  proved  DL-  and  both  this  PDA PDA  e_t a _ l . , 1 9 7 9 ) a r e  the  interpretation  of  results.  of antagonists  on PSs  and  synaptic The  stratum  effects  radiatum  pyramidale.  were  Consistent  reduced  decline  monitored  examined with  of  could,  normally  occurred  on  the amplitude  t h e PS  of  iontophoretically recorded  of effect t h e PS  t h e EPSP  Both  however, after  on  the DGG  reduce  brief  3  action  (4/4 s l i c e s ) or  on t h e EPSP, with  In contrast,  occasions  abolish  to  i n stratum  evoked  ( F i g .4.9).  o r a b o l i s h e d PSs and t h i s  concurrently.  slices)  applied  i t s lack  (0.05-0.1 Hz) s t i m u l i  reversibly  plasticity  o f A P V a n d DGG  d i d not affect  frequency  the  volley  10-30 s e c a f t e r  maximum  the synaptic  46.0+6.5%,  excitatory,  Effects  APV  Generally  2-4  (7/8 s l i c e s ,  -  (see  a  levels  fibre  of antagonist.  o(. aniinoadipate (12/12  reached  o f t h e EPSP o f  a l lbut 1 o c c a s i o n  was d e t e c t e d  to control  effective  and on  a f f e c t e d ( F i g . 4.8A).  the application,  Two  depressions  of the presynaptic  measured  o f t h e EPSP  recovered  caused  low DGG  paralleled  when  i t was  a n d APV ( 8 / 1 2 t h e LTP  tetanic stimulation.  which  110  (Ol  O-a-ferwwxt'pOle  I  PTxwfWrwjtoOW  COnA  <  \  1  KomSOnA  s  a  n  «  >-0'CWonV9lycint OwaSOnA  (cl  (01  i - O G t j t a m y ^ y c . * 0 0 nA  CO*A  COnA  C*2.]-Plocu»nt Ocwtn-ittK 5 0 nA  I  \ a  B  Fig. potential  4.8  and on t h e d e p r e s s i o n  A. T h e u p p e r the  lower  response tested, the  record  that  only  DGG  indicated  sweeps by  records  reduced  or  quisqualate ate were  (negativity  slice  abolished  obtained  effectiveness antagonists  both  DGG The  made  i n the presence  thus  upwards)  t h e two  effect  on  each  of  five  at  the  times  part  of  of the stimulus.  reduced of  i n  antagonists  uppermost  and s t r e n g t h  final  amplitude  taken  The  amplitude,  without  records  a n d PDA  with  the amplitude of  b u t was  the depressions  and k a i n a t e .  so that  the three  above.  i n d i c a t e s the time  and k a i n a i t e were  reduced  Of  Oscilloscope  ( a - c ) a r e shown  In another  volley  t h e EPSP  field  by e x c i t a n t s .  fibre  stimuli.  reduced  response  i s a p l o t o f EPSP  presynaptic  volley.  consecutive  B.  of  on t h e SC-evoked  of t h i s  of the graph  to successive  fibre  these  Effects of antagonists  t h e EPSP  t h e EPSP  and  caused  by  a p p l i c a t i o n s of quisqual-  the intensity  of s t i m u l a t i o n  o f t h e EPSPs matched  those  of the antagonists.  The  reduced  i n the presence  of the  excitants  i s not a t t r i b u t a b l e simply  which  to a smaller  EPSP.  Ill  When  i t was  ineffective  as  an  bath  applied,  antagonist  D-APV  of t h e SC-evoked  c h a n g e +S.E.M. - 0 . 5 + 0 . 3 % , n=5) d e s p i t e of  the depressant  4.10).  Under  stimulation washout shock the  actions these  had  no  of the drug  produced  o f NMA  on  conditions  effect  on  synaptic  a  brief  o f NMA  antagonism  activity high  responses, which  was (mean  (Fig.  frequency  amplitude.  a p o t e n t i a t i o n o f t h e EPSP period  M)  EPSP  the complete  t h e EPSP  and recovery  end of the r e c o r d i n g  (5X10  After a tetanic  persisted to  ( F i g . 4.10).  Discussion E f f e c t s o f e x c i t a t o r y amino a c i d a g o n i s t s •and a n t a g o n i s t s o n s i n g l e u n i t a c t i v i t y The  involvement  receptors  i n stratum  pyramidal  cells  the  existence  of postsynaptic  radiatum,  terminates of foci  and q u i s q u a l a t e  Biscoe  Straughan,  and  Andersen,  1975;  different same  site.  firing  may  The be  extracellular systems  short  ajL. ,  concentration  (Johnston  since  (Balcar  quisqualate  (Lodge  The a b s e n c e  e_t a _ l . ,  latency  and  1978). were of  or  as e x i s t  Qualitatively observed  at the  kainate-mediated increase  of the  neuronal  uptake  f o r L-glutamate,  1973) and p o s s i b l y f o r  1980 )  o f an uptake  L-glutamate,  1974; S c h w a r t z k r o i n  glial  and Johnston,  of  (cf.  rise  e t a_l. , 1979) s u c h  t o theC A l  firing  and k a i n a t e  continuous  input  acid  1 9 7 9 ) , was i m p l i c a t e d by  a t t r i b u t a b l e to the gradual  L-aspartate  agonist.  t o NMA  slow  elicited  e_t  t h e SC  iontophoresis  1966; Dudar,  Spencer  responses  (Laurberg,  a t which  L-aspartate  where  e x c i t a t o r y amino  do  not exist  mechanism  f o r this  f o r NMA may  also  112  DGG  nA  100  1 0 0 Hi 1 sec  !  APV  1 0 0 Hi 1 sec  •  100 n A  ri  25  .  •.S  •  mm  A'  mV  I  n  *  1  0 '  •  1 0 0 Hz 1 sec  Fig.  4.9  'generation stratum  of  by  frequency DGG,  in  for  arrows PSs  and  was  no  on  was  reduced  frequency  the  (lower  prevented.  LTP  100 which  responses  were  not  100  Hz  sthe  17  and  min,  for  3 min again  (45  .was  sec  completely min).  in  the  SC  the  bars.  sec  at  the  times  DGG  reduced  was  applied  stimulus  again  effects  over  of  had  LTP  to  high some  terminated  the  high  and  Although  able  of  intensity  resulted,  been  high  the ..actions identical  the  the  to  of  tested.  was  the  recorded  response  the  later  had  on  by  from  APV  potentiation  1  1  in  later  injection  recover  measured  for  LTP  and  iontophoretically  trace.  a  minutes  were  stimulation did  administered  period  APV  PS  s t i m u l a t i o n of  using  record)  the  Hz  recovery  Two  the  t h e IPS  Following  post-tetanic  After Hz  at  After  stimulation  short-lasting  min,  PS.  0.1  prevented  produced  stimulation.  of  on  durations, indicated  below  frequency effect  the  stimulation.  LTP  to  were  stimulated  the of  antagonists  response  •• mm  1  amplitude  antagonists  was  amplitude  of  The  radiatum  pathway  shown  LTP.  and  stratum  The  Effect  pyramidale  projection in  1 0 0 Hi 1 sec  was for  8  produce  time  that  113  D - 2 • Amino- 5-phosphonovalerote 5% 10" M 5  NMA 2 5 n A  S  N  S  s  s  s  s 10 mm  2-  >  0  ^  "' *'•'"'.•'• «-• \ ' — •  ......  J  3-1  t  100 Hz, I sec °  4.10  amplitudes  of  taneously  from  prevented excitation NMA  The  actions  SC-evoked 2  graph) (both  had  enduring  a  A  i n c r e a s e oE  dose  no e f f e c t  records)  brief  bath-applied  actions of  r e t u r n i n g t o normal  responses  of  dendritic  slices.  the depressant  (upper  After  t Hz, I sec '2 min.  J  Fig.  NMA  100  but  t h e EPSP  of  on t h e  recorded  simul-  the antagonist  which  iontophoretically-applied  on  low frequency  blocked  perfusate  tetanus  EPSPs  D-APV  LTP  and w i t h  ( 1 0 0 Hz  (lower  synaptic graph).  the recovery  f o r 1 sec) produced  amplitude.  of an  114  explain  its  tually  to  produce  reasonable bursting  pattern  has  of  of  spinal  of  in  in  the  latter  CAl  feline  spinal  been  vivo  of  even-  is  to  not  induce  bursts  i n dentate  (Flatman  a the  action  granule  and  cells  Flatman  to evoke  et  recording  of  (cf.  reported  (MacDonald  intracellular  but  ability  motoneurons  neocortical cells  study,  and  neurons.  neither  iji  neurons  firing,  composed  H o w e v e r , NMDA h a s interneurons  and  of  observed nor  of  i t s unique  response  been  adjacent  frequencies  firing  laboratory  a l . , 1983).  recruit  high  similar  potentials our  to  explanation  A  in  tendency  bursting  Nistri,  al. ,  et_  1978)  1983).  revealed  that  In burst  2 + activity  was  paroxysmal  triggered  by  an  depolarizing shift  1 9 8 3 a , b ; Wong a n d  P r i n c e , 1978)  for  bursts  In  the  rhythmic  this  connection  which  interacts  1981)  also  al.,  1983).  influx  i t is  with  produces The  for  the  of  NMA-induced (PDS)  and  (see  this  action  NMA  burst  significance modulation  of  of  Dingledine, basis  potentials reported  here.  of an  also  may  quinolinic  (Stone  CAl  NMA  cortical  a l s o be  that  receptor  firing  -dependent  the  interesting  the  Ca  and  neurons  acid  Perkins, (Ganong  receptor-coupled function  is  e_t 2 + Ca  discussed  below. As  i n other  selective  regions,  antagonist  hippocampal  neurons  potentiation  of  quisqualate.  Both  D-(-)-isomer.  The  of but  the types  APV the  was  excitations  effects  there  DAA  be  was  induced  a c t i o n appear of  to  NMA-induced  typically  of  found  were  to  by be  a  potent  and  excitation  of  a  concomitant kainate  exerted  qualitatively  by  the  and the same  115  as  those  other of  observed  CNS  regions  quisqualate  (Lodge  with  i t s  D A A has been  (Evans,  et. a _ l . ,  -phosphonate  1978);  reported  Francis  to increase  (Hicks  and  McLennan,  (Lodge  et. a _ l . ,  1978);  and  to  Francis  that  D A A has  ment  by  APV  suggestive  That  low  antagonized  but  currents useful  of  the  o f DGG as  potential  a  antagonists  reaction Hicks  of  concomitant  1982).  with  antagonist,  of  neurons imply  the  enhance-  here  i s also  APV  d i d not  evidence  f o r  there  when  that  NMA  a  (GDEE  them  quisqualate  significant ejecting  since  and  may other  PDA) useful  had as  of the L-glutam-  antagonism with  inter-  substance  less  L-  receptors  higher  this  DGG  and  proposed  especially  i s consistent  with  as  was  The d e c l i n e the  dipeptide  two d i f f e r e n t  rendered  with  the  the  receptors  i n the hippocampus.  and L i u ,  and  direct  supports  indicating  which  of L-glutamate  cord  observations  However,  quisqualate  these  excitations  spinal  (as w e l l  However,  of  quisqualate  actions  response  quisqualate  1981).  used,  doses  kainate  and q u i s q u a l a t e  were  of the  obtained.  to  action  effect  spinal  reported  no  In  L-glutamate  and  effects,  and  quisqualate  blockers  complicating  ate  not  These  kainate  ACh,  has been  and Watkins,  reduction  and  iontophoretic  of kainate  (Davies  to  responses  aspartate)  1978).  the  activity  frog  excitatory action.  responses by A P V  1979)  depolarize  excitatory  quisqualate  o f a weak  excitation  be  subliminal of  potentiate  action  and Watkins,  and  spontaneous  gyrus  APV.  t o enhance  e_t a _ l . , 1 9 7 9 )  dentate  (Evans,  analogue  a  by  DGG  of  preferential  receptors  (McLennan,  116  Previous responses  amino  Koerner  acid  indicate  the  isomers  et  a l . (1979)  and  those  of Hori  recent  spinal  demonstrated  APV  response  a  been  Blockade study  and  by  receptors.  APV  e_t a _ l . ,  through  t h e NMA  clearly,  the  antagonism  and  of  any  Our  one  of  CA3  of  spinal  cord,  of  of the  imply  More  t h e NMA  The  acid-  response  L-aspartate the  effect  responses  latter  (data by  not  has  shown). in  spinal  L-APB  and  action  D-APB  isolated that  1982)  differences.  Typically,  L-APB  cortex,  amino  i n the  region  in  1982)  this cord  excitation  receptor. of  formation the  observation  L-glutamate-induced  Evans  extent.  dose  by  of  Watkins,  D-APB  itself.  inhibition  hippocampal  frog  acid-induced  effects  (Evans  Quite in  the  the data  reflect  Our  are produced  and  by  1981).  by  antagonism  cause.  similar  smaller,  amino  agonist  D-APB  to a  antagonism  cells i n  with  i n the  simply  e_t a l _ . ,  synaptic  assumed  actions  (Davies  albeit  of  underlying  antagonism  may  APB  i n the r a t o l f a c t o r y  kainate  enhance  to excite  the  the  DL-APB  D-APB  Hori  to  of  mediated  L-APB  which DGG,  have  different  studies  observed  preparation is  and  by  e t a l _ . , 1 9 7 8 ; W h i t e et.  In agreement  decline,  was  be  with  cord  and  occasionally also  NMA  (but see  L-APB  quite  1981)  e_t a _ l . ( 1 9 8 1 )  responses  with  (Dunwiddie  to  non-specific  paralleled  of  that  obtained  antagonized  As  inhibition  Cotman,  o f APB.  D-APB  induced  and  receptors  results two  of  i n the hippocampus  a l . , 1979; of  reports  three that  excitations  synaptic cannot  transmission be  attributed  'conventional' L-APB also  d i d not argues  by  amino  to  acid  antagonize against  i t s  117  interaction preclude  with  a  novel  antagonism  glutamate  of  the  receptor,  receptor  b u t does n o t  f o r an  unidentified  neurotransmitter. Although antagonist, effect  on  agonist  i t was  PDA  also  had  C A l neurons  e f f e c t s o f PDA  (Davies,  Evans  synaptic  mechanisms  a potent,  e_t a _ l . ,  a  non-selective  direct  (cf.  Segal,  have  been  1981).  would  amino  APV-sensitive 1981).  excitatory  Partial  observed  acid  NMA-like  i n the spinal  cord  Interpretation of actions  be c o m p l i c a t e d  by t h i s  on  depolarizing  action. Although quisqualate (Davies Evans  responses  selectively  and W a t k i n s , e_t a _ l . ,  conclusion  reports  and  McLennan  dentate  because  e t a l . (1976) and Wheal  where  reasonable  the explanation i t s actions  antagonist  effects  excitations reported  of  of  CAl  tested  investigations,  This  study  neurons  DGG  while  conform  cord. and APV  others  such  may as  were  on  amino  in  general  be D-  found. may  o f GDEE  t o C A l seems  show  useful  Hicks  f o rthe  discrepancies  The data  to the  and of  (1980)  the value  the  i t s a c t i o n s as  specificities  the antagonists  support  i s i n contrast  and M i l l e r  input  in_ v i v o  1979; but see  f o rC A l neurons  are unclear  f o r the spinal  antagonists  of this  antagonize  cord  and Lodge,  for. these  of e x c i t a t o r y synaptic  The  to  e_t a l . ( 1 9 7 9 ) w h o d e s c r i b e d  (1979)  gyrus  Whatever  1979; McLennan  inexplicable".  of Spencer  ability  i n the spinal  1979) t h e r e s u l t s  of White  "capricious  and  GDEE h a s a d e m o n s t r a t e d  be,  as  an  limited.  acid-induced to  that  those of the  i n synaptic  and L-APB,  PDA a n d  118  GDEE  may  and/or  be  have  of  lesser  value  complicating  because  they  direct actions  are less  potent  on C A l c e l l s .  E f f e c t s o f e x c i t a t o r y amino a c i d a g o n i s t s and a n t a g o n i s t s on s y n a p t i c a c t i v i t y The  discrete  predominantly  Schaffer  ably  smaller  Both  projections  (Swanson  electrical  extent  collateral  commissural  appear  e_t a l _ . ,  produced  by  other  stratum  radiatum  and remain  terminals  originate  on  field  EPSP  membrane currents; involved  the  a  by  since  fibre  volley  EPSPs  also  i n some  was  essentially  1979).  identical  excitants  depolarizing produced  site  shunt  of action  may  reduction  of  a The  to  CAl  cells  reduced  the  increasing of  the  postsynaptic  also the  agonist-induced  sensitivity to that  a  and/or  through  proportion  et_ a T . ,  cases  a  a large  (Goldowitz of  so  cut slices  of  evident.  exhibited  course  i n transversely since  neurons  homogeneous.  dendrites  which  presynaptic  t o be  which  1981).  1981) and  apical  application  presumably  conductance  intact  e t a_l. ,  S0rensen,  systems  consider-  t h e same C A 3  assumed  consequence  f r o m CA3 n e u r o n s  Iontophoretic  be  fibre  be a t m o s t o f m i n o r  the  and  activated  to a  (Voneida  t o o r i g i n a t e from  Effects  of  fibres  can probably  used  and probably  1980; Laurberg  pharmacologically  would  stimulation  have  been  presynaptic  reduction  antagonism  which  of the e x c i t a t i o n of  of was  single  neurons. The various ficity  blockade  excitants of  an  may  of the depression therefore  antagonist  as  be  of EPSPs  used  i t i s tested  t o check against  induced  by  the specia  synaptic  1 1 9  response are  used  EPSPs to  (Figs. to  can  be  eject  be  synaptic  which  assumed  site  than  is  that  as  then,  not  the  is  then,  large  can  under can  the  be  from  can  which  be  inferred  in  mediating  antagonist  is  not  as  also  be  the at  synaptic  that  a  the  field  the  same  neuro-  responses  given  type  response,  reaching  the  such  excitatory  where  NMA,  assumed  antagonists  endogenous  conditions  or  generating  depressions  area  currents  kainate  excitant that  turn  whole  does  i t  the  as  high  I f exogenous e x c i t a n t s act  involved  the  In  when  such  region  these  i s produced.  unaffected,  hence  response.  reaching  example,  agonists  entire  be  transmitter"*",  receptor  the  For  and  prevent  subsynaptic  are  potent  can  to  potential  4.8B).  abolished  affecting  recorded APV  4.7,  the  of  rather synaptic  receptor. The  depressions  quisqualate due  to  uptake  contrast, and this PS  were  the  slowly  relatively  of  these  more  potent  by  was  locally  the  L-glutamate,  weak  and  (see  agonist  kainate  of  the  i n d u c t i o n or  (Fig.  4.6A)  L-aspartate  short-lasting,  compounds  reversible reduction  action there recorded  induced  or  probably  Introduction). produced  EPSP. increase at  and  the  a  massive  Concurrent in size cell  In  body  with  of  the  layer  R e c e n t l y L y n c h a n d h i s c o - w o r k e r s ( F a g n i et. a l . , 1983) reported a reversible densensitization of physiological r e s p o n s e s t o b a t h a p p l i c a t i o n s o f L - ( a n d D-) glutamate which was n o t p a r a l l e l e d by a r e d u c t i o n o f t h e S C - e v o k e d E P S P . This e v i d e n c e s u g g e s t s t h a t e x o g e n o u s g l u t a m a t e may act p r i m a r i l y on e x t r a - s y n a p t i c s i t e s and/or that L-glutamate i t s e l f does not serve a t r a n s m i t t e r f u n c t i o n i n t h e SC p a t h w a y . Nonetheless other evidence provided h e r e i n d i c a t e d an accessa b i l i t y o f i o n t o p h o r e t i c a l l y - a p p l i e d c o m p o u n d s ( e . g . DGG) to sub-synaptic sites.  120  (cf.. C h a p t e r of  synaptic  5).  excitation  Although there  were  comparison produced volley  with  presynaptic of  was  dentate  those  of  have  gyrus  First,  (but see  with  to a  Ferkany  ion  hippocampal  (Evans,  the amplitude from  to  be a s s o c i a t e d  an  effect  on shown  probably  due  to  2+ dependent A  inward  residual  observed  Ca  increase  increase  o f t h e EPSP an  with and/or  to  the this  f o r long  effect EPSP  which i n the  p o t e n t i a t i o n may  have  i n membrane p o t e n t i a l o r neocortical  the depolarizing an i n c r e a s e d  the  NMA  fibre  1982) due  i n  of  i n the  path-evoked  This  EPSP  Secondly,  motoneurons,  sometimes w i t h  doses  al. ,  depression,  n o t shown).  have  et  the  interaction  1980).  the perforant  Studies cells  high  concentrations  been due t o a l o n g - l a s t i n g a l t e r a t i o n conductance.  of  i t sactions  direct  recovery  (data  to  been  potentiated  obtained  aspects  reduction  due  chapter.  depression  APV-sensitive  depolarizations  following  the p o t e n t i a t i o n  i n the next  large  kainate.  extracellular  often  also  a  different  receptors  postsynaptic  periods  produced  very  may  underlying  are considered  pronounced  which  compound  NMA  two  a  changes  The mechanisms  actions  input  activation  of  and  of  NMA  resistance, a  voltage-  + and/or  Na  i n membrane  current resistance  (see Introduction). could  explain the  A l t e r n a t i v e l y , t h i s a c t i o n may 2+ r e f l e c t e v e n t s t r i g g e r e d by an i n f l u x o f Ca either pre- or p o s t s y n a p t i c a l l y s i n c e ( 1 ) NMA ( a n d L - a s p a r t a t e ) p r o d u c e s an 2+ 2+ increased i n t r a c e l l u l a r Ca concentration ( [ C a ]^) (Padjen and  Smith,  NMA  t o produce  i n t h e EPSP.  1981; M a r c i a n i "calcium  e t a l . , 1982) and (2) an a b i l i t y  spikes"  i n hippocampal  CAl cells  of  has  121  been  reported  a l , , 1983).  (Dingledine,  As  pointed  1983a,b;  out  by  see  also  Dingledine,  Flatman  this  et  rise  in  2+ [Ca  ]^  this  could  affect  connection,  antagonists  a  i t  DGG  number  is  and  of  biochemical  interesting  APV  that  prevented  LTP  processes.  the  NMA  since  In  receptor  postsynaptic  2 + Ca  entry  Lynch,  has  1981; The  that  unlikely  that  the  slice  lateral lower  ejection  the  this  pathway  unit  activity  reported  DGG,  of  a  the  and  by  at  the  not  also  slow  abolish  i n CAl  firing  It  highly  is was  due  since,  to  first,  gyrus and  the DAA  with  did  of  NMA  investigators implies  other  electrode not  affect  of  single  antagonist  transmitter  the much  third, same  of  second,  antagonized  i n d i c a t e s the  onset  in  terminated;  from  NMA  involved  APV  dentate  That  the  dendritic region  shown);  selective  endogenous  other  with  from  to  itself.  the  applied  the  increases  fibres  into  hippocampus  Furthermore,  here  and  Applications  receptors  response.  being  i s not  effectively  (data  .SC-evoked  i n the  APV  drawn  required  antagonism synaptic  input  example  while  (Baudry  pathway.  EPSP  stimulated  currents  interaction  receptor.  to  SC  those  of  of  be  receptor  EPSP o r  change  path  can  i n the  the  electrodes  for  NMA  administered  the  same  phenomenon  that  than  absence  perforant  depressed  no  were  where  substances,  any  or  of  accessibility  antagonists  the  greater  the  the  the  excitation  depressions  little  inadequate  conclusion  were  produced  in this  a l . , 1983).  i s that  synaptic  NMA-induced  using  et  studies  mediating APV  Lynch  implicated  firmest  antagonist  of  been  absence  with  the  of NMA  excitations that  i t would  122  not  be  involved Unlike  the  consistently ejection and  in fast  synaptic  selective  depressed  currents  the  excitation that  either  synaptic DGG  single the  no  the  separate  action  CAl  in  Furthermore,  or  the  the  the  evoked  by  dentate  gyrus  input  topical  reduced  the  potential  or  selective  1979)  cord did  await  antagonists  That spinal  receptor  not  GDEE,  an  the  to  the  any  the  synaptic  be  that  indicates  caused  to  by  a  terminals. blockade to  of  account  of  the  path in  EPSP  in  the  membrane  excitability  (Crunelli  has  mentioned  the  briefly  hippocampal affecting  that  synaptic of  quisqualate  affect  possible  volley  change  effective  Watkins,  the  mediates  unlikely  compounds.  and  and  observation  to  f o r these  (Davies  EPSP  perforant  development  kainate-  receptor  without  Confirmation is  high  reduction  (1981)  EPSP  the  unrelated  with  DGG  with  presynaptic  neuronal  a p p l i c a t i o n of  resistance.  of  medial  Segal  and  fibre  DGG-induced the  DGG  therefore  unlikely  effects  r e s i s t a n c e or  SC-evoked  must  is  APV,  The  the  associated  Finally,  quisqualate  pathway  the  not  a l . , 1982).  that  or  was  is  pathway. on  and  the  i n t e r a c t i o n s are  s t i m u l a t i o n of  potential, et  since  of It  excitability  postsynaptic  action  SC  DAA  i t also blocked  quisqualate  effect  neurotransmitter-receptor for  this  neurons.  antagonism  on  response  depressions  appreciable  synaptic  antagonists  t o do  kainate  excitation  had  that  of  NMA  synaptic  required  quisqualate-induced  transmission.  1979; response  membrane  e i t h e r the receptor more  does  not  kainate in  this  potent  and  antagonist  McLennan  slice  and  in  the  Lodge,  preclude  a  123  role  of  quisqualate  factory  antagonism  agonist  b y GDEE w e r e The  mechanisms to  but  of  less  excitatory  depression  of  APB  could  stereoisomers,  than  DGG,  of  slices  with  whereas  explain  EPSP  et al. ,  doses  an e f f e c t  why  (Dunwiddie  et. a _ l . ,  shown  that  On  1978; Cotman  Thus,  on  weakly  for  amino  although  the  following perfusion  o r both  hand, able  et_ a _ l . ,  mixture  1979)  t h e NMA  was  L T P c a n be p r e v e n t e d  similar  was  et al. ,  the other  the racemate  L-APB  weak  different  profile  of either  o f D-APB  to this  of the racemic  1978; White  be d u e t o a n a c t i o n  by  activity  observed  satis-  similarly  acted  antagonist  high  not involved.  here  each  had a pharmacological  devoid  (Dunwiddie  presumably  were  but probably  o f t h e SC-e,voked  hippocampal  might  o f APB  since  C A l neurons  e x c i t a t i o n s of CAl c e l l s .  of  etically  isomers  potent  and  acid-induced  obtained.  D-APB  pathway  of single  never  t h e EPSP  since  i n this  of responses  optical  depressants  receptors  theorof the  receptor i s  such  an a c t i o n  to abolish  1981),  since  b y A P V , a n NMA  we  LTP have  antagon-  ist . To aspartate  attribute  by t h e f a c t  interacting  amino a c i d activated 1981;  b y NMA,  to  either  transmitter that  to varying  receptors  McLennan,  relative  role  as t h e n a t u r a l  complicated of  a  both  extents  1981).  amino  sensitivities  acids  with  of  may  the three i . e . those  and q u i s q u a l a t e  I t has however  or  L-  i n t h e SC p r o j e c t i o n i s  so f a ri d e n t i f i e d ;  kainate  L-glutamate  (Watkins  be  excitatory selectively and Evans,  been suggested  L-glutamate  and  capable  from t h e  L-aspartate  to  124  blockade  by  a  range  preferentially qualate  and  1981).  Since  favor the  synaptic  the  related  receptors and  Evans,  plays  but  be  projection  no  the  role  CAl  demonstration sufficient  qualate  evidence  i n the  that  neurotransmitter  in  region  amounts  to  while  of  natural and  the  electro-  pathway.  other  (Stone  has  By  involved  in  on  NMA  the  acidic  amino  ligands  Perkins,  of  shown t h a t  for 1981;  the  having  of  no  pathway  the  pathway.  in  detectable  acid  the  NMA  SC  However,  when  kainate  excitability, amino  LTP  r a t hippocampus. this  the  synaptic excitation  generation  antagonize  excitatory in this  study  affects  postsynaptic an  SC  action  the  in  Introduction),  more  that  mediation  the  DGG  or  be  present  that  receptors,  presynaptic  may  excluded  the  involved to  be  Evans,  1981).  conclusion,  receptor  be  the  could  however,  compounds  cannot  in  quis-  seen  other  (see  acids  the and  is  with  predominant  possibility,  these  in  transmitter  a  with  antagonism  evidence  by  or  may  the  amino  (Watkins  together  plasticity The  In  data,  L-aspartate  acids  Watkins  of  these  cord  respectively  neurochemical as  that  spinal  pattern  present  and  the  reasoning,  receptor.  the  similar  L-glutamate same  in  receptors  a  physiological  antagonists  interact  NMA  hippocampus,  of  and  applied quis-  effect  provides  is utilized  on  strong as  the  125  CHAPTER E F F E C T S OF  K A I N I C A C I D ON I N THE  5 SYNAPTIC  HIPPOCAMPAL  PROCESSES  SLICE  Introduction Initially the  seaweed  Digenea  application  i n Japan  translation More its  of  excitatory  and  nervous  its  potency  be  10-200  al. ,  fold de  structurally property Konishi, and  of  1970)  name  has  acid,  generated  killing  of  an  by  extract  i t s  Ascaris;  i s "monster  received  of  literal  the  sea".  because  i n the  of  clinical  the  attention  properties  an  action  of kainate  Konishi  (1970)  excitant  was  greater  than  Montigny  kainate  Usherwood,  and  and  on  was  rat  first  of  mammalian  both the  1975)  that  None  of  produced  effect  i n the  is  the  the and  of  L-glutamate 1981)  vertebrate  to  facilitation acidic  underlying  (Biscoe  intriguing  junction  amino  et.  i t is  (Shinozaki  of  and  others to  which An  CNS  neuromuscular  other the  neurons  s u b s e q u e n t l y shown by  Tardif,  insect  demonstrated  cortical  ( J o h n s t o n e_t a _ l . , 1 9 7 4 ) .  L-glutamate.  unknown.  the  neurotoxic  related  this  was  kainic  system.  and as  1976;  simplex,  acid  excitatory  Shinozaki  in  i t s Japanese kainic  The  interest  in  recently  central  by  the  (Daoud  responses acids  mechanism  and  to  tested remains  )  126  Studies existence brain  of  and  of  a  a  low  affinity  second  high  predominantly B e r g e r and after  in  postsynaptic  receptors The  affinity  e_t  for  results  kainate  both  is  of  the  also  binding  have  isopropylene  usual  that  three  required  charged  for  Furthermore  the  the  kainate or  observation  which  from  Watkins, mixed 4).  or are  et  at  Together acid  that  quisqualate  these  data  receptor.  However  questions  which  Kainic  acid  is  an  effect  1974),  dendrites  indeed  the  remain  and  NMA  as -  well  Chapter  a_l. ,  of  as  the  4)  are  receptor. those  for  1980),  an  current  concept,  (McLennan,  receptors  i t s role  unique  1981;  behave  (cf.  as  Chapter  aspects  i n neuronal the  and  investigations  L-aspartate  the  1979).  1979b)  from  the  and Coyle,  al.,  distinct  and  nature  and  its  analyses  emphasize  pre-  with  et  1979a;  extrasynaptic et  chain  with  and  of  1974)  (Biziere  L-glutamate  and  neuronal  are  a  Coyle,  kainate  consistent  dysfunction  al. ,  of  sites  L-aspartate  1981a,b),  al. , side  pharmacological  agonists  kainic  binding  is  and  (cf. Introduction  interaction  L-glutamate  derived  groups  e_t  occurring  (London  (Davies  (London  rat  characteristics  indicate  existence  the  the  Coyle,  binding  likely  (Johnston  site  and  receptors  electrophysiological shown  (London  pathways  revealed  throughout  D  Changes of  a_l., 1981);  kainate  have  (K =10y*,M)  regions  afferent  l o c a t i o n of  Foster  binding  D  1983).  of  acid  (K =50yxM) s i t e  forebrain  Ben-Ari,  lesions  1979a;  [ H]-kainic  of  the  function  endogenous  ligand  unanswered.  also  powerfully  which  perikarya  is  neurotoxic  exerted  (Olney  primarily  et. a l . ,  1979);  (Olney against axons  127  of  passage  and  terminals  doses  of  the  amino  acids  their  injection into  compound excitant al. ,  compound cause  being  1981).  studies,  the  proposed  by  that  somata  synaptic  a c i d i c amino a c i d s .  per  i t  is  which  a  change  leads  by  nmole  and  axons  of  passage  tool  1978b). usefulness additional  for  in  the  was  of  some  the  i t  (Wuerthele local  to e_t and  et  than  years in  and  and  the  this  binding on  neurons  , 1978).  distant  The  lesions  the  conductance  most  of  toxic  ions  of  the (1-5  lesion  used  a  while neuro-  structure-function and  (cf.  was  for  producing  McGeer  e_t  from  mechanisms  a l .,  that  compromised  remote  are  of  injections  i t recognized  regard  and  depolarization  widely  system  was  e_t  excitatory  neuronal  diseases  later  destroy al.  the  focal  nervous  an  (1971)  located  intracerebral  became  as  death.  amongst  a  each  "excitotoxic"  membrane  cell  of  Nadler  ajL.  toxic  investigation  central  kainate  action  the  to  produce  neurodegenerative Only  the  receptors  increased  since  could  of  so-called  acid  1)  correlations  site  kainic  1  biological  models  the  of  intracellular concentrations  amino a c i d s  in ^a.  sparing  of  potency  e_t a l . , 1 9 7 8 ;  Rather  unknown p r o c e s s e s  Because excitatory  that  site  effectiveness  Olney  both  of  produces  the  electrophysiological  mediate  likely  by  large  excitatory  at  neurotoxic i t s  when  Other  observations,  originally  save  lesions  Schwarcz  actions se  the with  these  supported  and  neuronal  brain,  on  spared  administered.  e_t a _ l . , 1 9 7 8 ;  suggests  dendrites  normally  parallel  Based  subsequently  are  similar  in  (Coyle  hypothesis,  are  the  by  an  injection  underlying  produced  the  by  the  mechan-  128  istically  distinct  The  processes.  involvement  of  destruction  of  experiments  i n the striatum.  Biziere tion  and C o y l e  of  the  suggested the  neurons  indirect  by  (1978)  kainate  demonstrated  t o use L-glutamate of  kainic  acid  enation  not a t t r i b u t a b l e to a  factors  was  that  by  the prophylaxis  (McLennan,  at  change  The  confirmed  i n i_n v i t r o  studies  where  cells  co-cultured  (Whetsell  et a l  and McGeer  interaction cause  between  neuronal  against example,  (1978)  this while  kainate  striatum  injection  L-glutamate,  GABA of  (Biziere  and C o y l e ,  the serotonergic  kainate  i n the  hippocampal glutamatergic  perforant  that  a  also  pathway  presynaptic cortical  and  a  recent  co-operative  evidence  some  action  1980).  extent was  imparts et  a_l. ,  ablation blocked  For  i n the by c o -  produced  protection 1982).  to  argues  neurotoxicity  to  1980).  L-glutamate  and K o h l e r ,  was  striatal  In addition a chemical  (Berger  prior  of  neurotoxicity  proposed  kainate  of  deaffer-  1979; P a n n u l a ,  M  been  reduced  this  cortical  similar  1979).  striatum  formation  but  i s restored  input  has  two o r d e r s  by  transmitter  (Schwarcz  i t i s true  destruc-  for intact  kainate  and  destructions,  deafferented of  of  originally  hypothesis  from  of the postsynaptic  requirement  i n the expression  McGeer  which  least  afforded  afferents  were  the p r i o r  1980) t h e i n v o l v e m e n t  indicated.  local  first  as i t s n e u r o t r a n s m i t t e r ,  Since  excitability  the  suggested  projection,  magnitude. was  was  in  T h u s , M c G e e r e_t a l . ( 1 9 7 8 a ) a n d  corticostriatal  neurotoxicity  actions  by  lesion against In the  of  the  reputedly  the  destruction  of  129  dentate  granule  immunity  was  cells also  septohippocampal Furthermore,  although  commissural  pathway In  after  excitatory and  the  mitter  cases  specific lesion  uses  and  and  to  be  kainate  optic to  correlated  (Ferkany  with  presynaptic  (Streit  i n the  time  postsynaptic Although  1980).  releasing  lesion  of  kainate's  and  the lethal  toxicity  persists  possibly in  the  as  does  only  cochlea  i t s  not  protection  while  a  trans-  granule  cells  i n the  striatum  a  Collins 1980)  hippocampus  (Nadler  the  or  recruits  for  the  and  McGeer,  onset  of  an  was  slow  and  terminal  i t may  located  e_t a l . , 1 9 8 3 )  necessary  by  locally  fibre  observation  kainate  provided  (McGeer  for  receptor  in  from  e_t aJL. , 1 9 8 0 )  required  appear  septohippocampal  neurodegeneration  latter  with  factor(s)  the  inputs  tectum  (Evans,  prior  same  cholinergic  input  L-aspartate  dentate  or  et_ a _ l . , 1 9 8 2 ;  a  kainate  Thus  1980),  interaction  an  originates  or  for  other  the  be  transmission  protects  the  the  the  1979).  basis  with  to  major  which  impulse  the  From  depolarization  lethal  the  kainate-mediated  degeneration. an  of  but  Cuthbertson,  attentuate  deafferentations.  and  immunity  that  of  of  and  nucleus  pathway  Gulley,  Cuthbertson,  1978)  not  L-glutamate  immediately  injected  did  cochlear  Abolition most  (Nadler  L-aspartate,  afferent  (Bird  removal  i t appears  destruction  probably  injected kainate  by  projection  and/or  even  locally  conferred  L-glutamate  actions.  by  be on  by  and  inferred terminals  an  indirect  an u n i d e n t i f i e d  expression  of  i t s  effects.  other  acidic  amino a c i d s  also  destroy  neurons  130  when  they  are  mechanism  of  likely. (NMA)  In and  a  (Nadler  with  hippocampus  septohippocampal Schwarcz The  and  with  n e u r o t o x i c i t y of  the  excitotoxic requirement specific by  a  elements  in  i s not unique  is  The structures  hand  other  potencies  exists least  In  with  projections  CA3  sensitive  contrast  lesion  as  region  to  neurotoxicity  a prior  the the  local local  connection  other seem t o brain  indicates simply  ).  of  the  in  the  either  (Kohler  toxicity neuronal  presynaptic  in this  would  afferents  kainate  rather  theory  in  obvious  n e u r o n s ) and  excitations  the  et a l . ,  1980).  involved  cochlear  and  ibotenate  path  an  most  seem  pharma-  hippocampal  kainate  1978  perforant  one  kainate  to  the  aspartate  receptors  each  the  a_l. ,  Kohler,  of  into  being  with  in  kainate  exhibit similar  by  T h u s on  independently  4)  sensitivity  e_t  and  N-methyl  mediating  attentuated  or  factors  complex.  those  kainate,  i s not  compounds  interact  injection  neurons  differences  quisqualate,  Chapter  differential  situation  are  from  of  upon  respectively  1979;  which  distinct  dentate  these  hippocampus  Introduction  whereas  intracerebrally  of  ibotenate  neurotoxins  and  action the  cologically (see  injected  apply.  that  r e l a t e d to  the  kainate  death  occurs  contribution there  is a  On  the  (e.g.  (e.g.  similarity  e x c i t a t o r y amino a c i d s  regions  co-operative  of  other  and  the  hand  the  striatum)  destruction  of  neurons  i t s excitatory actions  interaction  with  kainate  various  for  but  presynaptic  suggested. direct such  injection as  the  of  amygdala,  into  striatum,  forebrain  thalamus  or  131  hippocampus  produces,  destruction  of  particularly  within  of  kainic  and  Ben  A r i , 1983).  the  neurotoxin  et  of  in  man,  of  distant  lesions  locally  claustrum-insula  a l . ,  1978;  (1-5  nmol)  the  et. a _ l . , 1978 ) .  Noting  associated with colleagues  that  intrinsic  processes  high-frequency  discharge  of  has  see been  abolish  by  reducing  and  attenuate acute  caused  supported  treatments (Fuller  diffusion  below)  Olney, and  transection remote  propagation convulsants  lesions  (Nadler such  as  limbic  distant  Ben-Ari et  excitatory indicating  allylglycine  lobe  similar most  in  the  epilepsy that  the  injections.  from  the  repeated  This  1979)  can  requirement 1980); e_t  site  hypothesis that:  or  (1)  enhancing  seizure  distant  (Meldrum  seizures  injection  observations  afferents  They  the  neurons d u r i n g  Cuthbertson,  and  electroencephalo-  a l . , 1982)  a  a  suggested  the  the  nuclei  similarity  damage.  respectively  and  a  et. a _ l . ,  of  amygdala,  among t h e  t r i g g e r e d by  kainate  Fuller  being  kainate  Berger  injection the  temporal  of  experimental  1979;  of  of  the  (e.g.  augment  by  the  high  Parenteral  (1978)  consequence  produced  a  thalamic  1982).  region  a  in  some  CA3  his  has  a d m i n i s t r a t i o n s produce  proposed  (but  and  and  Ruth,  lesions,  were  the  which  the site,  ( F o s t e r et. a l . , 1 9 8 1 ;  seizures  than  injection  system,  graphic  rather  the  lesions,  produces  and  lesions  limbic  receptors  lesions  Nadler  from  local  intrahippocampal  (Nadler  distribution  the  example  limbic  vulnerable  to  For  intraventricular pattern  remote  the  acid  cortex,  (Wuerthele  addition  neurons  density  pyriform  in  activity  damage;  (2)  reduce  or  for (3) a_l. ,  impulse other 1974  ),  132  bicuculline (Olney  (Meldrum and  e_t  limbic  a_l. ,  1980)  structures;  tions  of  the  pyramids  (4)  high  1980)  also in  the  focal  can  Damiano,  with  development  dipiperidinoethane lesions  1981a).  the of  throughout  electrical  destroy  deafferentation  consistent  and  frequency  pathway  and  e f f e c t s of  mechanisms  e l i c i t  perforant  protective  1973)  can  and  (Sloviter  are  Brierly,  hippocampal The  (Nadler  and  and  CA3  differential Cuthbertson,  involvement  distant  stimula-  of  separate  local  neuronal  degenerations. More seizure  activity  damage. reach  recent  In  the  sites  with  since  from  cannot  regard  remote  incorrect rapidly  alone  this  particularly  experimental  of  has  site  of  1979;  et. a _ l . , 1 9 8 0 )  produce  ibotenate,  local NMA  and  at  kainate  induce  only  Guldin  French tion  et  a l .  between  Therefore, the  and  production  least local  have  kainate's  of  brain  seizure  that  to  1982;  and  epileptogenic  and  appears  distant lesions probably  wide  McGeer,  too  small  occurring and  al.,  apparent neurotoxic to  a  of  electroencephalo-  (Zaczek  an  be  injections  A l d i n i o et  demonstrated  and  diffuses  achieves  those  degeneration  may  kainate  which produce  not  injections  in concentrations  comparable  does  administration that  that  distant  kainate  Intracerebral  activity  the  (Scherer-Singler  albeit  Markowitsch,  (1982)  while  the  quisqualate  seizures  explain  after direct  shown  lesions.  graphic  1982;  damage  indicates  i t s a p p l i c a t i o n and  throughout  to  assumption  been  distribution Zaczek  adequately  intraventricular i t  the  the  evidence  be  also  with Coyle,  1983);  and  dissociaactions.  a prerequisite involves  other  133  changes are  which  caused  only  The role  particularly  al. ,  the  actions  and  of  distant  local  or  of  acid  convulsants  e_t a _ l . , 1 9 8 1 )  of  and  actions  1981).  are  kainate.  that  kainic  ( L o t h m a n et.  synaptic  excitation  epileptogenic  i n the and  To  e x c i t a t i o n might  be  effects  p r o c e s s e s were  region,  cells  Therefore  the  pivotal  hippocampal.  of  slices  CA3  a  acid-induced  i n t h e CA3-CA4  neurotoxic  neurotoxicity  on s y n a p t i c  kainic  investigated.  synaptic  to play  i n hippocampal  the  were  of  potentiates  McLennan,  kainate  enhancement  amino a c i d s  the  greatly  mechanisms  of  to  b e e n shown  and  (Collingridge study  suggested  epileptiform discharges  1981)  amino  propogation  (Ben-Ari  sensitive  causes  been  e s p e c i a l l y those  has p r e v i o u s l y  acid  has  and  activity  cells,  excitatory  kainate.  onset  epileptiform  It  by  the  hippocampus  i n the  pyramidal  amongst  also  present  potentiating  determine  i f the  related  to i t s  of  other  acidic  assessed.  Methods In prepared Each  this and  study  maintained  experiment  obtained outlined  transverse  from  was  described  performed  separate  i n Chapter  as  hippocampal  at  animals.  2 were used  previously  least  3  times  slices (Chapter using  Conventional  to stimulate  were 2).  slices  techniques  and r e c o r d  evoked  responses. For  local  multibarreled cedures. solution  application  electrode  For experiments i n 100 mM  NaCl  using  kainate standard  was  ejected  iontophoretic  i n C A l and t h e d e n t a t e (pH 8.0)  filled  from  gyrus  a  pro-  a 50  t h e b a r r e l s ; i n CA3  mM 5  134  mM  kainate  used  i n 160  mM  for current -8  kainate  (10  NaCl  (pH  controls.  7.6)  With  was  used.  bath  NaCl  (1-4  M)  was  application solutions  of  -4 -10  M)  were  freshly  prepared  i n the  ACSF.  Results E x t r a c e l l u l a r l y r e c o r d e d e f f e c t s of on e v o k e d and s p o n t a n e o u s a c t i v i t y  kainate  CAl When sec)  applied  e i t h e r to  apical  dendrites  kainate  amplitude PS  tion,  In  following degree  pre-treatment ted  with  PS  interacting produce of  these  kainate  body  with  i t  20  anti-  or  2-4  latency  longer  appealing  synaptic  applied  layer indicated that  either  to  extra-synaptic  excita-  (50-300%  of  for  a  correla-  PS.  that  i t  was  dendrites  to  similar  dendrites sites  of  synaptic  the the  of  repro-  consider  the  the  was  f o r the  in  in  increase  of  processes  and  orthodromic  sometimes  to  EPSP  period  mV) , was  e l e c t r o p h y s i o l o g i c a l changes  when  a  and  slices  onset  the  P o t e n t i a t i o n of  and  of  to  radiatum,  increase  kainate-induced  min  10-120  or  stratum  i n t e r r u p t e d by to  for  SC-evoked  Enhancement  of  the  is  the  v a r i e d between  amplitude of  or  the  recovery.  a decrease  Although  slices  which  the  ( F i g . 5.1A).  both  lasted  of  of  PS  preceded  most  in  nA  pyramidale  long-lasting  orthodromic  excitation  the  distant  and  responsiveness  activation.  ducible  m  (50-100  in stratum  reduction  large  usually  depressed  synaptic  a  the  was  bodies  100-300  a  of  iontophoresis  cell  produced  concurrently  the  the  by  or  effects the  action  cell might  135  Koinole  Koinote  50 nA  50 nA •  I  o  >  E«  -y  (b) n  •to)  n  (c)  td)  10 msec  (I)  (e)'  n  n  Koinole  IO"'M  5  15mm  ^  Fig.  5.1  iontophoresis  A.  into  Actions  the  dendritic  responses.  The  in  pyramidale  stratum  measured  at  Kainate  was  Following was  an  The for  was  to  records upwards)  the  (5 were 15  of  min  the  period lag  CAl  the  medium  in  the  was  the  PS  measured  field  EPSP  radiatum.  by  the  bars.  increased  and  there  was  the  by  SC-evoked  stratum  PS had  The  effects  of  potentiated  in  been for  bar;  perfusion  graph  the  The  exchanged  times  of  B.  was  the  the  EPSP.  region  by  on  indicated  superimposed  at in  PS  CAl  of  in  CA3  the  consecutive  breaks  of  indicated  obtained  and  time  the  administered  of  amplitude  region.  the  time  regions  iontophoresis for  which  kainate  plot)  depression  normal  the  the  (upper  reduction in  from  kainate  two  initial  kainate  slice  due  show  administered  an  removed.  the  site  associated  perfused this  graphs  the  of  the  one  surgically containing  delayed  onset  system.  The  sweeps,  negativity  indicated  measurements  (a-f). were  not  filmed  During made.  136  exist.  This  manner.  Kainate  responses  from  electrode  and  200yx.m  away.  actions  were  the  applied  locus  central  second Since  observed  also  i f the primary  produced  experiments containing  could  of  the  site  the  brief  tiations  i t was  concluded  200ym.m  i t could  f o r kainate  from  n o t be occurred  (1-2  kainate.  min,  were  1-2  used  In ml)  preliminary  to  perfusate  t o mimic the  of  the  neurons  drug;  duration i n  later  —6  -10  administrations (up t o 100%  potentiating  e x c i t a t i o n of CAl  applications  10-20 m l o f 10  positioned  level.  ^ M kainate  iontophoretic  and  reason  simu-  iontophoretic  at least  of action  bath-applied  —7  studies  extend  field  recorded  the  site  For this  exposures  10 ^-10  following  electrode  recording  of SC-evoked  with  brief  of  the depressing  the d e n d r i t i c or somatic  was  C A l were  barrel  a t each  enhancement  of  single-barreled  both  i n the  iontophoretically while  of i t sa p p l i c a t i o n .  An  addressed  pyramidale  the  a  was  the e f f e c t s of kainate  determined at  was  i n stratum  ltaneously  that  possibility  M s o l u t i o n s were p e r f u s e d . 10  ^  M  kainate  f o r a pre-treatment  produced  response  With poten-  o f 2-4  mV)  -5  which  lasted  f o r 10-20  PS a m p l i t u d e s w e r e 1.5  h)  larger  ( F i g .5.IB).  treatments  with  m i n ; i n 10  10  5  (50-1300%)  On M  kainate  of  the longer the  i n i t i a l  observed.  10-20  ml  perfused  exposures  A  of  PS  t o 10  and  10  and t h i s  -10 period  with  of  but  duration  especially  —6  -10  M kainate, of  solutions  of depressed  of  potentiation  additional  o f S C - e v o k e d P S s was  M  increases  the short  ( F i g . 5.11A)  recruitment  reduction - •} -4  solutions  and l o n g e r - l a s t i n g (up t o  occasion  —7  with  M  produced  the ammo  cell  PSs  firing  acid  was  was when  were  followed  137  by  a  long-lasting  potentiation  of  synaptic  excitation  (not  shown). Simultaneous potentials  revealed  recording that  of  the  dendritic  and  somatic  p o t e n t i a t i o n of the o r t h o d r o m i c a l l y -5  evoked  PS  associated  by  1-2  with  a  ml  perfusions  no change  of the presynaptic  the  amplitude  of AD-evoked  the  experiment  (Fig.  i n some  to  the d e n d r i t i c f i e l d  an  artefactual reduction  majority  at  Furthermore,  intensities  threshold decrease evident (Fig.  f o r PS of  there  this  detectable;  unchanged effect  were  t h e peak  of  throughout kainate  is  clear contributions  EPSP  PS w h i c h  amplitude,  d i d not appear  a decrease  generation  additional  kainate  occurring  was  bursts  which  (Fig.  o f t h e EPSP  continued  5.3C:4  i n the  measured  t o be  and  sub-  5 V ) ; and  and t h e r e v e r s e d  field  caused  to explain the  o f t h e EPSP was a l s o  observation  EPSP  a  was  potentials respectively  i n CAl  the transient (SPB) which  Prince,  1978).  resembled  I t i s likely  e p i l e p t i f o r m discharge  1981; and see below)  in slices  appearance  i n the p e n i c i l l i n - t r e a t e d  spontaneous al.,  of  amplitude  5.4).  population  and  slices  stimulation  the slope  same  were  o f F i g . 5.3.  i n d e n d r i t i c and somatic  An with  of  v o l l e y was  remained  curves  kainate  o f t h e EPSP  fibre  This  M  p o t e n t i a l of a reversed  of the s l i c e s  effect.  PSs  5.2).  i n the input/output Although  10  reversible reduction  but  shown  of  these  o f CA3  spontaneous  those  CAl region  that  of  perfused  reported  (Schwartzkroin  SPBs m i r r o r e d  neurons  and t h e consequent  as  the  (Lothman e t  transmission  of  138  15 min I 1  2 5 min  i  1  > 4 e.  o  J  > E  "•'-'.•W.v'  I min  Koinote 10 M  F i g . 5.2  Graphs of evoked responses  i n CAl from above  downwards p l o t the b a s e l i n e t o peak amplitude of the o r t h - and a n t i d r o m i c PSs recorded i n the stratum pyramidale and, from a second e l e c t r o d e p o s i t i o n e d EPSP the (10~ was  i n the stratum radiatum, the f i e l d  and the p r e s y n a p t i c f i b r e v o l l e y . fibre 5  volley  were  unchanged  M, 1 ml) whereas a 4 - f o l d  observed  depressed.  a t t h e peak  The a n t i d r o m i c PS and  by the p e r f u s i o n  of k a i n a t e  i n c r e a s e of the SC-evoked PS  of the e f f e c t  and t h e EPSP was  139  Population Spike EPSP A.  Control  (*)  B  Kainate  10" M 5  (°)  Stimulus  Fig.  5.3  In  antidromic  (Sg)  intensities  of  this  recorded  simultaneously  field  antidromic  stimulations. and  Responses 10  M  5  elicited  PS  i n the  (7,  8  V)  decreased fibre  of with  from  after  stimulation appearance  dendritic  potential.  the as  volley  amplitude compared  amplitudes  of to  The  the  of  PS  various  which  were  PS  was  this  orthodromic  of  1  their  reduction  stimulus  responses.  and  at  ml  of  maximum  decreased  increased  unaffected.  (A)  below ( C ) .  a positive-going higher  stimuli  responses  and  and  in A  perfusion  B  EPSP  At  control  were  in  )  sweeps  are plotted  the  and  the  at  electrodes;  of.control  (o).  (S  alternated with  illustrated  in C  CAl  orthodromic  strengths  min  in  superimposed  separate  were  stimulus  orthodromic  evoked  the  amplitudes  are  graphed  intensities  and  10-17  (V)  Three  responses  The  kainate  correlated  EPSP  additional  amplitudes  were  stimulation.  the  these  Intensity  experiment  responses  indicate  evoking  Antidromic  Orlhodromc  at a l l  was  not  (reversed) strengths  i t s latency  Antidromic  and  140  EPSP  6 v  Orthodromic  —  pre-treatment kainate treated (IO  4  4  5.4  Fig. and  pre-treatment elicited  (right)  Note  field  EPSPs  various response  Superimposed  a t t h e end  EPSP a t e a c h cell  body  EPSP  and  treatment shown.  line  with  the decrease  sweeps,  M,IOml)  traces  intensities  increase  records t h e peak  a  (left  o f SC  are  responses  The r e s p o n s e s  decline of the slope  o f t h e PS  amplitude.  M  6  kainate. of the  recorded the  of 3 consecutive  negativity  the post-treatment  at the  reversed  P r e - and  are comprised of  lines)  amplitude of  and  i n stratum  (dotted  10 m l p e r f u s i o n o f 10 and t h e peak  radiatum  stimulation  simultaneously  the  column)  i n the stratum  recorded  of the slope show  illustrate  recorded  stimulus strength.  layer an  of a  2mV  msec  Solid  the f i e l d  pyramidale. evoked  _ 6  v +  the  PS  post-  superimposed PS  i s not  141  impulses  i n t h e SC  observed  in slices  using  a  never Aside  input  from which  hand-held  produced from  evoked  microknife  with  this  from  in  those  since:  t h e CA3 and  had been  the actions  slices  slices  region  were  i n which  of  removed was  kainate.  of kainate  otherwise  t h e CA3  not  bursting  applications  activity  intact  (1) SPBs were  (2) s p o n t a n e o u s  iontophoretic  spontaneous  responses  uishable  to CAl c e l l s  on  indisting-  region  had  been  removed.  CA3 With NaCl)  increase  stratum  the  Montigny  and  t i p of  in  ( 5 mM,  precise  control  iontophoretic  pH  stratum min)  CA3a  since  CAl  i n 160 the  leakage  probably  into  these  to kainate  excitations  or  the  more  NaCl) were of  used  t h e amino  the  ( c f . de and  d i d not  dentate  dilute  by  related to  t h e same e l e c t r o d e s  region  mM  simply  electrode  was  with  i n CA3  mM  iontophoret-ic  neurons  cells  currents  the  over  these  of  i n 100  gyrus.  s o l u t i o n s of to allow acid  from  more the  electrode.  Kainate  (10-20  1981)  7.6  of  observation of  ( 5 0 mM  a l a r g e and l o n g - l a s t i n g  iontophoretic  i n the experiments  kainate  the  the  backing  either  barrels  activity  This  Tardif,  kainate  to produce  sensitivity  identical  Therefore  to  possible  pyramidale.  of  peripheral  spontaneous  greater  occur  the  i t was  of  lowering  using  concentrations  filling  electrodes  the  high  applied  i n o t o p h o r e t i c a l l y (5-20 nA,  pyramidale  increase  i n the  i n CA3a  produced  amplitude  of  MF-  10-60  sec)  a long-lasting ( F i g . 5. 5A  and  142  FIM-evoked  ( F i g . 5.13Ai)  sometimes  preceded  by  o r t h o d r o m i c P S s , an a c t i o n  a  period  of  depressed  -8 Perfusion tiated  1°  10-15  synaptic  stimuli was  of  10  -10  consistently  excitation  was  mediated  component  by  i n CA3  of  containing penicillin were  low  within  of  10-15  [CI ]  only min  a  5-10  intervals  epileptiform  PSs  the spontaneous  analyzed,  over  a  rather  onset  of  those [ K  , increased  few  Prince,  and  MF-evoked of  kainate  ( F i g . 5.6B)  than  evoked  similarity  a  the  change  in  small  and  size  often  ]  q  in  medium  (see Chapter  PSs  was  qualita-  2)  Initially  SPBs  However,  comprising  SPBs  c o n t i n u e d t o appear h.  Though  spontaneous  noted  or  potential  amplitudes. of  activity  between  elicited +  there  were  extracellular  of  1-4  epileptiform  of kainate  1978).  the  the next  d i s c h a r g e s was  FIM  ( c f .Chapter 2).  from  t h e mumber  and  ously  q  PSs  ( F i g . 5.6Biv,vi) which  infrequently  increased sec  doses  and  responses  Since the  bath a p p l i c a t i o n s  ( S c h w a r t z k r o i n and  observed  consisted  ( F i g . 5.5B)  with  input  the  indistinguishable  poten-  t o t h e enhancement o f o r t h o d r o m i c  response  SPBs  also  and o r t h o d r o m i c  increased  with  following  appearance  tively  b y MF  o f AD  t h e MF  of that  responsiveness.  kainate  solutions.  attributed  Coincidentally  the  M  n o c h a n g e o f t h e a m p l i t u d e o f AD  former e f f e c t  firing  -5x10  produced  M  was  —7  10  excitation  by  was  producing  t h e AD  of  ( F i g . 5.6A); a d e c l i n e —6 —5  caused PS  ml  which  not and  at  rigourevoked  (e.g. F i g . 5 . 6 B i i i -  vi) . Large latencies  of  (up  to  evoked  100  msec)  responses,  and  variable  particularly  shifts with  low  of  the  inten-  143  Fig. recorded  5.5  from  positioned  A.  ( i ) The  t h e c e n t r a l b a r r e l o f an  i n CA3a.  sec) caused  an  immediate  the  positive  wave  which  of  potentation  the  of  the  observed  (iii).  elicited  by  respectively)  1° 1°  B.  MF  PS  and  10,  the  of  of  A  firing  of  the  somatic  25  and  65  of  min  a  10  (5 nA f o r  the  15  of  beginning  electrode  1°  PS  and  kainate-resistant  i s evident. the  response  kainate  both  followed.  Changes  stimuli  after  reduction  response  MF-evoked  iontophoretic  ( i i ) Iontophoresis  10  component  pre-treatment  a  min  later  second  PS  field  a was  potential  ( i i , i i iand i v ml  perfusion  with  —8 medium the  containing  i n i t i a l  depression records  PS of  10  and  evoked  are comprised  M the  kainate.  Note  development  activity o f 2-3  was  of  not  consecutive  the  potentiation  additional  observed.  In  superimposed  of  PSs.  A  A  B  and  traces.  144  Fig. evoked  The  activity  kainate and  5.6  i n CA3a.  orthodromic  PSs  a d d i t i o n a l PSs  SPBs  (e.g. B[iv])  evoked FIM  bursts.  stimuli  teral  stimuli.  which  Partial but  PS  a similarity was  after CA3. 2+  of  spontaneous elicited were  by  [Mg  after  number  of  the  with  iontophoresis.  latency  of  responses  to  discharges to  (cf.  Schaffer  the  C.  of  treatment  PSs  2  PSs  h  later  (vi) field activity  was  observed  stratum  (15 ml  pyramidale of -7 o f 10 M) i n l o w  t o n o r m a l ACSF c a u s e d  the onset  indistinguishable  E. W i t h i n  of  SPBs ( i v ) .  occurred  multiple  the  i n evoked  Spontaneous  i n the  colla-  amplitude  onset  had  Fig.  d i d not change  and  20 m i n  spontaneously-occurring o f PSs  of  (v) and s p o n t a n e o u s  than  were  onset  o f m u l t i p l e PSs  responses  kainate  increased  i n the  of kainate  the  rather  which  of bursts  were  With  response  evident.  kainate  by  SPB  and  a p p l i c a t i o n of  stimuli  shifts  appearance  ] a return  PSs  replaced  comprised  sec  evoked  s t i l l  Following 2+  ]/high  4  correlated  single  spontaneous  bath  ( i i ) .  large  between evoked  iontophoresis D.  were  The  of  FIM  recruited  increased  was  recovery  comprised  by  Perfusion  ( i i ) .  (iii)  potentials  [Ca  but  on  Following  ( i ) Pre-treatment  followed  activity  A.  kainate  i n ( i i i ) are 3 consecutive  delivered B.  antidromic  were  Shown  of  evoked  there  6.5-6.7). (SC)  effects  these field  from  of  those  single  PSs  potentials  (see t e x t f o r d i s c u s s i o n ) .  145  Koinote 2 5 x l O " ' M . lOml •  . . I mV(C.D) , I. 2 - IE) 10 msec ( C . E ) 20 • (D)  146  sities  o f FIM  SPBs  stimulation,  ( F i g .5 . 6 A i i i ) .  changes  of  response  chapter  in  relation  were  The  correlated  factors  latencies to  a  with  the onset  contributing  are  discussed  similar  action  to  of  these  i n the  next  folate  and  of  bicuculline. Field stimuli  potentials  were  sometimes e v i d e n t  particulary  when  However  this  produced  with  bath  short-lived  and  was mV  large  baseline  t o peak  was  was  a  iontophoresis employed.  different  at  acid  10-50  PSs o f v a r i a b l e  associated  electrical  were  o f t h e amino  negativity)  of synaptic  Since implicated  applications  of  kainate  currents  activity  of single  activity  potentiation  following  (2-5 m i n ) , o c c u r r e d  comprised  taneous  i n the absence  ejecting  spontaneous  was  5  occurring  that  i n that i t  msec  intervals  amplitudes  ( F i g .5.6C). with  from  (up t o  This  but outlasted  sponby t h e  excitation.  recurrent  i n the mechanism  excitatory underlying  circuitry SPBs  has  been  i n t h e CA3  region —8  (cf.  Chapter  2)  the effects  of  kainate  ( c a . 15 m l o f 10  -7 10 M) w e r e e x a m i n e d i n s l i c e s w h e r e s y n a p t i c transmission 2+ 2+ had b e e n b l o c k e d u s i n g l o w [Ca ] / i n c r e a s e d [Mg ] . I n each Q  of  3  slices  discharges response within  did  was 5-10  activity  treated  in this  not  min  of  comprised  those  recorded  (Fig.  5.6D).  after  fashion  occur  comprised  of a  and single  returning of  q  single  the PS  PSs  and  epileptiform  antidromically ( n o t shown).  t o normal  iontophoretic  As r e p l e n i s h m e n t  spontaneous  medium  evoked However,  spontaneous  indistinguishable  administration  o f n o r m a l medium  was  from  observed  continued  this  147  type  of  (Fig.  spontaneous  5.6E),  opment  an  effect  of a n t i -  Dentate  activity  and  which  doses  comparable  granulosum  stratum  or  stimulation LPP  of  and  latency  of  recorded  the medial  were PS  or  with  burst  by  the  SPBs devel-  firing.  potentiated  ( F i g . 5.16A). was  no  o f AD  the  input/output  evoked  were  (MPP  i n the  than  brief  evoked  by  relatively  reductions  for a  PSs  stratum  pathways  shorter-lasting Except  region  the  t h e PS  perforant  large  were  change  to  potentiations with  but  i n the CAl  either  lateral  These  discharge  there  by  those  i n i t i a l  MF  stimuli  shown). Shifts  following  the  of  bath  associated,  applications  again  middle  as  molecular (Fig.  particularly  striking  M  example,  solutions  V)  stimulations  an  EPSP  was  potentials. intensities  of  The was  in  the  an  the  of  4  elicited  of  of  10~  were  synaptic  kainate  where  10-15  were  8-10  MPP  was ml  perfused. low mV  occasional  secondary  PS  observation  at  of For  intensity  PS  even  high  (5  though  i n the d e n d r i t i c nor somatic a  PS  recorded  of  treatment an  kainate  of  action slices  M  5  o f EPSPs  region  acid  kainate  neither  f o r t h e MPP-evoked  reductions  amino  after  firing  ml  This  each  t h e MPP  evident  1-2  layer,  5.7A).  of  i n F i g . 5.8  of  curve  i n CAl, with  terminations  ^  replaced  required  kainate  associated  i n CAl  depression  of  moleculare  respectively).  greater  10  associated  to those  applications  in  was  orthodromically-evoked  iontophoretic  (not  gradually  Gyrus In  and  was  field  stimulus  ( F i g . 5.9:30  V).  148  18 -i  > 6 E  «  |,6J  5  "O  2  cu 14 H "O  4  1  ? 3  o  '2  Q.  o. to  2  10  00 0_  2  H  o  UJ 0  1  -1—  —I  I  30  20  10  —i 40  i 30  I  20  10  40  Stimulus  >  £  Intensity (V)  12 i  20  10  "a  V t  AD  8  Z>  6-  Q.  E <  30 V  4-  Q_  »•  2-  a  AD 5  10  15  20  25  J*4mV 4 msec  30  Stimulus Intensity (V)  Fig.  5.7  successive symbols)  relating  responses  10  kainate  M  measured  at  dendritic shift  stratum  and  of  ml  of  amplitude  of  apparent  (•)  precluded a  any  for  by  after  the  (o)  10  of  M  5  the  the  and  the  10  and third  had was  late  kainate.  curves  column)  there  and  been an  ml  PS  increase wave,  perfusion.  EPSP  both an  in  10  indicate 10  duration the  effect  peak  (•) from  control  and  20  ml  After  20  of  the  duration  which  of  ml  records  after  of  to  the  intensities  respectively). the  the  large  recorded  filmed  of  were  of  the  p e r f u s i o n of  obtained  perfused  of  3  of  2 ml  baseline  derived  columns  positive  amplitude  Note  various  The  were  those  PS.  slice,  of  than  contamination  the  evoked  before  and  p e r f u s i o n of  reversed  curve  mean  smaller  Amplitudes  PSs  of  after  after  antidromic  kainate and  stimuli  another  (second  increased  MPP  gyrus.  by  point  deviations  In  (first  10  (each  B.  (•)  perfusions -5 ml  potential  portions  response  and  dentate  granulosum.  20  (o)  input/output  stimulation  which  the  of  l a t e n c y which  the  amplitudes MF  a  intensity  in  curves  standard  before  field  of  Average  responses,  evoked 5  A.  was  PS and also  *  149  That  in  3/3  treatment [K ] +  Q  of  the  with  10-15  could  not  (Fig.  5.8)  tudes  to  In  connection except  a  observed  effect  of  kainate  5.7B).  the  of  either expel  was  to  or  by  brief  on  subsequently  potentiated  periods  of  15  When  39  min  applied  and  15  EPSP  and  PS  ampli-  both  the  not  cells.  bath-applied a  depressant  amplitude  followed of  to normal  the  AD  PS  kainate  and (Fig.  usually  ACSF.  CAl  excitatory  currents  where the  by  mM  were  granule  where  actions  periods  slices  or  which  in  other  investigated  10  I n t e r e s t i n g l y , an a d d i t i o n a l  returning  of  5,  dentate  exposures  enhance  amino a c i d s  subsequent  kainate's , actions  these  iontophoretic  kainate  of  or  were a l s o u n a f f e c t e d  from  actions  with  that  p o s i t i v e wave  other  The  changes  prolonged  w i t h i n 1 h of  Effects  containing  ( F i g . 5.7B).  Recovery  occurred  ACSF  similar  PSs  with  was  of  of  previous  d e p o l a r i z a t i o n of  AD  action  duration  slices  suggested  simply  kainate  ml  cause  related this  latter  amino  similar (1-3  to  min)  kainate  SC-evoked  acids those  of  by  used  perfusion  applied PS  applied  at  were  previously least  20%  NMA a  layer  produced  by  iontophoresis  long-lasting  both 5.9C).  in  the The  ibotenate  ( F i g . 4.7A)  only  a brief  depression  stratum  or for  longer. in  the  cell  body -5  dendritic  to  or or  i n the  no of  perfusate  increase  i n the  a l l evoked  pyramidale  and  (5x10 PS  stratum  -4 -10  followed  responses radiatum  or  M) by  recorded (Fig.  depressant a c t i o n o f NMA was m i m i c k e d b y perfused -5 -4 (5x10 -10 M) . As n o t e d p r e v i o u s l y ( c f . C h a p t e r  150  EPSP Pre-treatment  Recovery  I 0 " M Koinote 5  10 m M  K *  J 4 mV  )  4 msec  4 msec  Fig. tive  5.8  superimposed  somatic  field  Filmed  records  each  sweeps  showing  the  responses  perfusion  of  medium  increased  [K ]  the  containing  (15  +  in  ml).  comprised changes  5  Treatment  M  consecu-  gyrus  kainate  with  3  in dendritic  dentate  10  of  and  following (10  kainate  ml)  or  caused  a  o reversible enhanced  depression  synaptic  containing  10  either  EPSP  pressed  the  both  mM  EPSPs  the  excitation. [  or  of  K  +  ]  the and  Q  did PS. PSs  not In (not  dendritic  EPSP  Subsequent  exposure  produce the  same  shown).  and  comparable slice  15  mM  markedly to  medium  changes  of  [K ]  de-  +  o  151  4)  the  amplitude  recovery  of  the  EPSP  from NMA-induced  Quisqualate  was  often  potentiated  following  depressions.  applied  by  iontophoresis  or  perfusion  - 5 - 4 (10  -10  ally  preceded  (Fig.  M)  generally by  5.9C).  recovery  a  depressed  short-lived  although  this  duration  than  was  was  both  that  the  higher  commonly  smaller  elicited  PS,  an  effect  enhancement of  Especially with  from depression  the  by  in  the  doses  by  amplitude  kainate  i n the  SC-evoked  of  followed  occasionPS  quisqualate potentiation  and  shorter  same  in  slice.  -3  Both only  L-glutamate  short-lasting  and  changes  L-aspartate  i n the  PS  at  10  amplitude  M  produced  ( F i g . 5.9A);  at  -2 10  M  the  (100-400%, (Fig.  response  20-80  sec)  5. 9 B ) .  was  and  initially  then  was  Potentiation  briefly  depressed  was  not  potentiated  for  long  observed  periods  following  recovery. Kainate's  mechanism(s) of  It  has  Dingledine penicillin 1983)  been  and and  which  reported  that  bicuculline  (BIC)  (Wigstrom  reduce  epileptiform  discharge  activation applications hippocampal  GABAergic  of  produced  inhibitions kainate  region  by  kainate,  following were  using  the  examined paired  such  the  bath  as  Gustaffson,  can  produce  responses Since  1980;  and  inhibition  neurons.  Prince,  compounds  orthodromically-evoked  also  of  and  1980)  of  are  (Schwartzkroin  Gjerstad,  potentiation  actions  action  these  and  or  in  two post-  iontophoretic  extracellularly stimulus  cause  latter  changes  a  in  paradigm  each (cf.  152  LGLU 0 ' M 3  L ASP  0°M  > E  L-GLU O M  '.-'"v'^-V.  Ou.s  lO'^M  > E  lO'-M  C M  S3  ...v^r--  Fig.  5.9  A. I n t h e C A l r e g i o n  perfusion  f o r 1...min  with  -3  10  M s o l u t i o n s of L-glutamate  produced which  brief  increases  d i dnot o u t l a s t  (L-glu)  or L-aspartate  i n theamplitude the period  (L-asp)  o f t h e S C - e v o k e d PS  of application.  B. L -  -2  glutamate followed another  at by a  slice  long-lasting mimicked ca.  10  lengthy  elicited depression  a  short-lived 10  potentiation of synaptic  by a s i m i l a r  dose  orthodromic  treatment  activity.  PSs.  M kainate  5  excitation  of quisqualate  Subsequent  potentiation  of orthodromic  a 1 min perfusion with  40 m i n l a t e r .  depressed  M  (Quis)  with  C. I n  produced a  w h i c h was n o t administered  NMA ( l o w e r  graph)  153  Chapter  2)  as  w e l l as  by  intracellular  E x t r a c e l l u l a r a n a l y s i s i n CAl of r e c u r r e n t i n h i b i t i o n In  the  conditioning  of  preliminary  AD  and  test  analysis in  kainate-induced  experiments  orthodromic  the  stimuli  CAl.  changes intensities  remained  of  constant  _5 as  kainate  slices  was  examined  responses  in  was  inhibition  this  associated  amplitude  of  unlikely  that  this  to  the  inhibition  to  affect  Fig.  (t=20-25  min)  amplitude  of  s t i l l  potentiated. In  were  6  other  generated  by  a  that  constant  ( c f . Lynch  unconditioned  of  PS  larger  the  (25  PS  a  in  of  a  PS  of  the  stimuli  constant.  recovery  antidromic  of  It  PS  the  of  the  as  6.9a).  curves"  remained  conditions a  of  smaller  amplitude  After  was  orthodromic  revealed the  since  PS  activations these  of  inhibition  "inhibition  slices  conditioned  below)  is was  strength  orthodromic  Under  control  (see  3  SC-evoked  AD  constant  intensity  (Fig.  of  transient disinhibition  pre-treatment  increased  each  loss  msec)  unconditioned  the  In  a  remained  et_ a _ l . , 1 9 8 1 ) .  the  with  amplitude  shows  for  responses  decrease  AD  capacity  varying  while  percent  the  M).  p o t e n t i a t i o n of  preceding  slices  stimuli  stimulation  the  10  initially  limited  the  of  kainate-induced  simply  while  ml  manner  by  the  5.10  (1-2  activated  although  due  perfused  of  the  perfusion  of  —6 5-10  ml  of  10  M  was  associated  kainate with  inhibition  ( F i g . 6.9c),  appearance  of  Fig.  5.11A).  a  p o t e n t i a t i o n of transient  an  However  in  abolition  a c t i o n which  a d d i t i o n a l PSs  i n the  each  slice  synaptic  was  recurrent  paralleled  orthodromic the  of  excitation  by  response  inhibition  the (cf.  returned  154  (b)  (o)  (c)  (e)  (d)  v. 1.2  10 msec  IV.  (a)  f^—vyv_  (c)  (b)  n  n  10  mV  (e)  (d)  n  n  >  2-  Kainate IO" M, I ml 5  10  + 5  20  25  45  50  —i  60  65  Time (min)  Fig.  5.10  i n h i b ifc'io n response  in  was  The  uninhibited indicates  the  of  10  graph  (filled  a- b r i e f ^  appearance  PS  .In  M  potential  remained  of  period  (not  superimposed  the  peak  photographed. the  now  show  conditioning  row);  of  was  the  At  antidromic  PS.  3  in  in  absence  the of  any  1 of  transient  orthodromic  at  of  the (top  i n the  the (3  times  row)  PSs  responses  change  loss  records  orthodromic  field  of  unconditioned  filmed  obtained  test  PSs  reduction  unconditioned responses and  circles)  the  the  the  substantial The  the  application  with  an  of  apparent  min  by  inhibitory  amplitudes  an  test  msec)  (open  mV)  t=20-25 a  (20  bath  was  to  sweeps)  positivities the  -the  evident.  antidromic  Note  peak  correlated  (up  whereas  consecutive  the  PS  recurrent  recurrent  to  there  the  orthodromic  inhibited  was  shown).  PS  (a-e)  and  effect  potentiated  the  baseline  which  secondary  an  :  following  in  on  o c c a s i o n s •;preee!ded  the  conditioned  indicated  ;  circles)  This a  kainate  activating  kainate  of  of  t.he e x p e r i.m.ent  alternate  inhibition.  field  actions  stimulus  pathway.  ml  CAl.  on  antidromic  The  and  (lower  were  not  amplitude  155  far of  in  advance  kainate  was  of  recurrent  with  no  In  from  2  the  slices  change  or  potentiating effects  (not  an  shown) p o t e n t i a t i o n  apparent  increase  of  the  inhibition.  Intracellular o f k a i n a t e on  a n a l y s i s of the a c t i o n s CAl pyramidal cells  Stable  intracellular  obtained  activation passive  recovery  ( F i g . 6.9d).  associated  were  the  from  from  and  16  the  impalements  CAl  pyramidal  alveus.  In  a c t i v e membrane  m e d i u m was  switched  to  one  held  cells  the  f o r 30  -  identified  experiments  p r o p e r t i e s were containing  min  h  by  AD  changes  of  determined  kainate;  3  as  usually  the  1-2  ml  -5 of  a  10  were the  M  s o l u t i o n were  obtained stratum  Mean  (+S.E.M.) v a l u e s input  potential similar  (AP)  to  Dingledine of  longer)  was  ization  of i t  decreased or  was by  Except  RMP  by  PS  APs  in  kainate  was  those  (24jf2 mV)  in  others  R  positioned  the  impaled  and  mV, was  I N  normal  Prince,  and  action  either No  In  Poten-  30  small  were 1975;  1980).  a  (RMP)  medium  lasting  with  n=13).  (n=8).  megohms)  in  cell.  (Schwartzkroin,  intracellularly  4.8+1.0 megohms of  I N  (100-1000%  (4.3+0.3  measured  )  R  Hotson  SC-evoked  the  lOO^utm o f  (88 + 1  1980;  correlated  amplitude  removed,  a_l. ,  electrode  recordings  r e s t i n g membrane p o t e n t i a l (  reported  Extracellular an  ca.  f o r the  resistance  those  the  with  within  amplitudes  et  tiation  which  simultaneously  pyramidale  (-64+lmV),  perfused.  min  or  depolar-  those  cells  in  unchanged  ( n = 4)  or  change  i n the  duration  detectable. slices  initially  where  caused  the the  CA3  region  transient  had  been  appearance  156  (1-3  min)  20-100  of  spontaneous  msec  which  synchronized random  with  discharge  as of  APs  manner mV  R  of was  recover  synaptic  epileptiform  7  other  in  control  6  was  held  of  for  of  of  the  premV  than  in  mV  the  than 1  h  a  over  and  post-  the  to  same  8.9+1.2  reduction the  recordings  more  on  respectively  effect,  in  intra-  unchanged  measured  Although  more  intensity  subthreshold  13.0+1.5  suggested  for  were  EPSP  disproportionately  cells  or  SC-evoked  11.1+2.5  the  part  1  superimposed  cells  and  kainate.  factors  or  values  mV  an  the  RMP  values  explained  at  of  at  cells  with  have  other  pulse  discharged  previously  amplitudes  10.9+2.4  from  PSs)  was  either  cells  the  contri-  where  the  R  of  the  (Fig.  J N  EPSP  could  fully. A  and  In  In  and These  reflected  5/13  multiple  experiment:  were  reduced  5.11B).  of  peak  the  perfusion may  bution EPSP  of  declined  I N  potentials.  kainate-induced  which  measured  EPSPs  after  the  The  5.11A).  field  kainate  with  hyperpolarizing  treatment  hyperpolarization  likely  the  with  stimulation  EPSPs  course  (Fig.  of  lasting  cells.  5.11A).  mV  the  result  (associated  cellular  most  depolarizations by  extracellular  treatment  orthodromic (Fig.  a  CA3  After  5-10  enhanced  depolarizations  activation  more  were  membrane  the  more  IPSP  evoked  measured  stimulus  consistent  at  artefact  hyperpolarizing  action  either  the  RMP  (Figs. IPSPs  by at  kainate  orthoa  5.11A, were  of  or  5.12A,B). by  the  antidromic  latency  reduced  was  of  30 Thus  reduction stimulation  msec in  79+6.3%  from  13/13 from  the cells  a  pre-  .15 7  Fig. cellular  5.11  responses  superimposed (lower).  The  of  a  on  a  120  with of  the IPSP.  across  t h e membrane. enhanced  2  input mV.  the  B.  current EPSP  decreased. later.  2  ml  the PS  The At  although current  Lower on  kainate  action  intracellular ( t = 7 m i n ) was  t=37  a  nA)  poten-  was  correlated  EPSP  (t=5 m i n ) . intra-  10 m i n  substantially  reduced of  lower "seen"  excitation The  was graph  t o the magnitude  potential the  being  recovered.  potential  show  i n the  fields  indicates that  recovery  and  (0.8  correlated  synaptic  had  traces).  Partial  (middle)  i n t h e EPSP  min  IPSP  membrane pulses  intra-  o f two a c t i o n p o t e n t i a l s and t h e notches  the  traces  and  caused  which  kainate  after but R  t h e EPSP  by  perfusion I N  was was  only  (arrow)  produced  of  decreased  d e c l i n e d by  SC-evoked  a hyperpolarizing pulse  (upper  was  of  an  resistance; the resting  superimposed of  C£. of  potential  to the large e x t r a c e l l u l a r  steady-state  intracellular the  resting  ( t o p row)  hyperpolarizing pulse  the discharge  a r e due  relating  of  of a second  records s t i l l  at  msec  reduction  appearance  abolition  extracellular  t h e SC-evoked PS,  small  c e l l u l a r ^  The  evoked  Perfusion  tiation with  A.  EPSP  injection of  only evident  kainate slightly 20  min  158  Control  A.  Koinale (IO~ M.2 mm. 1 = 0 mm) 5  1=5  37  i  L  40 msec  40 mvf 4 0 msec nA  -50  Kainote o 2 min  -60  -i.o  - 7 0 mV -80 L  B.  Control  Kamate ( I 0 " M , 2 mm, t = 0 ) 5  10 nA  *J  20  -90  159  Control  Kainate ( I 0 " M . 150 sec. t = 0 mm 5  t =7  A  13  18  4mV [ +  40mV  [  Kainate ( I 0 ~ M , 120 sec. .t=0 mm ) 5  t=l7  24  B 40mV [ +  100 msec  Fig.  5.12  intracellular region. after a  A.  responses  recorded  In the c o n t r o l  situation  the beginning  hyperpolarizing  of a  of kainate  same  the  time  inhibition and  Recovery continued but  with  reversible  of  IPSP  7  min  SC  t h e PS was  and  cell  of  at  t=18  of  synaptic  antidromic reduction cell  IPSPs  firing.  o f 10 c o n s e c u t i v e  there  cell  min  was  and In  the  was  associated B.  kainate  their  inhibitory  A  B  and  superimposed  filmed  sweeps.  bath  At the enhanced  t = 13 was  msec  of  an  . At  CAl  elicited  beginning  firing  Again  and  d e l i v e r e d 60  excitation.  stimuli. of  the  firing.  inhibit  potentiation  in  greatly potentiated.  to  IPSP  stimuli  after  persisted  records)  depolarizing pulse  i t s ability the  (upper  simultaneously  msec  current-induced  current-induced comprised  200  IPSP.  application  IPSP  Extracellular  min  the  reduced. with As  the  for A  caused actions records  a on are  160  treatment greater  value than  suggested not  a  could  by  of current  when s y n a p t i c onset  cells  o f t h e IPSP was  induced  cell  however w i t h  also  (see above).  The  analyses  firing  inhibi-  ( F i g . 5.12A).  The  o f 2 o r more  A P s was  ( F i g . 5.11A), with  paired  of kainate-treated CAl  of depolarizing current  of trains  poten-  time  i n the experiments  t o 200-300 msec p u l s e s  an  was  at a  o f t h e IPSP  response  A s was  and  persist  enhanced  «  there  producing  could  a depression  reached  be t h e d i s c h a r g e  IPSP  firing  was s t i l l  I N  disinhibitory  a prominent  excitation  conclusion  stimuli  to  Thus  kainate's  of orthodromically-evoked  correlated a  of e x t r a c e l l u l a r  between  actions.  The d e p r e s s i o n  be a t t r i b u t e d t o t h e c h a n g e o f R  the results  parallelism  tiating tion  o f 5.9+0.8 mV.  o f s i n g l e APs ( F i g .  continued  5.12A,B).  CA3 In  addition  inhibition a  late  (cf.  there  onset  Chapter  described ularly  to  phoretic  3).  Using 3  20  o r bath  and  t h e same  inhibition  200-300  3  min)  potentiation effect  to of  on t h e i n h i b i t i o n  which has  stimulating of  the  C-T  procedures and  FIMQ  intervals  t o BIC  particPS  after  as  were  ionto-  of kainate.  i o n t o p h o r e t i c a l l y ( 2 . 5 - 1 0 nA f o r u p  the stratum FIM-evoked  an i n h i b i t i o n  of FIM^ orthodromic  msec  administrations  BIC-sensitive  and i s i n s e n s i t i v e  the changes  When i t w a s a p p l i e d to  GABA-mediated  but long-lasting effect  i n Chapter  at  early  i s i n t h e CA3 r e g i o n  t h e MF^-evoked  assessed  an  pyramidale orthodromic  occurring  i n doses  producing  PSs, kainate  20 m s e c  after  MF  had  no  stimuli;  161  pre-,  post-treatment  inhibition and  (mean  72.5+11%  early  (n=8).  may  of  concurrently of  71.6+4.2%  reduced (Fig.  to  the the  5.13Aiii).  within  15-25  actions  of  slices,  Recovery  min  of  kainate  the  were  the  a  small  the  late  termination  not  when  C-T  mimicked  by  the this  in  the  measured inhibition  interval  was  administration  inhibition  of  of  increase  msec  (n=9)  however  pre-treatment  300  early  increase  following kainate of  the  80.7+9%  contrast,  the  at  (n=ll)  (n=9),  treatment,  In  measured  3.5+3%  to  PS.  same  (n=ll)  only  1°  for  i n d i c a t e s an  kainate  related  MF^  values  72.7+6%  F i g . 5.13Aii  been  in  were  following  have  amplitude  recovery  +S.E.M.)  inhibition  effect  and  occurred  treatment.  current  These  controls (Fig. —8  5.13B).  In  virtually FIM  2  process  inhibition  could  synaptic  the  late  only  GABAergic  indicates be  The inhibitions  that  produced  the  the  reduction  independently  suggesting  the  between  two  of  the  ( F i g . 3.4A,B)  unconditioned  (2-5x10  produced  reducing  excitation  event  kainate  inhibition  slightly  s i t u a t i o n s where  inhibition  bath-applied  M)  with  paired  early  GABA-  ( F i g . 5.14A).  5.14B  relationship to  the  while  Fig.  of  slices  abolished  stimuli  mediated  other  was  not  response  e f f e c t s of i n the  CA3  actions  any  absence  reduced  the  e l i m i n a t i o n of  ( F i g . 5.13B; and  region  with  of  a  In  early  burst  late  the  causal contrast  GABAergic late  firing  in  nonthe  5.14A).  kainate are  the  the  enhancement  kainate.  BIC  correlated  BIC  of  of  on  the  summarized  early  in Fig.  and 5.15.  late  162  Fig.  5.13  input/output CA3a  and  curves  20  msec  stimuli.  late  with  tiation  of  (a)  recovery the  was  changes  inhibition  kainate  Kainate producing 2.5  msec  nA  current  the  iontophoretic  responses curve  in ( i ) .  increased  ( [•]  (iii).  early  (25 msec)  reversibly  nA c u r r e n t  carried  reduction by C I  a  an  the  a  shift  min  B.  Filmed  and  late  the  the  late  was  records (225  from  3.4.  6 to 5 V PS  of  an  situation. while  inhibition.  d i d not mimic kainate's  of  kainate  inhibition  of the early  a  msec)  as f o r F i g .  unconditioned late  poten-  time  application of  decreased  of  to the  there  i n the pre-treatment  abolished  small  25  Conventions  t o evoke  was  i n i i ) whereas  values.  to that  as  ejection  A t t h e same  After  l a y e r o f CA3a.  treatment  there  revealed  o f F I M s t i m u l a t i o n was  a  explanation)  ( i i i ) after "conditioning  f o r 3 min  to pre-treatment  comparable only  f o r  the  abolished  body  intensity  amplitude  300  following the iontophoretic  the c e l l  after  (see text  response i n  Following  was  i n  orthodromic  amplitude  (o) r e v e a l comparable e a r l y and  the evoked  inhibition  The  10  inhibition  intensity/PS  responses  of the input/output  early  in  a  curves  ( i i ) and  Control  kainate  stimulus  f o r the FIM-evoked  inhibitions.  left  ( i ) The  the inhibition  generated MF  A.  A  actions.  164  Fig. 20  msec  paired  5.14  (centre FIM  17 m l ) . actions  To a v o i d of  test the  which  PSs  was was  stimulation  depressed  at  parameters 300  msec  (t=30  produced  min) and  inhibition.  larger  a  similar  caused  dose a  a  MF  M, used  on  of  and  kainate  late  normal  of kainate  caused  orthodromic  PS  the i n h i b i t i o n After  i n record) dose  the  with  stimuli.  C  response of  was  of  wash-out  FIM-evoked  effect  (break  the unconditioned A  by  (2x10  (15 V)  reduction  after  no  with  iontophoretic applications  the  having  column)  by t h e p o t e n t i a t i n g  iontophoresis  of  (20 msec)  inhibition. activity  slice  produced at  of kainate  stimulus  selective  reversible  while  preceded  inhibition  a  (right  introduced  As w i t h  potentiation  circles)  msec  supramaximal  In another  immediate  (filled  a  there  B.  300  complications  the experiment.  kainate  medium.  and  following perfusion  kainate  inhibition an  column)  stimuli  throughout of  A. T h e c h a n g e s o f t h e i n h i b i t i o n  to  changing  assess  kainate  decreased  applied  substantial .depression  longer-lasting depression  min of  of  the  briefly the  15  of  late later  evoked  the  late  165  166  Pretreatment  BIC treated  Recovery  100 (10) (10)  JL  (3)  50  (10)  LU  0  A  L  20  200-300  . 20  200-300  20  200-300  c o  OJ  E  Pretreatment  100  o  Kainate treated  Recovery  (9)  la  19>  (8)  1  (II)  _c  JL  (©)  20  300  50  (ill) 0  L  20  Fig. and  kainate  dromic uli  5.15  PSs  20  Summary  (bottom) elicited  (see text  group  300  on 20  of  the e f f e c t s  the  inhibition  and  200-300 msec  for explanation).  i s indicated  300  i n brackets  The  above  of b i c u c u l l i n e of  FIM-evoked  following  number  of  the e r r o r  a MF  slices bars.  (top) ortho-  C  stim-  i n each  167  Dentate  Gyrus There  by  kainate  intervals 1-2 in  was of  with  the  inhibition  Damiano,  MPP  same  A  the systemic 1981b).  synaptic  had  of a  20  orthodromic was  PS  of  Approximately  PSs  i n the  events  3 min  continued and  5.16C  kainate but  produced  a  whereas  pre-treatment  i s seen  later  while  was  of  (Fig. 5.16Cii)  observed  C-T  intensity  administration of kainate  recovered  msec  i n Fig.  reduced  conditioning  sequence  excitation  inhibition  at  depression  of the iontophoresis  amplitude  similar  brief  For example,  stimuli  activity  for a  occurring  stimuli.  granulosum  of t e s t  following  inhibition  of the t e s t  situation.  in 3 slices  the termination  the stratum  potentiation  of  the paired  min a f t e r  evoking  evidence  iri v i v o  (Sloviter  and  the potentiation  the p o s t a c t i v a t i o n  i n fact  increased ( F i g .  5.16Ciii) . In  4  slices  ically-applied clear  the potentiating  kainate  were  disinhibitory  increase  of  of  iontophoret-  not c o r r e l a t e d a t any time  action.  MPP-evoked  actions  cell  Similarly firing  the  produced  with  a  substantial by  10-15  ml  -5 perfusions  of  persistence  of the i n h i b i t i o n  msec BIC  C-T  intervals  (10 ^  firing  10  M)  kainate  the  were  of t e s t  ( F i g . 5.16Bi).  abolished  i n both  5.16Bii).  M  responses Subsequent  inhibition  the conditioning  correlated  and  and test  with  elicited treatment  also  caused  the a t 20 with burst  responses ( F i g .  168  Fig. stratum  5.16  granulosum  stimulation body  activity -5 M  B.  at  14  PS  Addition  reveal  a  lower a  intensity  shorter  C.  of  a  In  min  of  PS;  6-7  p o t e n t i a t i o n of  later  stimulus) large test  but  field  recovery comprised  test  inhibition  depression  of  the  of  of  5x10  (C)  of  of  activity  test  kainate  test  was  V)  had  of  had  occurred.  superimposed  sweeps.  ml  of  test  EPSP.  the  in  ( i i )  and  caused  MPP  stimuli  the  control ( i i ) a a  PS  the  ( i i i ) (11  recovered. EPSP  at  to  continued  min  of  kainate-  evoked  reversed  ( i v ) 25  (T)  of  potentiated.  activity  evident.  10  i n amplitude  c o n d i t i o n i n g PSs  cell  discharge  paired  (7  comparable  test  of  iontophoresis  stimulation  actions  and  inhibition  slice  peak a m p l i t u d e  3 consecutive  to  (MPP)  the  the  the  path  in  reversed M)  pulse  in  inhibition  ^  response  potential is also  from kainate's  kainate  another  but  pre-treatment min  of  detectable  after  onset  the  perforant  after perfusion  paired  conditioning  latency  medial  recorded  conditioning  (BIC,  inhibition 1-2  PS  conditioning response  absence  firing.  ( i ) .  the  persistence  bicuculline  potent  situation  a  medium a b o l i s h e d  epileptiform  to  interval  i n the  i n the  of  iontophoresis  showing  Note of  containing  response  m s e c C-T  kainate  mV  in  ( i ) MPP-evoked  a 20  responses. a  Potentiation  following  layer.  10  A.  in  later  A l l records  V A the  full are  169  170  Discussion Although hippocampal  regions  physiological in  low  kainic  changes  PSs.  At  activity  was  perfusions  dentate  reported  a  -6  reduced  in  -5  M  was  i n CA3  sustained  cells  kainate,  seems  region this  with by  effects likely  was  due  regard  evoked  10  that also  the  respones  the  on  dentate  to  lower in  greater  and an  The  responses  following  related  an  to  neurons of  There disinhibiting trations  of  were and  evoked  10-15  ml  i n CAl  and  (1981)  of  CA3  have CAl  concentrations  of  is  evoked  analysis  not  dose vis the  available,  activity  in  In of  the  depression  vis  CAl  is  CA3  neurons  (de  of  AD  iontophoresis  consistent  to e x c i t a t i o n  Montigny  both  this  neurons.  for  a  of  and  Tardif,  and  synaptic  were  probably  depolarization.  several  observations  potentiating  kainate  large  and  cells  of  depressions kainate  excessive  M  d e p o l a r i z a t i o n of  threshold  sensitivity  transient  -4  intracellular  iontophoretically-applied kainate  1981).  -5  M  granule  large  CA3  as  of  with  Deadwyler  10  depression a  enhanced  orthodromic  dose  depolarization  While  the  and  Thus  -5 M  respectively.  kainate's it  at  gyrus.  of  -10  major  electro-  depression  10  and  -6 pyramidal  a  and  the  reflected  amplitudes  threshold  Robinson  and  dentate  inhibitions  levels  the  between  comparable  the  which the  greater  -10  and  effect  gyrus. large  produced  kainate  fold  10  varied  CA3  observed,  were  acid  increases  50-1000  doses  in CAl,  e x c i t a t i o n , an  long-lasting  the  threshold  concentrations  synaptic and  the  were  not  actions related  which of to  implied  much a  lower  that  the  concen-  depolarization  of  171  neurons. PSs.  First,  Second,  potency  NMA  to  of evoked  Finally,  processes  change  that  ml  i n CA3  (10-15  ml  of  intracellular  recording  depolarization  (this  Deadwyler  study  [K ]  ml  (8-  +  affects 1-2  gyrus,  synaptic  of 10~  5  M)  —7 -10  to  M;  6  a  b u t d i d cause  i n the dentate  kainate  7  AD  cause  by i n c r e a s e d  of 10~ -10~  10  d i d not  excitation,  at which  of  e x h i b i t an e x c i t a t o r y  Third,  —8 and  the amplitude  kainate,  not mimicked  the doses  i n C A l (10-15  of  which  of  activity.  p o t e n t i a t i o n s were  mM).  no  enhancement o f s y n a p t i c  depressions kainate  was  and q u i s q u a l a t e ,  comparable  long-lasting  15  there  M)  cause  have  little  i n CAl only  1981; Westbrook  and Lothman,  the CAl region  kainate  been or  and  shown  no  by  membrane  see Robinson  1983; F i s h e r  and  and A l g e r ,  1983 ) . In been  shown  to  be  mediated  by  GABAergic  interneurons  (Anderson  Langmoen,  1980 ).  decline  that  which  The  could  be  blocked a  recurrent  which  has  a c t i v a t i o n of  e t al-. , 1964; D i n g l e d i n e of  ascribed  t h e IPSP  the  IPSP  was  t o the decrease  greater of  and than An  R-r...  IN antagonism receptor since:  Cl  of  ionophore  of any  e_t a l .  reported not  kainate  r  that  inhibition  1980 );  and  by  a  that  by  kainate  (2) F i s h e r  decline  iontophoretically-applied a change  events basis  of  GABA.  of  and A l g e r  GABA  an  binding  (1983 )  have  o f t h e IPSP  hyperpolarizing latter  the  indicated  [ H]-GABA  depression  The  at  f o r the effect  i n v e s t i g a t i o n s have 3  the kainate-induced  paralleled  implies  postsynaptic  i s an u n l i k e l y  (1) n e u r o c h e m i c a l  absence (Ruck  -  by  was  responses  observation  of the e q u i l i b r i u m p o t e n t i a l f o r C l  to  also was  172  not a  involved.  Therefore,  presynaptic  disynaptic basket  blockade,  pathway  cells, In  hippocampi  there  al.,  Franck  infer  that  a  the  actions  of  the  was In  is  a  the  virtually Chapter  region  i t was  electrophysiological the  slow  onset,  CA3  shared  a  hypothesis potassium (Fisher the  the  the  depression That  in  actions  of  the  CA3  of  no  IPSPs  1983) to  (Lancaster  et  i t i s appealing  to  kainate  cells  may  of  GABAergic  predispose  the  neurotoxic  BIC-insensitive  late  inhibition  in  was  the  The  not  decline was  of  was  also  both  an  that  by  kainate  in  the  GABA-mediated may  CA3  of  passive with  also  inhibition  detectable  in  analogy  may  this  "slow,  i t is possible  interneurons  and  the  blocked  By  CAl  with  a l t e r a t i o n of  i n CAl  early  similar  in  s e l e c t i v e blockade  involved.  of  their  Consistent  recently  that  kainate  late  of  kainate.  c h a r a c t e r i s t i c s , that  origin.  CAl  implies  of  because  reported  function  region  of  hyperpolarization  1983).  properties  impairment  acid-lesioned  kainic  pharmacological  IPSP"  inhibition  disinhibitory  in  the  suggested,  been  Alger,  GABAergic  iontophoretically-applied  inhibitory  i t has  of  the  consequently  common m e c h a n i s t i c  and  membrane  the by  and  dependent  late  and  in  disinhibitory  absence  pyramidal  that  acid.  abolished 3  synapses  1  sensitivity  depolarizing  CA3  the  appear  kainate s  Schwartzkroin,  vis  amino  in  since  virtual  and  at  i t would  overdepolarization  regard,  v i s \  to  an  involved  greater  former  elimination  either  by  be  this  interneurons  In  or  may  actions.  1983;  by  that  the an  underlie  CA3.  have  inhibition  been  due  to  173  differences trations  the  prevented  bath-applied  in  and  magnitude  iontophoretic of  kainate by  effects  paired  Fisher by  of  the  of  Alger  perfused  concen-  administrations.  firing  pulse  and  kainate  bath  epileptiform  the  on  depression  vs  i n t e r p r e t a t i o n of  determine  analyses  reversible  would  data,  no  various  have  attempt  doses  inhibition. (1983)  kainate  of  have  the  of  Intra-  revealed  a  GABA-mediated  CA3. No  iness  by  meaningful to  cellular  duration  induction  a  made  IPSP  the  achieved  Because  was  in  explanation  of  kainate  the  may  dentate  which were  be  offered  gyrus  not  to  the  expressed  here  for  the  refractor-  disinhibitory  even  after  actions  perfusion  of  of  10-15  -5 ml  of  10  M  reported granule  solutions.  kainate-induced cells  synaptic  acid  and  left  intensity fire  APs  in  of  enhanced  to  these The  1983). CA3  (1981b) of  have  dentate  the  amplitude;  reduction was  McLennan,  action  dentate curve  was  gyrus  the  usually  which  EPSP  but  1981;  as  a  shift  stimulus  cells  could  previously  were  recorded  not  to  manifest  plotting  intracellularly,  stimuli of  responsiveness  and  This  and  orthodromic A  neuronal  input/output  PS  extracellularly  with  Damiano discharges  (Collingridge  CAl,  the  against  subthreshold. or  also  Lothman,  extracellularly the  epileptiform  excitations  Westbrook  and  in. v i v o .  Kainic  to  Sloviter  either  invariably  intra-  associated  potentiations. simplest  concommitant  with  explanation a  decline  of  for the  the  increase  EPSP  would  of  the  be  a  PS sub-  174  threshold  d e p o l a r i z a t i o n of  threshold  for  same  time  tances.  firing  orthodromic kainate  having  as  were or  accommodation)  force  effect  of  for  with on  treatments  at  these  RMP  at  the  conduc-  changes  of  only  of  measured  somata.  best  the  concentrations  the  neuronal  but  synaptic  previously,  produced  no  impalements  slices  of  following  approached evoked)  driving  them c l o s e r t o  In  with  addition,  weakly  mimicked  potentiating actions.  In noted  bringing  no  discussed  little  depolarizing  kainate's  the  the  responses  intracellular other  (assuming  reducing However,  cells  the  point  present  acid  of  spike  evident  increased  of  preferential  the  in  EPSP  by  d e p o l a r i z a t i o n and  since  evidence  gyrus.  increase  for a greater  "as  maximum  may  a l .  (1982)  the  EPSP  (orthodromically-  sequence  dentate kainate  a  et  that  the  to  this  the  Auker  treatment  abolishment  investigation  particularly  dendrites  olfactory cortex  kainic  population  reduction  the  in  events  The  substantial  been  of  was  due  conductance  density  and  of  have of  size"  to in  a the  d e n d r i t i c vs  3 somatic  kainate  binding  studies  Although  no  detected  this  a  receptors  decrease  in  the  change  of  does  has  of  not  been  indicated  hippocampus the  transmitter  the  in  the  H]-kainate al_. ,  fibre  possibility  release  [  (Foster et  presynaptic  preclude  by  1981).  volley of  a  was  role  for  of  the  reduction  EPSP. The of  neurons  mechanism(s) exposed  to  underlying doses  membrane d e p o l a r i z a t i o n and  of  the  increased  kainate  decreasing  causing  the  EPSP  excitability little i s not  or  no  clear.  175  The  reduction  expected  to  Wigstrom  and  between For  orthodromic  the no  the  dentate  the  in  GABA-mediated inhibition  i n the could  CA3  some  the  cases  of  these  would  orthodromic  CAl  and  there  the  despite  and  the  increase  Quite  cases  basket  cells  to  mechanism  by  which  of  be  poor  the  Indeed,  in  with  postactivation the  to  input  of  apparent  associated  due  synaptic  (e.g. a  the  IPSP.  conceivably  could  was  be  enhancement  p o t e n t i a t i o n was  an  itself  potentiating actions  region  persist  in  excitation  However,  inhibition  inhibition.  in  of  disinhibitory  recurrent and  inhibition  1983).  example,  gyrus or  discharge  increase  excitation  of  change  an  Gustafsson,  kainate.  recovery  GABA-mediated  cause  correlation of  of  increase an  from  of  enhanced collateral  pathways. One probability producing then  a  cell  via  mechanisms  firing  to  could  orthodromic  an  coupling effect  (Traub  and  between  or  1979;  the  is  dendrites  dendrites  regenerative  Llinas,  increase stimuli  l o n g - l a s t i n g d e p o l a r i z a t i o n i n the  facilitates  possibly  et  of  kainate  by  which  and  somata  dendritic  spiking  a_l. , 1979;  Benardo  Wong e t  a l . , 1982).  C o r r e l a t i o n b e t w e e n e l e c t r o p h y s i o l o g i c a l and n e u r o t o x i c a c t i o n s of k a i n a t e Ibotenate,  NMA  and  L-aspartate  depressed  the  periods,  effect  an  depolarizations of  these  which  in  as  doses  SC-evoked was  (intracellular  compounds  higher  PS  associated data  not  depressants  L-glutamate in  with  CAl large  shown). of  for  evoked  The  and long  neuronal potency responses  176  parallels  their  directly  into  accordingly  ability  the hippocampus  i s  originally  t o cause  consistent  proposed  parable  t o those  similar  excitatory potencies  4] ) k a i n a t e  also  responses.  attributable to a direct  be  the case  That  with  i n both  striatum  (Zaczek  kainate-induced below  that  o f NMA  interaction tion of  kainate  that  Coyle,  (McGeer  1982).  mechanisms imposition  of  In  ongoing  increased kainate the  1982)  may  evoked  sufficient  i n some  et  cases  input  as  stimulation nucleus  e f f e c t s of acute  1981)  and  dose f o r  of  magnitude  but  undefined  terminals  since  transec-  increases  the dose  to a  level  and T a b l e the  similar  1 i n Zaczek  to and  i n t e r p r e t a t i o n of  complicated  actions  1979).  t o a unique  damage  be  a_l. ,  2 orders  to the striatum  may  e_t a _ l . ,  the threshold  i s ca.  o f e p i l e p t o g e n i c and l o c a l  auditory  in  (Kohler  the hippocampus  excitatory  depressed  that  (Nadler  e t a l . , 1978b  i n the cochlear  sparing  a n d NMA  f o r local  toxicity  and  of kainate  synaptic  Some o f t h e n e u r o t o x i c by  the  ascribed  with  pathways  required  f o r NMA  reflecting  (presumably  implies  damage  has been  of kainate  of c o r t i c a l  NMA  d e p o l a r i z i n g a c t i o n as appears t o  Coyle,  local  mechanism com-  hippocampus and  excitotoxic  CAl cells  ibotenate  the  1981) and  In doses  toxicity  be  e_t a _ l . ,  e_t a _ l . , 1 9 7 1 .  observation  the local  the  injected  o f t h e two compounds [ c f . C h a p t e r  depolarized  This  concentrations  with  l e s i o n s when  (Nadler  with  by O l n e y  required  local  by  the  super-  damage. of kainate  indicated  the a b i l i t y  of  t o augment t h e t o x i c i t y  of  (Mattox  by  are modulated  e t a_l. , 1980) and by  l e s i o n s of a f f e r e n t pathways (see  177  Introduction). the  The  excitatory  prolonged major  enhancement  ability  induce may  amino  hippocampal  unique  focal  be  results  a  distant  of  that  i t would  concentrations  of  cells  the  of  recurrent has  been  nmoles  to  circuitry  c_a. the  in  would  to  kainate  a to  there  an  would  acid  by  cells.  This  to  the  that  low  augment  limbic and  al. ,  favor  progressive  diffuses  ( S c h e r e r - S i n g l e r and  the  of  the  sequence  e_t  support  kainate's  promoting  (Foster a  intriguing  have  1981)  a  and  high where  increase  this  rapidly McGeer,  of  system  theory  following i t s intracerebral injection  1 jxl)  a the  by  That  suggest  nmoles/1)  amino  In  of  kainate  enhances  epileptiform firing  excitation.  brain  is  discharge  of  each  of  of  causes  is paralleled  kainate  applicable  receptors  shown t h a t  throughout et  prone  synaptic  in  the  discharge  kainate  neuronal  kainate  that  e l e c t r o p h y s i o l o g i c a l and  reasonable  of  show  l i m b i c system.  these  (10-100  particularly  are  only  action  the  of  seem  capabilities  seems  study  excitation  this  neuronal  kainate  synaptica1ly-evoked  density  and  actions  Given  neurotoxicity  where  synaptic  between  neurotoxic  events  tested  l e s i o n s throughout  possibility.  present  intracerebral injections  correlation  neurotoxic  the  acids  regions  of  of  of i t (2-5  and  widely  1979;  Zac'zek  a l . , 1980 ) .  Mechanism(s)  of  Previous (Dingledine have  and  shown t h a t  kainate's epileptogenicity studies Gjerstad, a  with 1980)  reduction  of  convulsants and  BIC  such  (Wong a n d  GABA-mediated  as  penicillin Traub,  inhibition  1983) may  be  178  a  crucial  factor  Consistent  with  intracellular showed an  the  induced  of  applied  of with  kainate  orthodromic be  While  cannot  hypothesis  did  of  et  slices  in  (Traub  the  since  CAlb did  which  to  firing  induction  CA3  the  seems  kainate-  Alger,  dentate  recurrent  PSs; BIC  the  of  1983).  gyrus  bath-  inhibition  and  continued  subsequent  blockade  did  epileptiform  cause  not  of  membrane  pyramids. burst  other  and  activity  Hotson  and  Lothman  transmission  had  CA3  Prince,  (1983) firing  that in  in  for  properties  and  burst  of  atten-  requirement  1978;  firing  actions  the  biophysical  However  underlying  enhancement  the  current-evoked  cause  of  epileptiform  Westbrook  synaptic  factor  1983)  explain  (Prince,  of  major  Wong,  may  evoke in  a  simultaneous  and  inhibition  changes  possible  in  with  also  greatly potentiated,  with  Thus  kainate  burst  1983)  a l . , 1982).  discounted.  of  It  and  contribution  kainate-treated in  in  reduce  single  and  epileptiform discharges (Fisher  although  extra-  associated  involved  the  to  stimuli  not  inhibition  Finally  the  evoked  this  be  epileptogenic!ty  neurons.  noted  be  neurons  to  activity.  Alger,  inhibition.  may  CA3  recurrent  are  i n CAl  of  and  be  doses  leading  and  excitation  of  firing  Fisher  may  be  uation  also  results  disinhibition  recurrent  1981)  burst  (cf. Misgeld  kainate's  (see  epileptiform  the  GABA-mediated  comprised  firing  of  hypothesis  responses,  GABA-mediated  low  of  spontaneous  Consistent  onset  disinhibition  populations  to  this  onset  that  the  recordings  abolition  likely  in  have in  antidromic  kainate-treated been  blocked  179  suggests  that  relatively  changes  of  unimportant.  passive  membrane  characteristics  are  180  CHAPTER  6  F O L I C A C I D - I N D U C E D CHANGES OF NEURONAL  EXCITABILITY  Introduction Kainic elicits  a  acid  complex  intracerebrally.  destruction  system. also  A  at  disease  similarity Implicit acid  at site  t o those  receptors  neurological  with  which  observations may  be  disorders  ligand  of  electroto the  o f t h e i n j e c t i o n and i n i n the limbic  diseases  i n humans a r e  lesions  of s p e c i f i c  the neuropathologies  temporal  lobe  epilespy,  degeneration  may  be  induced  bear by  and t h e r e f o r e  f o r the receptor  evident Hunting-  a  striking  kainic  i s the possibility  involved  which  injected  i n addition  p a r t i c u l a r l y those  or cerebellar  i n these  endogenous  seizures  F o r example  i n patients  when  the production  by l o c a l i z e d o r d i f f u s e  of neurons.  excitant  reactions  number o f n e u r o d e g e n e r a t i v e  autopsy  ton's  structures  characterized  classes  include  both  neuronal  of  and b e h a v i o r a l  of neurons  brain  powerful  spectrum  These  encephalographic  distant  i s a  that  i n the etiology  than  kainic  of  the i d e n t i f i c a t i o n i s o f more  acid.  some  of the  heuristic  interest. Although it  sharing  seems u n l i k e l y t h a t  endogenous a g o n i s t  structural  similarities  L - g l u t a m a t e may  f o rkainate  fulfill  receptors.  with  kainate  the role of the  Thus a n t a g o n i s t s  of  181  L-glutamate  excitations  responses  (McLennan,  do  1981;  not  affect  Watkins  and  kainate-induced Evans,  1981);  L-  binding  to  3 glutamate  weakly  neuronal  membranes  conductances excitants 1980); to  (Evans  kainate  charged  1979);  the depolarizing  actions  et. a _ l . , 1 9 7 7 ; M a c D o n a l d  groups  and L - g l u t a m a t e  ligand  acid  requires  side  et. a _ l . ,  suggest  i s likely  chain  properties of  (Nadler  relationships  t h e two  i n addition  isopropylene  (pteroy1-L-glutamic  of L-glutamate  acid;  the reduced  folic  acid  to  1981).  that the  incorporate  a  form  coupled of  by  acid)  an N - a c y l  the molecule,  consists bond  to  of a  pteroic  5,6,7,8-tetrahydro-  (THF) a c t s as a m e t h y l group a c c e p t o r t o y i e l d t h e 5 5 N - f o r m y l - T H F ( f o l i n a t e ) and N -methyl-THF (MTHF).  co-enzymes  of  modulation reported  of  ionic  and Wojtowicz,  and t h e n e u r o t o x i c  receptor  the  conformation.  F o l i c  number  an i n t a c t  are dissimilar  f o r the kainate  molecule  of kainate  structure-activity  glutamate-like  [ H]-kainate Coyle,  e t a_l. , 1974 );  Nonetheless  studies  of  potent,  of  folic  confirmed  by  strations  of folate,  Spector, folate  enhancement  have  neuronal  the  injections  brain  and  the e x c i t a t o r y potency  the three  1960;  specific  (London  underlying  differ  (Johnston  A  inhibits  others  suggested  role  excitability.  convulsant acid using  1971; H o m e s concentrations  of  and t h i s  systemic  or  a n d MTHF  and Obbens, has a l s o  (Goff  f o r folate  Hayashi  actions  i n dogs  folinate  of kindling  a  e t a_l. ,  (1959)  first  intraventricular effect  intracerebral i n rats  (Noell  1972).  An  been  i n the  been  adminiet. a l . ,  increase of  correlated  1978) o r  has  with  an  sound-induced  182 seizures  (Cooke,  measurable al.,  i n cobalt-induced  1971).  change  1978) and an e l e v a t i o n  Originally  however  this  (Meller  epileptic  a mechanism  of neurotransmitter hypothesis  of folate foci  (Mayersdorf  of action  methylation  reactions  has  been  since  levels i s e_t  involving a was p r o p o s e d ,  proven  incorrect  e t a l . , 1974 ) . Recently  inhibited  Ruck  et  (K.= 0.88jXM)  a l . (1980)  the binding  reported  of  that  [ H]-kainate  MTHF to r a t  3  3 cerebellar glutamate al.  membranes; binding  (1981,a,b)  injections death  of  folate  of f o l i c  activity  lesions.  Folic  orders  may  of magnitude  be  kainate acid  with basis  derivatives They  (50-100  was a p p l i e d  data  of  neuronal  intrastriatal  induced  neurons than  prolonged  that  remote  locally  and i t s  o f MTHF b u t 1-2  f o r kainate. that  i n part  by  the actions an  of the three  major  changes  properties  of folate  interaction  i n the present  (1) understanding  f o r i t s convulsant  that  the  the kainic acid-like  suggest  t o determine  with  nmoles)  that  Accordingly  t o each  slice  found  was g r e a t e r  at least  receptors.  t h e aims  its  l e s s than  these  mediated  hippocampal  of intracerebral  d i d not destroy  as a neurotoxin  [H ] -  the neurotoxicity  and produced acid  Together  Aware  antagonized  f i n d i n g , Olney e t  and  acid  weakly of this  by k a i n a t e .  seizure  potency  only  ( K ^ = 83yA.M). compared  produced  injection  MTHF  study  regions  i n neuronal  with folic  of the r a t activity  the electrophysiological (2) a l l o w i n g  a  comparison  183  with  kainate-induced  relating  the  a l t e r a t i o n s of  synaptic  e l e c t r o p h y s i o l o g i c a l and  processes  neurotoxic  and  (3)  actions  of  folate.  Methods For 52  male  the  experiments  albino  incubation  for are a  maintained  and  slices  in_ v i t r o in  recording  obtained using  Chapter extra-  2  and  from  standard  where  the  intracellular  outlined.  5-10  the  hippocampal  described  evoking also  After responses  were  conditions  techniques responses  rats  73  min  period  of  recording  perfusing  medium  was  changed  stable  submaximal  to  containing  one  - 4 - 3 10  -10  M  istered  folate  before  and  i n most  returning  to  cases  normal  10-15  perfusate.  ml  were  The  admin-  effects  of  — 6  BIC  (1-5x10  drug pH  of  M)  solutions  were  were  also  examined  i n some e x p e r i m e n t s .  f r e s h l y prepared  and  adjusted  to  a  A l l final  7.3-7.4. Results  Effects  of  f o l a t e i n CAl  Because  folate  activity  which  synaptic  a c t i v a t i o n of  induced which  changes  the  CA3  and  region  dentate  elicited  produced  i n CAl  the  via CAl  in  Schaffer cells  responses had  CA3  been  (see  were  gyrus neurons  spontaneous  collateral below),  assessed  fibres  the  a  folate-  in slices  removed.  Perfusion  of  large  long-lasting  from  folate  -3 (10  M,  10-15  ml)  produced  a  and  (up  to  184  Pre - treatment  Folate  Recovery  A  I  —  6.1  pathway.  ,  ,  ,  i  0  -10  20  .30  from  stratum  following B.  recorded  (3  intensities  records  and  courses  of  simultaneously  at  a  rate  of  was  c a . 1 ml/min.  V  —,  i 45  on  50  responses  i n CAl  superimposed  the peak M),  evoked  In  this  the  PS  in strata perfused  by C.  and  6  of  three  depression  pyramidale during  the  EPSPs  different  arrow  In another  figures  below slice  of and  time  and  V.  subsequent an  sweeps)  effect  sweeps)  with and  indicated  of  stimulus  superimposed  i s downwards.  p o t e n t i a t i o n of  Folate  at -3  (10  radiatum,  is  m V  Schlatter collate.ral-commissural  folate  stimulation  respectively.  folate  before,  stimulation.  2  ( mm )  (3 c o n s e c u t i v e  consecutive  negativity  of  the  Records  stratum  of  of  recorded  of  p e r f u s i o n with  from  time  actions  pyramidale  Records  the  The  A.  *  ,  by . s t i m u l a t i o n  (SC)  3  -5  evoked  J  4 msec  B.  Fig.  ,  the  the time EPSP  radiatum indicated  185  Fig. tions  as  for  stimulation from  the  absence courses  6.2  Actions  Fig.  of  the  middle of  of  any the  6.1.  of  folate  A.  Records  medial portion  change changes  in  in  perforant of the  elicited  from path  stratum EPSP by  area  in  dentata.  stratum (6  granulosum;  V).  moleculare; this  folate.  Conven-  slice.  B.  Records  note C.  the Time  186  Stimulus Intensity (V)  B. -—-  8 -i  > E  4 -  Q_  00  Q_ LU o -  "T~  4  7  5  Stimulus  Intensity  Pre- treatment  Fig.  6.3  successive symbols) evoked  relating  A  the  EPSP  too  small  dentate  in in to  at  elicit  EPSPs.  standard  before  CAl, CAl  curves  intensity  responses  folate,  F o l a t e (10  Average  responses,  B  in  of  and area  PS;  and  (each  M , 10 m l )  point  deviations  stimulation at  the  dentata.  a l l stimulus a  (V)  peak  of  intensities,  the  lesser  of  smaller  and  Note  mean  than  amplitudes the  the  effect  reduction  including  action  of  3  of of of  those  folate  on  187  1.5  h) p o t e n t i a t i o n  n=8) e v o k e d of  typical  reduction in  part  responses  recorded  similar  stimuli  6.1B:3  V).  markedly  o f t h e SC p a t h w a y .  i n Fig.  have  course  together  6.1.  The  attributable  o f a s m a l l r e v e r s e d PS  i n the f i e l d  radiatum  also  gyrus  ( F i g . 6.1B:7  observed  subthreshold  In the dentate  increased  The t i m e  been  i n the stratum  were  +S.E.M.,  o f t h e PS a n d E P S P  i n F i g . 6 . I C may  r e d u c t i o n s were  which  467+59%, mean  are illustrated  t o the appearance  however  (maximum  i n the amplitudes  o f t h e EPSP  potential  by  by s t i m u l a t i o n  the changes  with  o f t h e PS  f o r PS  of  evoked  generation( F i g .  similar  the amplitude  i n EPSPs  V) ;  doses  t h e PS  of  evoked  by  i n c r e a s e s o f 760+177%  (Fig.  Depression  o f t h e EPSP r e c o r d e d  simultaneously i n  molecular  layer,  of  the  middle  termination,  was  6.2C) a n d w a s the  change  the  peak  spike  Effects  than  reduced.  of  observed  MPP  three  in CAl.  synaptic  slices (Fig.  F i g . 6.3  between  hippocampal  PS was s o m e t i m e s  of f o l a t e  shows  the stimulus  i n C A l ( F i g . 6. 3 A )  In both  of a second  The  b y AD s t i m u l i  one  relations  a n d PS  ( F i g . 6. 3 B ) .  appearance  region  that recorded  t h e EPSP  of the action  was  evoked  and  gyrus  transient  less  i n only  of the input/output  intensity dentate  observed  the  been  MPP  stimulation, 6.2).  (n=3) h a v i n g  folate  and t h e  areas noted  during  and t h e l a t e n c y o f t h e  amplitude  and waveform  were u n a f f e c t e d  of  the  first  responses  (data not shown).  i n CA3 -4 With  the  lower  amplitude  and  doses  (10  decreased  M, the  10-15 ml) latency  of  folate  increased  orthodromic  PSs  188  evoked  by  FIM  stimuli  ( F i g . 6.4b,d;  progressive  increase  i n t h e number  immediately  followed  i t was  MF-evoked  1°  PS  stimulation  affected  following  PSs  appearance  of m u l t i p l e  with  6.8).  amplitude of  of  6-12  mV  with  a  varying  6.8a-lower  trace).  persisted  were  positivity  5-10  SPBs  until  t o peak  the experiment  was  of  small  t h e SPBs  were  n e g a t i v i t y ) and with  i n every  The  coincided  and  in different  occurred  of  (SPBs) ( F i g .  later  regularity  sec  also  bursts  min  collateral  amplitude  infrequent  5-10  which  6.5c,f).  responses  population  clock-like from  and  a  neither the  Schaffer  6.4a,c,e,g;  however  (peak  by  number  i n evoked  SPBs  duration;  intervals  and  PSs  spontaneous  amplitude  discharged  (Figs.  Initially  and  the  of PSs  In contrast  elicited  but  increased  the onset  6.5g;  PS  Thereafter  and amplitude  observed.  n o r t h e AD  was  6.5Ab,e).  interburst  slices ( F i g .  subregion  terminated  of  CA3  (up t o 4  h  _3  later). only  When  10  detectable  M  solutions  d i f f e r e n c e was  of  responses With  variations responses were  (up  first  folate  100  by  of  the output  of  msec)  low  of  intensities  be  an  SPB  signal  to SPB was  and gated  the  of evoked  and  depression  amplitude  the latency  appeared  perfused  of  CA3  observed.  large  6.6A;  an  A direct  never  ( F i g . 6.5h;  following  spike  was  appearance to  evoked  bursts  stimulus  and  the  observed  delayed  by  were  the f a s t e r onset  spontaneous e p i l e p t i f o r m a c t i v i t y . evoked  folate  ( 5 - 1 5 V)  6.7).  related  The to  to test with  an  the  this  of of  SPBs  orthodromic FIM  stimuli  latency  of  the  timing  of  the  hypothesis  adjustable  o f t h e d i s c r i m i n a t o r was  large  used  DC to  the level  trigger  189  Fig. responses  6.4  Effects  recorded  stimulation  o f MF  or  below upwards, p l o t the  primary,  sample the  graph  the  are  i n the  terminated  of  course  of at  above.  b,d,f,h.  The by  potentiation  stimulation.  Peak-to-peak  were measured  in this  the  evoked  by  graphs,  of  Folate  of  the  PSs;  in  responses: msec  the  of  the  stimulus  application PS  of and  indicated  activity  primary  from  amplitude  times  "ringing"  the  on  quaternary  1.5  amplitudes  experiment.  The  initial  bursting of  and  CA3  MF-evoked  r e c o r d i n g system. spontaneous  in  changes of  and  sweeps) t a k e n shown  M)  inputs.  tertiary  contaminated  tape  before lack  time  (10  pyramidale  orthodromic  are  FIM-evoked:  records  artefact Note  (single  lowermost  sample  FIM the  folate  stratum  secondary,  records  a,c,e,g,;  in  of  was  developed.  evoked  by  MF  population spikes  190  191  'Fig. AD-evoked  6.5  Changes  activity  (first,  respectively)  in  campal  slice.  Filmed  each  horizontal  [c])  in  same  time.  The  FIM-evoked were the  and  AD  PS  (•  in  SPBs  latency  the  If  as  subthreshold  be  observed were  (j).  virtually  intensity  identical  a  of  the  and  hippo-  except  at  folate  was  PSs absence  AD an  the  PS  evoked  (g,h,i,j).  and  approximately  the  to  of of  was  delayed  in  increase AD any ca_.  large  AD-evoked  stimulation AD  [b]  (e);  coincidentally not  and  traces  administration  but of  column  sweeps  the  FIM-evoked of  obtained  for  for  Spontaneous  of  additional  and  FIM-(h)  occurred. to  the  were  save  (g)  third  (single  effect  After  appeared of  row  recruit  f ) .  and  region  records  affected  spontaneous,  second  CA3  initial  similarly  folate the  PS  the  in  the  responses change 13  ml  of  shifts  of  response  (i)  decreased bursts  epileptiform  in  so  were  discharges  192  A  (o)  8. (a)  2.6 sec  3 sec (b)  (b)  3.6 sec (C  )  5 sec  -^vV^-  (c)  4.6 sec  6 sec (d)  (d)  8 sec  5 6 sec (e)  (e)  (f>  (f)  10  10 m V 10 m s e c •20 •• 10 sec  Fig. responses  6.6  in  CA3  at  and  and  representative given  burst-shock tion  a  latency  latency  for  a  B of  2.5  f o r A; the the  sec  PS  a  decreased the (not  bursts  differences  the  in  spontaneously  onset  ([e],  and  at  Two  FIM  At  a  one  5.3 to  and much  shock 3.6  of  (b);  V.  In  another,  resembled slower  sec  stimulathe  increased  Stimulation at  folate  successive a  intensity  was  in  a c t i v a t i o n was  B.  latency,  FIM-evoked  to  occurred  (pre-drug)  comparable  (a)  burst.  interval  i n t e n s i t y constant were  A.  same  illustrated).  of  response  discharge  as  control  no  the  sec  intervals  spontaneous  with  (o-d) (e) (f)  latency  slices.  showing  after and  the  burst-shock  epileptiform  stimulus  evoked  occurred [f ] ) .  mV  in  in  treated  :  traces  interval  further  The  as  2.6  delayed  (f)  various  b i c u c u l l i n e ' (B)  V)  20 10  Changes  (A.)  (9.0  10'msec  (b-e)  (o)  mV  burst (c,d).  2.8  msec  conventions both  A  and  regardless those  sweep  which  speeds,  193  a  single  of  stimulus  this  (3-15  cycle  V)  and the  but constant  interval). the  SPB  burst  shock  burst  decreased  so  6.6Ae).  that  between  b u r s t s were  stimulation  V).  When  CA1/CA2 latency were  from  SPBs  were apparent  3  cases  treated  slices  was  between  the dentate  gyrus,  of  delayed  intensities  of  by  a  shock  applied  i n the  response  of stimulation of  an  AD  i n  which  PS ( F i g .  of latency d i d not appear t o  were  before  fields  relationship  ( F i g . 6.7:10.5  o f a n SPB relatively  since  for a  constant and  o f t h e expected SPB.  the spontaneous  studied  and t h e  situation  variations  f a r i n advance  experiments  changed  w a s t h e same a s  appearance they  were  response  f o r the generation  interval  and were  constant  At the higher  elicited  the shifts  response  or duration  a direct  only at intensities  any c o i n c i d e n t a l  i n some  As t h e  the  and t h e l a t e n c y  comparable  represent  In  of  amplitude  remained  varied  strength  I n any case  occurred  was  were  the hilus  burst-shock  low  (burst-shock  one t o a n o t h e r  i n.their  i n the pre-treatment  subthreshold  a  any response.  Regardless  interval  6.5i,j).  given  t o evoke  demonstrated.  responses  or  of  t h a t up t o 3 s e c f o l l o w i n g  the l a t e n c y of the evoked  observed  varied  When t h e c o n d i t i o n s o f s t i m u l a t i o n  be  that  stimulus  then  comparable  of stimulation  could  the beginning  was i n c r e a s e d t h e l a t e n c y o f t h e d e l a y e d  the stimulus  bursts  was  illustrates  the burst-shock  intensity  between  of the  ( F i g . 6.6Ab-d).  indistinguishable (Fig.  interval  delivery  not possible  interval  evoked  The  intensity  F i g . 6.6Aa  i t was  latency  cycle.  activity  and a f t e r  in  folate-  the connections  CA3 a n d C A l w e r e  cut using a  194  Fig.  6.7  orthodromic plotted  The  changes  response  f o r several  i n the latency  at various  intensities  of the FIM-evoked  burst-shock  intervals are  at stimulation.  Epileptiform  -3  activity  was  (+S.E.M.) (except stimulus the  was  where  induced based  intensity  stimulus  the  on  indicated  burst-shock  constant  by  the  the by  of  (10  results bracketed  latency  interval;  latency  strength  folate  when  was the  the evoked  increased.  M, of  15  ml).  five  successive  numbers). inversely  At a  mean tests  constant  proportional  burst-shock burst  Each  interval  decreased  to was  as t h e  195  hand-held  microknife.  following side by  of  this  the  in  the  Stimulation orthodromic  PS  CA3  the  slice  but  CAl  and  in  p e r s i s t e d only  intact  and  CAl  no  completeness  on  they  responses  intervention.  b e t w e e n CA3  PSs  indicating  SPBs  surgical  cut  multiple  Evoked  1  of  3  i n the were  produced  of  the  of  the  slices  recorded  an  AD  was  CA3  followed in  CAl  transection.  evoked  smaller  both  the  evident  cut  CA3  on PS  axonal  a  isolated  unchanged  Stimulation  response  side  were  a  single  secondary  region  whereas  i n CA3  and  PS.  in  CAl  the  (Fig.  6.8d-f) . To  determine  transmission  the  i f  SPBs  e f f e c t s of 2+  contain  0.5  indicates blockade  mM  Ca  the of  were a  dependent  perfusing  on  medium  synaptic  altered  to  2 + /8  mm  Mg  elimination  MF-evoked  were SPBs  examined.  coincidentally  epileptiform  orthodromically-evoked  activity  Fig.  in  firing;  CAl  was  6.8a-c  with  the  spontaneous also  and  abolished  by  the l a t t e r treatment (data not shown). These e f f e c t s of low 2+ 2 + Ca / i n c r e a s e d Mg were observed i n each of 4 s l i c e s t e s t e d and  were  reversible within  15-20  min  medium.  of  returning  to  normal  ;' Intracellular  perfusion  from mV  10  than  -60  and  cell  spontaneous  SPBs  recorded  6.8g)  .  CA3  neurons  spike  of  with  bursts  extracellularly  membrane  were  amplitudes  firing  I n j e c t i o n of  spontaneous  recordings  with  obtained  during  folate  resting potentials  greater  of  at  of  APs  an  least was  80  mV.  In  synchronized  adjacent  hyperpolarizing  current  depolarizations  (Fig.  electrode revealed  6.8h)  each with (Fig.  sudden  (associated  196  7*  10 msec (c)  (b)  (a)  10 sec  CA I (f)  (e)  (d)  mV  |*4  my  CA3  -^WiYrr,  4 0 m s e c (e) 10 sec (d.f)  mV  1  Fig. medium  and  folate.,  normal  in  abolished pathway  for  action  this  was cell  spontaneous a  (lower  The  (g):  potentials synchronized to  -80  mV  to  low  following  (e):  of  in  between the  occurring 2+  [Ca  MF  1°  different  pyramidale  after  a  CA3  faster  in  folate of  the  burst  records  in  ]/high  PS  the  in  the  slice  spon-  CAl  (upper  CA3-CA1  (e).  a  folate-treated  with  SPB.  the  (h):  revealed  follower (See  neuron  text firing (lower  Hyperpolarization PDS  spontaneous  prolonged  discharges.  a  CA3  a-re  axonal  Intracellularly-recorded burst in  underlying burst  ]  of  treatment  and  CA3 2+  [Mg  restoration  the  a  transection  ( i ) : a  show  epileptiform  in  RMP  discharge.  during  traces)  latency shown  traces)  stratum  following  of p e r f u s i o n with a 2+ Mg on e v o k e d (upper  mM  persistence  in  is  baseline  follows  the  8  (lower  recovery  recorded  CAl  and  (b)  and  in  effects  bursts  (d)  explanation.)  trace)  on  (b).  CA3  CAl  Ca  The 2+  before;  Note  and  (f).  in  mM  partial  bursts  traces),  of  (c)  trace  burst  (a)  medium.  taneous  0.5  spontaneous  perfusion;  top  (a,b,c):  containing  traces) after  6.8  40msec(q.h) 100 (i)  of the  superimposed  hyperpolarization  which  197  extracellularly  with  SPBs)  which  depolarization  shifts  icillin-treated  hippocampus  in  Marsan  (1964).  Spontaneous  bursts  (0.5-1  sec)  (Fig.  hyperpolarizations  of  folate  Folate excitatory  on  has  by  in  Matsumoto  were  with  paroxysmal  described  vivo  recurrent  been  properties  depressant  that  first  the  followed  amplitudes  the  pen-  and by  of  Ajmone  prolonged  up  to  10  mV  6.8i) .  Actions  any  (PDSs)  resembled  i t was,  Therefore  on  actions i f at  i t  epileptogenic  found  have  folate  a l l , a  on  weak  of  folate  inhibitory  processes.  This  comparing  the  of  actions  determining  the  changes  accompanied  folate-induced EPSPs  or  at and  evoked  excitant that  in  be  due  possibility  was  with  recurrent  best  very  the  the  weak  absence  responses  the  might  folate  in  no  neurons  possible  actions  Although  to  cortical  of  seemed  inhibition  suggested  hippocampus.  potentiating to  a  of  BIC  inhibition  and  reduction  addressed  those  of  in  of  by  (1)  and  (2)  CAl  which  potentiations.  were  not  monitored  perfusion  of  BIC  — 6 (1-5x10  M)  potentiated  epileptiform been  reported  Misgeld of  discharges  the  evoked  e_t  1982). of  orthodromic  recruited; recorded  MF  In  folate. PS  responses  after  in  previously  a_l. ,  effects  orthodromic  and  the  PSs  dentate  gyrus  (Schwartzkroin the  CA3  Thus  region the  increased comprised Schaffer  and of  and  caused and  CAl  evoked as  has  and  Prince,  BIC  reproduced a l l  amplitude  of  additional  bursts  collateral  of  PSs  1978;  the  FIM-  PSs  were  were  also  stimuli  but  in  198  either  situation  (data  not  FIM  of  were  previously  stimulus  treatment  responses  and  difference  was  unchanged  variations  responses  evoked  by  with  and  the  these  low  were  burst-shock  in  the  intensities  correlated  interval  PSs  of  5/5  slices  the  p e r f u s i o n of  from  an  as  and  the  the  10-15  ml  the  509+72%  for  the  subsequent  disinhibition potentiating inhibition the be  responses.  from  had  of  which  amplitude attributed  to but  kainate  recovered of to  as  of  the  the  unconditioned against  F i g . 6.9a).  been  removed,  t h e r e was inhibition  amplitude the  data  kainate as was  the an  AD  PS  any  of  in  also  ( F i g . 6.9d).  of  a  change  alteration  of  after  either at  a  6.9  time  mV, were  transient 5  the  when  the  neither of  no  (maximum  Chapter  apparent Since  In  1.7+0.2  Fig.  caused  noted  AD  percent  plotted  (e.g.  folate  stimuli  SC-evoked  p o t e n t i a t e d ( F i g . 6.9b)  exposure  ( F i g . 6.9c) action  M  pre-treatment  slice  PS  folate  conditioning  was  r e g i o n had -3  10  was  a  c o n d i t i o n e d and  enhancement  u n c o n d i t i o n e d PS  by  inhibition  CA3  of  occasions  calculated  unconditioned  which  mean  msec)  'was  percent  when t h e  In  (20-30  following  using paired  alternate  between  the  apparent  a  on  inhibition  and  amplitude  or  Briefly  the  i n CAl  extracellularly  preceded  the  cannot  PS  large  observed  i n amplitude  orthodromic  affected  first  recurrent inhibition  2).  were  stimulus  derived  the  addition,  were determined  Chapter  n = 5) .  of  s t r e n g t h ( F i g . 6.6B).  Changes of  change  In  orthodromic  stimulation  discussed  (cf.  amplitude  shown).  latencies of  the  acid  inhibition  conditioning  199  S  (c)  80-,  (o)  8CH  40  40-  0-  4  6  8  2  4  6  8  Population Spike Amplitude ( m V ) Koinate. I0' M D  Folo 1 6 , 1 0 %  miiiiuiiiHii > 6 (o) »  (d) »  (c)  (b)  6 <  5  ao  CL  0  J  rV/n  + 10  20  50  60  75  85  100  -rVA,— 110 120  130  kainate  on  Time (min)  Fig. recurrent (see  The  inhibition  text  preceded mV  6.9  antidromic  intensity  PS  and  constant  by  a  the  produce  t h e PS  after  only  (10 ^  M);  with  upper  ditioned  folate  lettered  graphs  test  were  orthodromic  stimulus  (10  M);  below  arrows  varied  inhibited  to The (b)  perfusion  with  was  reduced  course  of the  the times  Solid  was  stimulus  (a) c o n t r o l ;  t h e time  indicating  PS  the abscissa.  inhibition  shows  circles  was  a 3  stimulus  the  (c) a f t e r  The  constructed.  PS; open  on  when  evoking The  responses  of i n h i b i t i o n , -3  (d) r e c o v e r y . The graph  response  whereas  indicated  the degree  with  by k a i n a t e .  changes, the  show  perfusion  kainate  amplitudes  inhibition  of the antidromic  experiment,  f o r the unconditioned  (above)  AD  test  on . t h e o r d i n a t e .  voltage  graphs  and  .The p e r c e n t  conditioning  the amplitude  throughout  folate  o f an S C - e v o k e d  i n plotted  thus  of  of C A l neurons.  f o r details) (20 m s e c )  effects  at  circles  which uncon-  responses.  200  Discussion In of  CAl  folate  neuronal  in  the  Similar  by  for  kainate,  the  MF-evoked  and  CA3 the  1°  input this The  PS  was  was  doses  manifest associated of  the  as  an  with  no  dendritic  synaptic  excitability  dose w i t h  bath  c_a.  i n CAl of  10-fold  are  applications  1000-100-fold  with  of  related  the  and  less  represent  CA3  were  bursts  of  was  a f f e c t e d by  not  be  the  (Schwartzkroin  case and  EPSPs  with  the of  multiple  SPBs.  medium  which  folate-induced of  synaptic  large  intra-  (Johnston  observed  neurons.  to  Since  and  underlie recurrent  f o l a t e i t seems u n l i k e l y  epileptiform activity to  in  hypothesis  summed  1982)  of  of  modulation  this  but  antidromically-evoked  neurons  a  folate  stimulation  indicated that  to  of  response  appearance  CA3  with  a_l. ,  dose  Eventually  spontaneous  e_t  lower  p o t e n t i a l comprised  transmission  which  appears  penicillin  field  of  was  spontaneous  as  and  orthodromic  discharge  Lebeda  onset  a  associated  PDSs,  inhibition  tion,  a  Consistent  1981;  the  similar  synaptically-evoked was  being  unaltered.  elicited  orthodromic  cellular  that  of  min)  of  amplitude  threshold  region  burst  processes.  the  PS  peak  changes  of  which  the  the  FIM-evoked  epileptogenesis  Brown,  of  (10-15  abolition  synchronous blocked  perfusion  folate.  potentiated  PSs  in  the  durations  In  either  effect  amplitude  EPSP.  gyrus  probability  an  reduction  similar  than  the  a  produced of  dentate  increased  or  field  the  discharge,  increase change  and  was  due  kainate  Prince,  1978).  to  disinhibi-  (Chapter  5),  However,  and this  201  possibility  cannot  assessment entirely instead  of  changes  plausible  excluded  without  of  i n CA3  that  IPSPs  the  l i e i n i t s capability  Compounds  such  diisopropyl possibly their  be  as  burst  firing  pyramids.  K  l  +  t o enhance  excitation.  recurrent  o f CA3  2)  neurons  may  tetraethylammonium,  (Lebeda  (Chapter  0  I t i s  folate  4-aminopyridine,  t  intracellular  e p i l e p t o g e n i c i t y of  phosphof l u o r i d a t e  increased  an  et  a_l. ,  appear  1983)  also  by p r o c e s s e s  to  and  produce  other  than  disinhibition. The CA3  triggering  region  isolated due  since  CA3  this  island.  to synaptic  indicated  mechanism  by  activity That  excitation  their  persisted  v i a Schaffer  at a constant  CA3  ance  following transection of the synaptic  CA3 and  T h e same  has been  reported  An the  intriguing large,  kainate  bursts in and  have  vivo more  fibres  latency  following  (Wong  (Schwartzkroin  disappear-  connection occurring  was  from  CA3  i n C A l and  and Traub,  1983)  Prince,  1978)  i n t h e CA3  region  and  slices. feature  variable  dromically-evoked or  i n C A l were  immediate  between b u r s t s  in bicuculline-  penicillin-treated  hippocampal  was  ( F i g . 6.8e) a n d t h e i r  relation  bursts  t o the  surgically  collateral  each  to C A l .  intrinsic  i n the  the spontaneous  appearance  discharge  f o r S P B s was  burst  shifts  firing  (Fig.5.6Aiii) been  reported  (Matsumoto recently  observed of  the  following  treatment.  as o c c u r r i n g  et_ a l . ,  only  (Wong  folate, Delayed  of  ortho-  bicuculline synchronized  i n the hippocampus  1969; D i c h t e r  ir\ v i t r o  latency  and  and S p e n c e r , Traub,  both  1969a)  1983).  In  202  agreement bursts  could  various and  with  with  appeared  that  the delayed  the time  required  networks  i n an  from  SPBs.  latency  of  a correlation with  of burst  firing  a n d Wong,  fashion  addition  to  delayed  the timing  an SPB was d e m o n s t r a t e d .  onset  delayed  a c t i v a t i o n of  In  f o rsignal processing  (Traub  (1983)  all-or-none  the onset  strength,  following  and Traub  low i n t e n s i t y  the r e l a t i o n s h i p of  the stimulus  local  inputs,  to the stimulus  appear to  elicited  o f Wong  indistinguishable  confirming  of  be  afferent  were  bursts  the results  I t would  i sa t t r i b u t a b l e within  complex  1983).  Mechanism o f A c t i o n It  i s u n l i k e l y that  ically-evoked  P S s by  folate  metabolism.  First,  f o l i c  slices 1977)  of brain  and the r e d u c t i o n  forms  (THF)  amethopterin has  acid  (Makulu  orthodrom-  t o i t srole  i s very  e_t a _ l . ,  slowly  in  cellular  taken  1973; Spector  up by  et al.,  of folate to i t s metabolically  probably which  i s due  of  occurs  blocks  intracellularly.  the enzymatic  active Second,  reduction  of  folate  no e f f e c t on e i t h e r t h e e l e c t r o p h y s i o l o g i c a l ( C l i f f o r d  Ferrendelli, of  tissue  the potentiation  1983) o r n e u r o t o x i c  the vitamin.  activity  (Noell  Ferrendelli, with  e_t aJL. ,  1983) a l b e i t  amethopterin 1960;  (Olney itself  e t al_. , causes  and see a l s o  i n higher  doses  than  1981a) seizure  Clifford  and  are  required  an improbable  explan-  folate. A  ation  Indeed,  actions  and  of  direct depolarization the large  increases  i s also of  synaptic  excitation.  Thus  203  even w i t h folate  prolonged perfusions  AD  PSs were  orthodromic regions. also  reported  folate, with  a weak  to  structure produce  itself  death  5)  folic  acid al.,  uptake  by f o l a t e  hippocampal EPSP  action  either  tissue  neurons  by  i s i n agreement acids  moiety  or quisqualate However,  the e x c i t a b i l i t y n o t shown)  within  receptors  L-glutamate  changes  i n CAl  nor the pattern  presented  4)  (see  of  cell  1981).  for a  could  since  blockade  of  amino  uptake  as a mechanism  explain  the  that  However  L-glutamate  may  the amplitude  of the  gyrus  as might  Furthermore,  be  expected.  o r MTHF  Thus  on L - g l u t a m a t e  measurable a  of action  blockade  was  be a  i n CAl o r the dentate  has been  observations).  conceivably  neurochemical evidence (see  indicates  folinate  lished  precluded.  NMA  implied.  an a c t i o n  t o increase  hippocampal  be  neurotransmitter.  of folate,  have  i n c e r e b e l l u m ( R o b e r t s , 1974; R o b e r t s e t  Chapter  measured  observed  with  o f PS r e s p o n s e s  Introduction,  which  a glutamate  (Nadler et a l . ,  h a s been  1981) and such  enhancement  (1982)  of N - a c y l a t e d amino  contains  may  (data  by f o l a t e  Evidence  and McLennan  hippocampal  of r a tcentral  studies  acid  i t s effects  caused  (1973)  an i n t e r a c t i o n  o r CA3  depression of  1965).  reproduces neither  Chapter  direct  o r MTHF, a n o b s e r v a t i o n  and W a t k i n s ,  concentrations of  i n any of t h e  excitation  structure-activity  Because  and a  not observed  and Watkins  folinate  (Curtis  its  unaffected  PS was  Davies  of m i l l i m o l a r  f o r folate  no  uptake i n  (H. McLennan, of  never  unpub-  neurotransmitter would  seem  t o be  204  Since PSs  the actions of f o l a t e  and induce  duplicated events tion  spontaneous  with  by  folate  Antagonism cuneate  may  suggested  by  have  a blockade occurred  itory  mechanism  reduction On  of  discrepancy  presynaptic  has  been  literature. i n ther a t  folate  has  and Watkins  produced  been  (1973)  by  exogenous  On  the other  against a  disinhib-  neurons.  arguing  were  disinhibi-  inhibition  of i n h i b i t i o n  presented  of  i n the  1974) and D a v i e s  evidence  firing  GABA-mediated  the matter  on c a t c o r t i c a l  hand e l e c t r o p h y s i o l o g i c a l  orthodromic  epileptiform  iontophoretically-applied  and M i l l e r ,  that  GABA may  i s some  of GABAergic  (Hill  a  be p r o p o s e d .  there  nucleus  reported  and evoked  bicuculline  by f o l a t e  to potentiate  (Hill  et. a _ l . , 1 9 7 4 ) a n d  3 folates  affect  neither  uptake  (Roberts  H-GABA b i n d i n g  e_t a _ l . , 1 9 8 1 a n d H.  observations).  The  study  changes  assessing  consistent  (Ruck  with  results  McLennan,  of experiments  i n recurrent  t h e absence  e_t a l . , 1 9 8 0 ) n o r  of  any  unpublished  i n the present  inhibition  disinhibitory  are  also  effect  of  folate. In  view  of  curves  which  6.3B),  a mechanism  between  were  the shifts especially  dendritic  of folate  that  a  produced  evidence  f o r active  dendritic  by  t h e EPSP/PS  involving  as w e l l  depolarizing  as  an  and s p i k e  process  enkephalin  input/output  i n the dentate  possibility.  similar  tiations  campal  large  depolarization  soma seems a r e a s o n a b l e suggested  of  Lynch may  gyrus  increased coupling initiation  events  underlie  conduction  (Traub  i n the  e t a l . (1981)  i n C A l , and there passive  (Fig.  the  have  poten-  i s growing of  and L l i n a s ,  hippo1979;  205  Benardo  et  a_l. ,  1982).  Folate  dendritic  processes, although  currently  available.  The provide  insight  neuropathologies folate  striatum  with the  with (Olney  excitatory Olney's  the  much  kainate,  excitotoxic  vitamin direct  B  time  depolarization distant with  neurotoxic  actions  (see  there  above),  epileptiform system. the  doses  local appear  as o c c u r  both  i s  to  a  uptake  to the onset various was  and f o l a t e  i t s  consistent  cells  and  In  reduces However  suggests  that  rather  of  than  a  acid  appear  a  t o be  epilepticus  since  i t s convulsive  e_t aJL., 1 9 8 1 ) .  no  of kainate  t o be d u e  1981),  of status  reduces  there  i n the  death  et_ a l . , 1 9 7 1 ) .  by f o l i c  i s virtually  Unexpectedly  cell  i s involved.  produced  within  of  to the  effect  et. a l . ,  whether  activity  of  i n the neuropathology  t o determine  contributes  may  i n contrast  which  Purkinje  f o r this  of neurons,  (Fuller  i s  injections  ( R o b e r t s e_t a _ l . , 1 9 8 1 ) .  (Brennan  diazepam  not possible  no  observation  the induction  with  which  such  lesions  treatment  for  course  deficiency  correlated  folate,  destroy  f o r GABA  alterations  The  that  hypothesis (Olney  does  markers  1 2  an  for this  i n the patterns  intracerebral  causes  actions,  active  data of the present study  et_ a T . , 1 9 8 1 b ) w o u l d  longer  metabolic  is  The f a c t  c e r e b e l l u m MTHF  biochemical  by  these  evidence  the differences  produced  and k a i n a t e .  situation  weak  into  alter  no d i r e c t  electrophysiological  an  may  As  and  f o r kainate i t  diffusion process  of  folate,  i n the  brain  and maintenance  regions  considerable  of  the  limbic  variation  causing epileptiform  of  in  activity  206  in  vivo  (3 n m o l e s  (10~ -10~ 8  M  7  The  and  and  4  of  and  their  actions  on  change  observed  discrepancy binding  to  in  of  of  Indeed  to  the  and  the  MF-evoked  1°  in  CAl  e_t a _ l . ( 1 9 8 2 )  the  are with  caused and  This  reflect  receptors  shown  PS,  was  apparent  electrophysiological results  Longoni  folate  situation  responses.  kainic acid  the  processes  inhibition  evoked  that  kainate  synaptic  e_t a _ l . ( 1 9 8 0 ) may  of  doubt  i n v e s t i g a t i o n has  contrast  recurrent  the  Ruck  d i f f e r e n t types  regions.  l i t t l e  present  potentiate  depress  between  data  leave  excitatory  f o l a t e d i d not  detectable  never  the  However  jln v i t r o  M).  3  others  similar  kainate,  r e s p e c t i v e l y ) and  d i s t a n t c y t o t o x i c e f f e c t s of  comparable.  of  10~ -10~  broadly  that  no  50-100 nmoles  results  convulsive are  and  and  the  existence  in different brain  reported  that  compared  3 to  cerebellar  hippocampus folate. types,  tissue  was  which  powerful  neuronal  agonists  and  ization,  possibly  excitation. the  connection changed of  binds  kainate  triggers  evoked  kainate  e_t  nor  olfactory  two not  and  to  an  )  other  cortex.  the  binds  than of  considered  folate  kainate  found  blocked  which  are  binding.  that  folate  potentiating  the by  receptor  mediates  enhancement  of  inhibit  acid  f o l a t e and  changes  in  inhibition  kainic  another  actions  to  [ H]-kainate  w h i c h m u s t be  a _ l . ( 1982  responses  i n the  but  leading  proposal  its ability Auker  be  membrane  those  of  s e n s i t i v e to  may  excitations  Another  to  less  there  electrophysiological  unrelated  binding  1000-fold  Conceivably one  the  the both  depolarsynaptic is  that  entirely In  this  neither actions  207  Questions and of  folate folate  or a related  compound  especially  would  connection ly  Brennan  derivatives  from  potassium.  folate  levels  regard  e_t a _ l . ( 1 9 8 1 ) h a v e release  slices  of  of  Folate  fractions  al. ,  Finally,  1980 ). of  folate  diphenylhydantoin that  decline  of f o l a t e  (Bridgers i t has  long  are reduced  and  the therapeutic  present  remains  and  with  and  by  known  "partial-  i t s  controversial  reduced  elevated  brain  (Miller (Goff  1972; that  however  and  et_ a l . ,  Carl  serum  anticonvulsants  of these drugs  this  concentrations  McClain,  been  a  activity  i n high  In  of  depolarization  to kindling  phenobarbitone,  effects  etiology  demonstrated  animals  1978).  synaptosomal  kainate  investigation.  are predisposed to seizure  i s also  the  r a t c e r e b e l l u m by  Furthermore  of  of the involvement  folate  1975) and a r e more amenable  levels  to  further  Webster,  CSF  homologies  i n the modulation of neuronal  with  seem t o w a r r a n t  calcium-dependent"  with  the receptor  notwithstanding, the p o s s i b i l i t y  excitability epilepsy  regarding  such  i n e_t and as  the hypothesis  i s related  (Reynolds,  to  1972).  this  208  References A L D I N I O , C , F R E N C H , E.D. & SCHWARCZ, R. 1 9 8 3 . 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(left  excitants  L-aspartic  acid,  are  acids  the  from acid,  ibotenic antagonists  2-amino-4-phosphonobutyric  2-amino-5-phosphonovaleric ester  The  acid,  quisqualic  same  amino  column).  L-glutamatic  In  acid  of  (APV), (DGG)  acid Dacid  glutamate and  piperi-  


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