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The peripheral vascular effects and supraspinal control of a spinal cardiovascular reflex Szeto, Peter Ming 1975

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THE PERIPHERAL VASCULAR EFFECTS AND SUPRASPINAL CONTROL OF A SPINAL CARDIOVASCULAR REFLEX hy PETER MING SZETO B.Sc., University of B r i t i s h Columbia 1 9 7 4 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF "THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE i n the Department of PHYSIOLOGY We accept t h i s thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA September, 1 9 7 5 i In present ing th is thes is in p a r t i a l fu l f i lment of the requirements for an advanced degree at the Un ivers i ty of B r i t i s h Columbia, I agree that the L ibrary sha l l make it f r ee ly ava i l ab le for reference and study. I fur ther agree that permission for extensive copying of th is thes is for s c h o l a r l y purposes may be granted by the Head of my Department or by h is representa t ives . It is understood that copying or p u b l i c a t i o n of th is thes is for f i n a n c i a l gain sha l l not be allowed without my wri t ten permission. Peter M. Szeto. Department of Physiology The Un ivers i ty of B r i t i s h Columbia 2075 Wesbrook Place Vancouver, Canada V6T 1W5 Date October 1/1975 ABSTRACT It has been reported that a mechanical stretch of the walls of the thoracic aorta without obstruction of a o r t i c blood flow induced s i g n i f i c a n t increases i n a r t e r i a l blood pressure, heart rate and cardiac contract-i l i t y (indicated by LV dP/dt max), and that these r e f l e x hemodynamic changes were present i n both i n t a c t and spinal cats (Lioy et a l , 1974)• We decided therefore to study (1) the r e l a t i v e contribution of the external i l i a c , the superior mesenteric and the renal vascular beds to the pressor response of thi s spinal thoracic: aorta r e f l e x (TAR), and (2) the effects of the carotid baroreceptor and chemoreceptor reflexes on the TAR pressor response. Experiments were carried out i n adult cats with i n t a c t brain-stem and anesthetized with chloralose-urethan. The c e r v i c a l vagi were cut and the common carotids were occluded or the carotid sinuses denervated. The animals were a r t i f i c a l l y v e n t i l a t e d and the blood gases and pH were kept within physiological l i m i t s . i i (1) By measuring blood flows in the external i l i a c , superior mesenteric and renal arteries using ultrasonic Doppler flowmeters, we found that aortic stretch (AS) induced, in a l l the vascular beds a vasoconstriction which could be prevented by the administration of an alpha-adrenergic blocker (phenoxybenzamine). The increase in vascular resistance was most marked in the superior mesenteric bed, and least in the renal bed. The lat ter maintained a constant flow in the presence of a higher perfusion pressure, probably because of autoregulation. After alpha-adrenergic blockade, the external i l i a c vasoconstriction was reversed to vasodilatation that could be blocked by atropine. The poss ibi l i ty that AS reflexly activates the sympathetic cholinergic vaso-dilator system is suggested. (2) The baroreceptor and chemoreceptor reflexes were activated by stimulating the carotid sinus nerve or by perfusing the sinus. We found that both these supraspinal reflexes inhibit but do not abolish the pressor effects of the TAR. The possible mechanisms of these inhibitions and the implications of these results on the nervous control of circulation are discussed. i i i TABLE OF CONTENTS Page ABSTRACT i i LIST OP TABLES v i i i LIST OF FIGURES x ACKNOWLEDGMENTS . x i i i PART I. INTRODUCTION 1-13 SPINAL CARDIOVASCULAR REFLEXES 1 THE SPINAL THORACIC AORTA REFLEX 10 AIMS OF THE PRESENT STUDY 12 PART II. METHODS OF STUDY 14-33 EXPERIMENTAL GROUPS 14 BASIC PREPARATIONS 16-23 Anesthesia 16 Controlled Respiration 17 Body Temperature Regulation 18 P0 2, PC0 2 and pH Analysis 18 Thoracic Aorta Cannulation 19 Blood Pressure Measurements 22 SPECIAL PREPARATIONS 24-33 Blood Flow Measurements 24 Vascular Resistance Calculation 25 Pharmacological Blockade 26 iv Page Carotid Sinus Nerve Stimulation 27 Carotid Sinus Perfusion 28 Carotid Baroreceptor Activation 31 Carotid Chemoreceptor Activation 31 Norepinephrine Infusion and 32 Change of Blood Volume EXPERIMENTAL PROTOCOL DATA ANALYSIS 34 36 PART III. RESULTS 37-83 REFLEX CHANGES IN ARTERIAL PRESSURE 37-45 Control 37 Pharmacological Blockade 41 Effects on the TAR Response 41 REFLEX CHANGES IN ARTERIAL FLOWS 46-62 Control 46 Pharmacological Blockade 47 Effects on Reflex Changes of Blood Flow 60 THE BARORECEPTOR REFLEX & THE TAR 63-71 Effects of Carotid Occlusion 63 and Sinus Denervation Carotid Baroreceptor Stimulation 66 Effects on the TAR Response 66 v THE CHEMORECEPTOR REFLEX & TEE TAR 72-77b Carotid Chemoreceptor Stimulation 72 Effects on the TAR Response 73 EFFECTS OF PREVAILING ARTERIAL 78-83 PRESSURE Lower Blood Pressure 78 Higher Blood Pressure 79 PART IV. DISCUSSION 84-119 REFLEX CHANGES IN ARTERIAL 84-103 PRESSURE AND FLOWS The Doppler Flowmeter Method 84 The External I l iac Flow (IF) 85 The Superior Mesenteric Flow (SMF) 96 The Renal Flow (RF) 100 THE CAROTID BARORECEPTOR REFLEX 104-111 & THE" TAR Effects of Carotid Occlusion 104 and Sinus Denervation Inhibitory Effects on the Carotid 105 Baroreceptor Reflex on the TAR v i Page THE CAROTID CHEMORECEPTOR REFLEX 112-119 & THE TAR Methods of Chemoreceptor Activation 112 Inhibitory Effects of the Carotid 116 Chemoreceptor Reflex on the TAR CONCLUSIONS 1 2 0 BIBLIOGRAPHY 122-138 APPENDICES 1 5 g v i i LIST OP TABLES  Table Page I E f f e c t s of Phenoxybenzamine, Propranolol, 39 and Atropine on the Reflex Pressure . Changes induced by Stretching the Walls of the Thoracic Aorta. I I E f f e c t s of Phenoxybenzamine, Propranolol, 48 and Atropine on the Reflex Changes i n Blood Plow and Vascular Resistance i n the External I l i a c Artery induced by • • ' A o r t i a Stretch. I l l E f f e c t s of Phenoxybenzamine, Propranolol, 50 and Atropine on the Reflex Changes i n Blood Plow and Vascular Resistance i n the Superior Mesenteric Artery induced by A o r t i c Stretch. IV E f f e c t s of Phenoxybenzamine, Prop r a n o l o l , 52 and Atropine on the Reflex Changes- i n Blood Plow and Vascular Resistance i n the Renal Artery induced by A o r t i c S t r e t c h . V E f f e c t s of A o r t i c Stretch on Blood Plows 58 and Vascular Resistance i n the Intact and Skinned Legs. v i i i Table Page VI E f f e c t s of Carotid Baroreceptor 64 Stimulation on the r e f l e x pressor response induced by A o r t i c Stretch. VII E f f e c t s of Carotid Chemoreceptor 76 Stimulation on the r e f l e x pressor response induced by A o r t i c Stretch. VIII E f f e c t s of Low and High Basal Blood 82 Pressure on the r e f l e x pressor response induced*by Aorti c Stretch. i x LIST OF FIGURES Figure Page H 1 Schematic representation of the 21 s p e c i a l cannula used to stretch the walls of the thoracic aorta without obstructing a o r t i c blood flow. 2 Schematic reprenentation of the 30 perfusion system used to activate the carotid baroreceptors and chemoreceptors. 3 Reflex changes i n a r t e r i a l pressure 38 induced by st r e t c h i n g the walls of the thoracic aorta. 4 Reflex changes i n blood flow i n the 43 external i l i a c a r t e r i e s of the i n t a c t and skinned legs induced by a o r t i c s t r e t c h . 5 E f f e c t s of a o r t i c s t r e t c h on blood 45 flows i n the renal and the superior mesenteric a r t e r i e s . x The time-related changes i n vascular resistance during the period of a o r t i c s t r e t c h under normal conditions and under pharmacological blockade. Correlations between changes i n vascular resistance and i n a r t e r i a l pressure induced by a o r t i c s t r e t c h . Reflex cholinergic vasodilatation i n the external i l i a c vascular bed induced by a o r t i c s t r e t c h . E f f e c t s of stimulating the central end of the carotid sinus nerve on the pressor response induced by a o r t i c s t r e t c h . E f f e c t — o f increasing the c a r o t i d sinus perfusion pressure on the pressor response induced by a o r t i c s t r e t c h . Stimulus frequency—Response curve (O) f o r the baroreceptor f i b r e s i n the car-o t i d sinus nerve and the graded supp-ression on the pressor response ) induced by a o r t i c s t r e t c h . x i Figure Page 12 Stimulus-Response relationships f o r 71 the c a r o t i d haroreceptors i n two groups of animals and the graded suppression on the pressor response induced by-a o r t i c s t r e t c h . 13 E f f e c t s of stimulating the central end 74 of the c a r o t i d sinus nerve with short duration (0.07 msec) pulses on the pressor response induced by a o r t i c s t r e t c h . 14 E f f e c t s of c a r o t i d chemoreceptor 75 stimulation by cross-perfusion of the sinus on the pressor response induced by a o r t i c s t r e t c h . 15 Suppression of the TAR response by 77b chemoreceptor stimulations, and by higher control blood pressures. 16 E f f e c t s of a higher basal blood pressure 80 on the pressor response induced by a o r t i c stretch (norepinephrine i n f u s i o n ) . 17 E f f e c t s of a higher basal blood pressure 81 on the pressor response induced by a o r t i c stretch (blood i n j e c t i o n ) . x i i ACKNOWLEDGMENTS I would l i k e to express my gratitude and appreciation to Dr. Franco Lioy f o r h i s guidance i n s t r u c t i o n and h e l p f u l c r i t i c i s m i n th i s work, and my thanks to Miss Susan Powell and Mr. K. Henz f o r t h e i r able assistance. x i i i 1 PART I. INTRODUCTION SPINAL CARDIOVASCULAR REFLEXES This thesis i s concerned with one s p i n a l c a r d i o -vascular (CV) r e f l e x e l i c i t e d from the thoracic aorta. A s p i n a l CV r e f l e x can he considered a 3 a c i r c u l a t o r y control mechanism which i s subserved by an afferent nervous pathway coming from sensory receptors outside or within the CV system and projecting to the sp i n a l cord, and by an efferent nervous pathway from the cord to target organs of the c i r c u l a t o r y system. I t i s a s p i n a l mechanism because the r e f l e x response i s mediated at the spi n a l l e v e l and i s operative without any neuronal connection with supraspinal structures. Experimental evidence f o r the presence of s p i n a l CV reflexes i s a v a i l a b l e : Goltz ( 1 8 7 4 ) noticed that i n spinal dogs, a f t e r the a r t e r i a l pressure had returned to a normal l e v e l from spin a l shock, further destruction of the cord reduced t h i s pressure to or below the l e v e l character-i s t i c of spinal shock and precluded a l l remaining CV r e f l e x changes. Sherrington ( 1 9 0 6 ) observed that following 2 a c e r v i c a l section of the cord, vascular tone and pressor reflexes e l i c i t e d by stimulation of somatic nerves were l o s t at f i r s t , but reappeared a f t e r a short time. In acutely prepared s p i n a l animals, s u b s t a n t i a l increases i n systemic blood pressure were obtained by e x c i t a t i o n of somatic nerves (Langley, 1924; Brooks, 1933), and the responses were potentiated a f t e r intravenous i n j e c t -ion of strychnine (Langley, 1924). Similar vasomotor reactions were also produced i n s p i n a l animals by mechanical or noxious stimulation of the skin (Sahs & Fu l t o n , 1940; Kuntz, 1945; Eichins & Brizzee, 1949) and of the v i s c e r a (Downman & McSwiney, 1946; Mukherjee, 1957). Kuntz- & Hazelwood (1940) demonstrated that some of the pressor responses to somatic nerves stimulation were dependent upon the i n t e g r i t y of the sympathetic efferent supply to the c i r c u l a t o r y system; Alexander (1946) and Schaeffer ( i 9 6 0 ) , however, observed that following acute or chronic s p i n a l section, the change i n sympathetic a c t i v i t y evoked by stimulation of somatic nerve was abolished. In contrast, ^  Koizumi & Suda (1963) were able to record sympathetic s p i n a l r e f l e x discharge induced by stimulation of somatic nerves. In a s i m i l a r but more extensive study, Beacham & P e r l (1964) 3 were also able to record background discharge from some, but not a l l , preganglionic f i b r e s at each of the s p i n a l l e v e l s (T1 to L4) i n acutely decapitate cats. These workers also showed that the background discharge could be f a c i l i t a t e d or i n h i b i t e d by noxious, thermal and mechanical s t i m u l i , as well as by e l e c t r i c a l stimulation of dorsal roots, s p i n a l nerves and limb nerves. Fernandez de Molina & P e r l ( 1 9 ^ 5 ) then demonstrated that some of the s t i m u l i which r e f l e x l y excited preganglionic neurons also evoked transient increases i n systemic a r t e r i a l pressure, and/or alt e r e d blood flows i n the femoral, i n f e r i o r mesenteric and br a c h i a l a r t e r i e s . The above CV reflexes e l i c i t e d from stimulation of somatic afferents are also referred to as sp i n a l somato— sympathetic r e f l e x e s . They have been extensively studied (see — Koizumi & Brooks, 1 9 7 2 ; Sato & Schmidt, 1 9 7 3 ) , and a l l the r e s u l t s from these studies indicate that the i s o l a t e d thoracolumbar spin a l segments are capable not only of i n i t i a t i n g a tonic discharge but also of mediating r e f l e x a l t e r a t i o n of t h i s spontaneous a c t i v i t y ; although 1 the functional s i g n i f i c a n c e of the spinal sympathetic reflexes under normal conditions i s not known' (Koizumi & Brooks, 1 9 7 2 ) . 4 Spinal CV reflexes e l i c i t e d from receptors within the CV system were studied by Heymans and h i s associates. They described vasomotor reflexes produced by v a r i a t i o n s of blood pressure i n the mesenteric vessels of s p i n a l dogs (Heymans, Bouckaert, Parber & Hsu, 1 9 3 6 ) , and found that the nerve endings responsible f o r these reflexes were situated i n the mesenteric vessels (Heymans, Bouckaert & Vierzuchowski, 1 9 3 7 ) . Heymans et a l ( 1 9 3 6 ) pointed out that ... These spin a l vasomotor reflexes which adapt vascular tone to the general blood pressure, however, do not appear to play an important part i n the auto-matic and proprioceptive regulation of the general blood pressure, as do the vasosensitive zones i n the a o r t i c arch and the c a r o t i d sinuses... I t i s probable that the s p i n a l vasomotor reflexes under discussion play a r o l e i n the d i s t r i b u t i o n of blood i n the deep abdominal c i r c u l a t i o n and i n the periphery... and i n a minor and accessory manner, may j o i n with the vascular zones i n the a o r t i c arch and the c a r o t i d sinuses i n the regulation of the c i r c u l a t i o n and the general blood pressure. These workers also suggested that other vascular areas such as the thoracic may 'possess r e f l e x s e n s i t i v i t y to endo-vascular pressure*. 5 In 1 9 5 4 , Gruhzit, Freyburger & Moe studied the nature of the frequently observed r e f l e x v a s o d i l a t a t i o n i n s k e l e t a l muscle of the l e g induced by intravenous i n j e c t i o n of epinephrine, and found that, i n baroreceptor deafferented animals, the femoral v a s o d i l a t o r response to the sympathomimetic was diminished by the a p p l i c a t i o n of a l o c a l anesthetic around the aorta, while the response to affe r e n t vagal stimulation was not a l t e r e d . Gruhzit et a l ( 1 9 5 4 ) then i s o l a t e d one of the hind legs of the dog and perfused i t from a donor animal. They demonstrated that occlusion of the aorta by a l i g a t u r e above the diaphragm resulted i n an increase of femoral blood flow i n the perfused l e g . Next, they compared the e f f e c t of epinephrine upon femoral blood flow i n the perfused l e g before and a f t e r clamping the descending aorta d i s t a l to the l e f t subclavian artery, and found*that the d i l a t o r r e f l e x was greatly reduced or completely prevented when the drug could not reach the thoracic aorta. F i n a l l y , Gruhzit et a l ( 1 9 5 4 ) cannulated the aorta d i s t a l to the l e f t subclavian artery and t i e d i t just above the diaphragm, and perfused the thoracic aorta with blood from a r e s e r v o i r . The perfusate apparently entered the animal v i a thoracic a r t e r i e s . Except f o r the r i g h t l e g which was again cross-perfused, a l l the sub-diaphragmatic organs were supplied with blood from a by-pass 6 connecting the l e f t subclavian to the l e f t i l i a c a r t e r y . When the perfusion pressure within the thoracic aorta was increased, they observed vasodilatation i n the r i g h t femoral vascular bed, although of a smaller magnitude than that induced by epinephrine. Gruhzit et a l ( 1 9 5 4 ) therefore concluded that there 'may be mechanoreceptors located i n the wall of the aorta or i t s thoracic branches or i n the tissues surrounding these vessels' which were responsible f o r the r e f l e x v a s o d i l a t a t i o n i n the l e g . Although t h e i r study was done i n dogs with i n t a c t central nervous system (CNS), Gruhzit et a l ( 1 9 5 4 ) suggested that * i t i s possible that the response i s mediated i n the cord i t s e l f rather than at higher l e v e l s . ' The nature of the 'receptors' was studied by Leitner & P e r l ( 1 9 6 4 ) . They made a systemic survey of afferent f i b r e s entering the thoracic l e v e l s (T3 to T12) of the spina l cord which were responsive to c i r c u l a t o r y changes. These workers found that some dorsal root f i b r e s discharged synchronously with cardiac systole, p a r t i c u l a r l y a f t e r the intravenous i n j e c t i o n of adrenaline (epinephrine). They observed also that the rate of discharge i n these f i b r e s was rela t e d to the amplitude of the pulse pressure and was not affected by changes i n d i a s t o l i c or mean pressure. 7 These workers then recorded afferent a c t i v i t y of mesenteric nerves coming from Pacinian corpuscles i n the cat's mesentery, and found that both intravenous and i n t r a -a r t e r i a l (close to the corpuscle) in j e c t i o n s of epine-phrine induced s i m i l a r increases i n afferent discharge. Leitner & P e r l (1964) therefore concluded that 'the response i n association with the cardiac cycle was i n i t i a t e d by a mechanical transient related to the pulse wave i n l a r g e r blood vessels and came from receptors s i m i l a r to Pacinian or paciniform corpuscles.* These workers also suggested that *intravascularly i n j e c t e d adrenaline may a l t e r the s e n s i t i v i t y of this type of receptor.* After Leitner & P e r l (1964)» the afferent pathway of these spin a l reflexes were studied by several groups of workers. In 1967* Brown reported that coronary occlusion i n the cat activated a large contingent of afferent f i b e r s , some of which were i n the cardiac sympathetic nerves entering the spinal cord v i a the upper f i v e dorsal roots. Other workers subsequently confirmed the presence of these •afferent cardiac sympathetic f i b r e s ' and showed that mechanical as well as chemical s t i m u l i were capable of modifying the spontaneous discharge of this type of f i b r e s , whose mechanosensitive endings were located i n the p e r i -8 cardium and the thoracic aorta (TJeda, Uchida &. Kamisaka, 1969) t "the main coronary artery (Brown & M a l l i a n i , 1971; M a l l i a n i , Recordati, Schwartz & Pagani, 1972), the ve n t r i c l e s and the a t r i a ( M a l l i a n i , Recordati & Schwartz, 1973) and the pulmonary artery ( N i s h i , Sakanashi & Takenaka, 1974)* Furthermore, e l e c t r i c a l stimulation of these 'afferent cardiac sympathetic f i b r e s ' induced i n spinal animals, a r e f l e x r i s e i n blood pressure which could be abolished by phenoxybenzamine (Peterson & Brown, 1971)» and a s l i g h t but s i g n i f i c a n t increase i n LV dP/dt max (maximum rate of r i s e of l e f t v e n t r i c u l a r pressure) ( M a l l i a n i , Peterson, Bishop & Brown, 1972), and a s i g n i -f i c a n t increase i n heart rate ( M a l l i a n i , Parks, Tuckett & Brown, 1973). The efferent nervous pathway of these s p i n a l reflexes was studied by M a l l i a n i , Schwartz & Zanchetti (19^9). They confirmed the previous observations of Beacham & P e r l (1964) that spontaneous a c t i v i t y , c o n s i s t i n g of a few spikes/second, could be recorded from single pre-ganglionic sympathetic f i b r e s i n spinal cats. M a l l i a n i et a l (1969) then demonstrated that occlusion of the coronary artery caused e i t h e r an increase or a decrease i n sympathetic discharge. Similar responses were obtained when the 9 systemic a r t e r i a l pressure was increased ei t h e r "by occluding the aorta at various s i t e s or by i n j e c t i n g pressor drugs (angiotensin and norepinephrine) i n spinal cats ( M a l l i a n i , Pagani, Recordati & Schwartz, 1971a). These workers observed that a given single f i b r e consistently responded to r i s e s i n a r t e r i a l pressure e i t h e r by decreasing or increasing i t s f i r i n g rate but never i n both d i r e c t i o n s , and the majority of the preganglionic sympathetic f i b r e s responded by decreasing t h e i r discharge. M a l l i a n i , Pagani, Recordati & Schwartz (1971b) therefore proposed the presence of spina l sympathetic reflexes e l i c i t e d by coronary artery occlusion and by blood pressure changes. The afferent and efferent pathways of these reflexes were both i n the sympathetic system. The above experiments have been c r i t i c i z e d because the s t i m u l i used to activate these reflexes could hardly be considered as ph y s i o l o g i c a l . The use of the term 'afferent sympathetic f i b r e s * has also been opposed, since i t i s generally accepted that 'the sympathetic nervous system consists by d e f i n i t i o n , of a pure outflow' ( M a l l i a n i , Lombardi, Pagani, Recordati & Schwartz, 1975). The second objection appears to be an argument over a semantic problem, 10 and c e r t a i n l y does not i n i t s e l f over-rule or deny the existence of afferent f i b r e s i n nerves which are anatomi-c a l l y located i n the sympathetic system. Thus, the above experiments have provided these . information on spinal CV re f l e x e s : a) the CV receptors involved are possibly 'mechanoreceptors 1 located i n d i f f u s e areas of the c i r c u l a t o r y system; b) one of the s t i m u l i capable of a f f e c t i n g these receptors i s endovascular pressure; c) the afferent and efferent nervous pathways are i n the sympathetic system, and d) these reflexes are integrated at the spinal l e v e l . THE SPINAL THORACIC AORTA REFLEX I t remained to be demonstrated that sp i n a l r e f l e x hemodynamic changes could be induced by a mechanical stimulus which would not a f f e c t the very c i r c u l a t o r y functions to be studied. This problem was solved by Lioy, M a l l i a n i , Pagani, Recordati & Schwartz ( .1974). These workers devised a sp e c i a l cannula which could be used to stret c h the walls of the thoracic aorta without obstructing a o r t i c 11 blood flow. Since the same type of cannula i s to be used i n the present study, i t s detailed description i s given i n the section of METHODS OF STUDY and i s not repeated here. Lioy et a l (1974) found that by s t r e t c h i n g the aorta between T7 and T 1 0 , s i g n i f i c a n t increases i n a r t e r i a l blood pressure, heart rate and LV dP/dt max were observed i n cats with i n t a c t CNS and with spi n a l transection at C 1 . They further demonstrated that these responses were abolished by i n f i l t r a t i n g the walls of the thoracic aorta with xylocaine, while CV responses to a strong nociceptive stimulus (clamping of the paw) were not a l t e r e d . These workers also assessed the contributions of cardiac and peripheral vascular mechanisms to the r e f l e x response by carrying out adrenergic blockade and cardiac denervation, and provided i n d i r e c t evidence that the adrenal glands were probably activated by t h i s spinal r e f l e x . Lioy et a l (1974) found that a s t r e t c h causing a 10% increase i n a o r t i c diameter was s u f f i c i e n t to e l i c i t a clear r e f l e x response, t h i s amount of stretch was equivalent to that induced by an a o r t i c pressure increase of 5 0 - 6 0 mmHg and therefore considered to be within the physiological range (Peterson, 1 9 6 2 ) . 12 As suggested by Lioy et a l (1974). t h i s spinal CV r e f l e x appears to operate by a p o s i t i v e feedback mechanism, i . e . , a stimulus l i k e l y to duplicate the eff e c t s of an increase i n a o r t i c blood pressure produces a further increase i n blood pressure, heart rate and LV dP/dt max. However, the r e s u l t s of t h i s p o s i t i v e feedback mechanism are not necessarily detrimental, since Pagani, Schwartz, Bishop & M a l l i a n i (1974) have shown that the thoracic aorta i s also a target organ f o r t h i s kind of spina l r e f l e x e s . They have demonstrated that stimulation of afferent sympathetic nerve f i b e r s reduces the compliance of the thoracic aorta; therefore stretching of the aorta may induce a feedback mechanism which re s t r a i n s any further d i s t e n t i o n of the same ve s s e l . This protective process i s p h y s i o l o g i c a l l y s i g n i f i c a n t , since according to one of the theories of atherogenesis, a t h e r o s c l e r o t i c lesions can develop as a re s u l t of l o c a l increases i n l a t e r a l ( s t a t i c ) wall pressure which may injure the intima or lead to l o c a l l i p i d accumulation within the intima, suppression of l i p i d secretion from within the a r t e r i a l w a l l , or both (Vesolowski, Sabini & Sawyer, 19&5)« 13a Pagani, Schwartz, Banks, Lombardi & M a l l i a n i (1974) made use of the specia l cannula, and recorded the a c t i v i t y of sing l e sympathetic preganglionic f i b r e s (T3-T4) i n spinal cats. They found i n 18 f i b r e s that when the thoracic aorta was stretched, the rate of discharge increased i n s i r , decreased i n ten and did not change i n two, Thus, the experiments of Lioy et a l (1974) demonstrated that stretching the walls of the thoracic aorta can induce CV r e f l e x responses i n spina l preparations. In t h i s t h e s i s , we s h a l l r e f e r to t h i s r e f l e x as the spinal thoracic aorta r e f l e x or the TAR. AIMS OF THE PRESENT STUDY I t i s known then that stimulation of receptors located i n the walls of the thoracic aorta e l i c i t e s a pressor response due to active vasoconstriction i n some unknown vascular bed3 (Lioy et a l , 1974), and probably also induce* some vaso d i l a t a t i o n i n the muscular bed of the l e g (Gruhzit et a l , 1954). We decided therefore to measure blood flows i n the external i l i a c , 13b the superior mesenteric and the renal a r t e r i e s before, during and a f t e r the TAR, so as to assess q u a n t i t a t i v e l y the contributions of these vascular beds to the r e f l e x response. Ve have also studied the nature of these vasomotor changes by pharmacological blockade. Since the TAR i s a spinal r e f l e x , an understand-i n g of the i n t e r r e l a t i o n s h i p s between t h i s s p i n a l mechanism and some known supraspinal mechanisms would contribute to our knowledge concerning r e f l e x control of CV a c t i v i t i e s . Ve decided therefore to study the ef f e c t s on the TAR of two well—known supraspinal r e f l e x e s , namely the c a r o t i d baroreceptor and chemoreceptor r e f l e x e s . 14 PART II . METHODS OF STUDY EXPERIMENTAL GROUPS A total of 58 cats of either sex with "body weight "between 1,88-4.70 kg were used i n the experiments. The animals were divided into four major groups according to the objectives of the present study. (I) in 22 animals, the contribution of different vascular beds to the spinal thoracic aorta reflex (TAR) were assessed. The blood flows in the external i l i a c , the superior mesenteric, and the renal arteries were measured during an aortic stretch under normal circum-stances and under pharmacological blockade of the adren-ergic and cholinergic systems. (II) in 15 animals, the effects of carotid baro-receptor activation on the TAR were studied. The baro-receptor reflex was activated either by stimulating the carotid sinus nerve (CSN), or by changing the sinus perfusion pressure (CSP). 15 (III) i n 12 animals, the e f f e c t s of c a r o t i d chemo-receptor a c t i v a t i o n on the TAR were studied. The chemo-receptor r e f l e x was evoked by stimulating the CSN, or by perfusing the sinus with hypoxic, hypercapnic and a c i d o t i c blood from donor animals. (IV) i n 1 0 cats, the influences of p r e v a i l i n g a r t e r i a l pressure on the TAR were studied. In these animals, the TAR was induced before and a f t e r changing the basal a r t e r i a l pressure e i t h e r by an i n f u s i o n of norepinephrine, or by a change i n the blood volume of the animals. 16 BASIC PREPARATIONS The following general procedures were performed i n a l l of the animals used in the present study: Anesthesia The cats were anesthetized with an intraper i -toneal ( i .p . ) injection of urethan and alpha-chloralose at 25O and 60 mg/kg respectively. The anesthetic was prepared "by dissolving 2000 mg of urethan in 40 ml of d i s t i l l e d water, then adding 480 mg of alpha—chloralose. The solution was then heated to about 65 -75 °C for alpha-chloralose to dissolve, and was f i l t e red . The f ina l concentrations of urethan and alpha-chloralose in the solution were 50 and 12 mg/ml respectively. The solution was kept at about 65 °C i n a water bath before inject ions . Complete anesthesia was usually induced within 30 min after the i n i t i a l in ject ion. One vein (usually the femoral, but in some blood flow measurement experiments, the brachial was chosen) was always cannulated with polyethylene tubing (Incore Electro-Plast ics , SLV 105, #20 or 22). for 17 la ter Injection of drugs or supplementary doses of anes-thetic whenever necessary. Controlled Respiration The vago-sympathetic trunks and the depressor nerves on both sides of the neck were isolated, l igated and sectioned. (These were not performed in the six donor animals in the chemoreceptor cross-perfusion experiments). A tracheotomy was performed in a l l cats, and after an intravenous ( i . v . ) injection of a paralyzing dose ( 2 . 5 mg/kg) of gallamine triethiodide (Flaxedil ) , positive-pressure respiration was in i t i a ted using a respirator pump (Harvard Apparatus, Model 614) connected to the tracheal cannula. The respirator drew gas through a demand valve from a cylinder of compressed 100% 0^ (This cylinder could be easily replaced by another cylinder of a different gas mixture, as required in the chemoreceptor cross-perfusion experiments — see below)• The 100% 0^ gas was then mixed with room a i r in an elast ic compensator to y ie ld a high P0 2 gas of approximately 5 0 % 0 2 and 5 0 % Ngi *> e ; f o r © pumped to ventilate the animal. 18 The volume (about 100 ml) and rate (about 20 strokes/min at an output r a t i o of Inspiration/Expiration = 40/60) of v e n t i l a t i o n were adjusted to maintain a r t e r i a l gases and pH within physiological l i m i t s (P0 2 135+15 mmHg, PC0 2 24+4 mmHg, pH 7.40+0.05). In case of acid-base inbalance i n the a r t e r i a l blood, i t was corrected by i . v . infu s i o n of sodium bicarbonate solution (1 mEq/ml). . Body Temperature Regulation The body temperature of the animal was measured by an esophageal thermal probe and maintained at 37.5 °C by a feedback heating u n i t (Yellow Springs Instrument, Model 73TG-). P 0 2 < PC0 2 and pH Analysis End t i d a l P C ^ i n the tracheal cannula was continuously monitored by an assembly (Beckman Instruments, Model LB-1) which included a microcatheter sample pump, a CO^ pickup u n i t and an in f r a r e d analyzer. For blood gases and pH analysis, samples of 1.5-2 ml were c o l l e c t e d as often as required, or at l e a s t once every hour. The samples were then analysed at 37 °C i n a blood PO^, 1 9 PCO^ and pH analyzer (Instrumentation Laboratory, Model 113). Correction factors were allowed f o r any d i s c r e -pancy between th i s temperature (37 °C) and the body temperature at the time when the blood sample was taken. The a r t e r i a l cannula from which the blood sample had been co l l e c t e d was always flushed with i s o t o n i c s a l i n e , and the l o s t blood volume was sometimes replaced with Dextran sol u t i o n . Thoracic Aorta Cannulation For thoracic aorta cannulation, the animals were placed on t h e i r r i g h t side, and the skin and muscles over the l e f t hemithorax were dissected using an e l e c t r o -s u r g i c a l cutter/coagulator (Birtcher, Model 755) to expose the r i b s . Each r i b , fourth to tenth i n c l u s i v e , was l i g a t e d on both the ventral and dorsal ends and removed. The cut ends were sealed with bone wax to stop any bleeding. A wide pneumothorax was thus produced. Two loose l i g a t u r e s were then placed around the aorta, one caudal to the l e f t subclavian artery, another r o s t r a l to the diaphragm. The r i g h t p l e u r a l cavity close to the diaphragm was also exposed by making a hole i n the mediastinal pleurae, and 20 a t h i r d loose l i g a t u r e was placed around the i n f e r i o r vena cava. A special cannula ( F i g . 1) was used to s t r e t c h the walls of the thoracic aorta without obstructing a o r t i c blood flow. I t consisted of a s t a i n l e s s s t e e l tube surrounded by a thin rubber cylinder which could be i n f l a t e d v i a another small metal tube mounted perpendicular to the tube close to one end of the cannula. A l l cats, except the s i x donor animals used i n the chemoreceptor cross-perfusion experiments, were cannulated with t h i s a o r t i c cannula, which came i n d i f f e r e n t dimensions (5-7 cm long, 3-5 nun, i.d.)-, and was selected according to the body size of the cat so that i t f i t snugly into the aorta. After the animal was heparinized (3 mg/kg), the loose ligatures around the i n f e r i o r vena cava and the aorta were pulled to minimize blood l o s s , the aorta was completely divided below the tenth i n t e r c o s t a l artery and the cannula inserted. The two cut ends of the aorta were l i g a t e d around the lower end of the cannula, with the side tube passing out between them. The l i g a t u r e s on the i n f e r i o r vena cava and the aorta were then released, and normal c i r c u l a t i o n resumed. The cannulation procedure 21 Figure 1. Schematic representation of the s p e c i a l cannula used to s t r e t c h the walls of the thoracic aorta without obstructing a o r t i c blood flow. 22 took between 1 and 1.5 min. The pressure drop across the cannula measured by comparing the c a r o t i d and femoral a r t e r i a l mean pressures was between 5-15 mmHg. When the ballon on the cannula was i n f l a t e d with 0.6-1.3 m l of i s o t o n i c saline from a 2.5 ml syringe, the a o r t i c walls were stretched between the seventh and the tenth i n t e r -costal a r t e r i e s , thus evoking the spinal thoracic aorta r e f l e x (TAR). Blood Pressure Measurements In the experimental animals, a r t e r i a l pressure was always measured both above and below the a o r t i c cannula to ensure that a o r t i c blood flow had not been obstructed. In most cats, the pressure above the cannula was measured i n the carotid artery (CAP), while the pressure below the cannula was measured i n the femoral artery (FAP). The only exception was the group of cats i n which the external i l i a c blood flows to both hind limbs were assessed simultaneously. In these animals, the pressure below the a o r t i c cannula was measured i n the i n f e r i o r mesenteric artery (IMP). In the donors of the chemoreceptor cross-perfusion experiments, only the FAP was measured. 23 A l l a r t e r i a l pressures and perfusion pressure were measured by s t r a i n gauges (Statham, P 2 3 D b ), which were connected to a transducer/converter (S.E. Labor-a t o r i e s , Type SE905) and to a d i r e c t - p r i n t , u l t r a - v i o l e t l i g h t (UV) recorder (S.E. Laboratories, Type SE3006). The pressure measurement system was ca l i b r a t e d by a mercury manometer at the beginning of each experiment, and the true zeros of the transducers were adjusted to the l e v e l of the heart. 24 S P E C I A L PREPARATIONS Blood Flow Measurements The abdomen was opened by a midline i n c i s i o n , and the contents were kept moist and warm i n a p l a s t i c bag and pushed aside to expose the external i l i a c , the superior mesenteric and the l e f t renal a r t e r i e s . Perivascular tissues were cleaned under a d i s s e c t i n g microscope (A.O. Instrument, Model 570) so that nerve f i b r e s accompaning the blood vessels were not damaged. In some experiments i n which blood flows to the i n t a c t and the skinned hind limbs were compared, both the l e f t and the r i g h t external i l i a c a r t e r i e s were prepared. One of the hindlimbs i n these animals was then skinned from the hip to the footpads, which i n turn were t i e d o f f i n d i v i d u a l l y so that blood flow to the footpads was minimal. The skinned l e g was smeared with a thin l a y e r of vaseline, covered with moisted gauze and kept i n a p l a s t i c bag. Blood flows were measured with polystyrene flow cuffs (2-4 mm, i.d.) placed around the prepared a r t e r i e s . A i r bubbles between the vessel and the probe were eliminated 25 by f i l l i n g the intervening space with vaseline. The probes . were connected to Doppler u l t r a s o n i c flowmeters (Parks E l e c t r o n i c Laboratory, Model 803), which conveyed the signals to a transducer/converter (S.E. Laboratories, Type SE423) with an e l e c t r o n i c device capable of converting the p u l s a t i l e flows to t h e i r integrated means, and recorded by a UV recorder (S.E. Laboratories, Type S E 3 O O 6 ) . At the end of each experiment, the u l t r a s o n i c probes and flowmeters were calibrated with a p e r i s t a l t i c pump (Harvard Apparatus, Model 1210) using blood taken from the same animal. The c a l i b r a t i o n curves so constructed were l i n e a r between 0-60 ml/min i n most experiments. A d e t a i l e d discussion of the p r i n c i p l e and i t s l i m i t a t i o n s of the Doppler technique f o r measuring blood flow i s found i n Appendix A. Vascular Resistance Calculation Vascular resistance was calculated according to the formula T T -, - O - J . Mean A r t e r i a l Pressure (rnmHg) Vascular Resistance = r: r — : — x — •" " m ,—, , . — r Mean A r t e r i a l Blood Plow (ml/min). The FAP or the IMP was used because the a r t e r i e s i n which 26 blood flows were measured and the femoral or the i n f e r i o r mesenteric artery are a l l located downstream of the a o r t i c cannula. As a f i r s t approximation, the venous pressures i n the external i l i a c , the superior mesenteric and the renal beds were taken to be constant during the period of blood flow measurement, and were n e g l i g i b l e i n magnitude as compared to the a r t e r i a l pressure. Pharmacological Blockade In some experiments, the TAR was e l i c i t e d before and a f t e r sequential administrations of phenoxybenzamine hydrochloride (PhB), propranolol hydrochloride (Prop) and atropine sulphate. A l l these drugs were injected intravenously. They were, with the exception of PhB, prepared f o r admini-s t r a t i o n by d i s s o l v i n g d i r e c t l y i n i s o t o n i c s a l i n e . PhB, which produced a turbid suspension i n s a l i n e , was prepared by f i r s t d i s s o l v i n g i n 1-2 ml of propylene g l y c o l , a c i d i f y i n g with a few drops of 0.1M HC1, and then d i l u t i n g to about 10 ml with i s o t o n i c s a l i n e . The re s u l t e d c l e a r solution was used f o r infusion over a period of 30 min 27 at a dosage of 5 mg/kg. The alpha-adrenergic blockade was tested by in j e c t i o n s of single dose (0.15 >ig/kg) of norepinephrine b i t a r t r a t e ( N E ) . I f the pressor e f f e c t of the sympathomimetic was dimished by at l e a s t 70%, the blockade was considered to be e f f e c t i v e . This us u a l l y occured 20-40 min a f t e r PhB administration. The TAR was then e l i c i t e d 2-4 times. Following t h i s , Prop (1 mg/kg) was infused also over 30 min to achieve beta-adrenergic blockade; which was not tested by any pharmacological agents. The TAR was again evoked 2-4 times before administration of atropine (0.5 mg/kg). The cho l i n e r g i c blockade was tested by in j e c t i o n s of single dose (0.15 mg/kg) of acetylcholine bromide. I f the depressor e f f e c t of the para-sympathomimetic was dimished by more than 70%, the blockade was considered to be e f f e c t i v e . This usually took place 5-10 min a f t e r atropine admini-s t r a t i o n . Carotid Sinus Nerve (CSN) Stimulation In cats i n which the baroreceptor or the chemo-receptor r e f l e x was i n i t i a t e d by e l e c t r i c a l stimulation of the CSN, the nerves on both sides were cut close to 28 the sinuses under the dis s e c t i n g microscope. The central end of one (usually the l e f t ) CSN was placed on a bipo l a r electrode within a pool of p a r a f f i n o i l . Pulse s t i m u l i were delivered from a stimulator (Grass Instruments, Model S8). The parameters set f o r baroreceptor f i b r e stimulation (1.5-3 V, 3 msec, 5-15 Hz) and chemoreceptor f i b r e stimulation (1.5-4 V, 0.07 msec, 8-40 Hz) were based on a study by N e i l , Redwood & Schweitzer (1949)* i n which they showed that long duration pulses activate mainly the baroreceptor f i b r e s while short duration pulses activate the chemoreceptor f i b r e s i n the CSN of cats under chloralose anesthesia. Carotid Sinus Perfusion Only t h e 2 l e f t c a r o t i d sinus was prepared f o r perfusion. A l l blood vessels connected to th i s sinus except the common caro t i d , the external and internaljcarotid, and the o c c i p i t a l a r t e r i e s , were t i e d o f f with very f i n e l i g a t u r e s under the dis s e c t i n g microscope. The l i g a t e d vessels were cut, i f necessary. The CSN was however l e f t i n t a c t . A fter the cat was heparinized, the common carotid and the external carotid a r t e r i e s were cannulated with 29 J-shaped metal cannulas made from #15» 16 or 17 hypodermic needles. These two cannulas respectively formed the perfusion i n l e t and outlet of the sinus. A var i a b l e speed p e r i s t a l t i c pump (Harvard Apparatus, Model 1210) was used to draw blood from a source (to be described below) and . pump to the i n l e t cannula v i a a silicon-rubber tubing ' engulfed i n a c i r c u l a t i n g hot water jacket. The pressure i n the sinus (CSP) was measured by a transducer (Statham, P23Db) connected to a side-arm of this cannula. The perfusate came out of the sinus v i a the cannula i n the external c a r o t i d artery. The outflow tubing was f i t t e d with a screw-clamp which allowed variable resistance to flow before i t returned to i t s appropriate source ( F i g . 2). The i n t e r n a l c a r o t i d artery was also l i g a t e d a f t e r the perfusion pump was turned on, so that the l e f t c a r o t i d sinus was then almost completely i s o l a t e d from the rest of the c i r c u l a t i o n . 30 Per is ta l t ic Pump |0 Blood Source Baroreceptor Expt.: Own Blood Chemoreceptor Expt.: Donor 's Blood o Statham Transducer H t Screw Clamp Water ( 4 5 ° C ) Figure 2. Schematic representation of the perfusion system used to activate the carotid haroreceptors and chemoreceptors, 31 Carotid Baroreceptor Activation A t o t a l of 10 animals were perfused i n the l e f t c a r o t i d sinus f o r baroreceptor a c t i v a t i o n . The r i g h t common ca r o t i d artery i n 5 of these animals was occluded, and the r i g h t CSN i n the other 5 animals was sectioned. In a l l of these animals, blood was drawn by the p e r i s t a l t i c pump from the l e f t common ca r o t i d artery to perfuse the sinus on the same side. The perfusate re-entered the animal v i a the l e f t jugular v e i n . To activate the baroreceptors, the screw-clamp regulating the e f f e r e n t flow or the speed of the pump was manipulated, so that CSP increased from a normal range of 95-110 mmHg to an intermediate range of 140-155 mmHg, or to a higher range of 195-215 mmHg. These changes provided adequate s t i m u l i to evoke the carotid baroreceptor r e f l e x as showed by a f a l l of more than 20 rjimHg i n PAP. Carotid Chemoreceptor Activation Six cats were perfused i n the l e f t c a r o t i d sinus f o r chemoreceptor a c t i v a t i o n , the r i g h t CSN was cut i n a l l these animals. Blood was drawn from the common car o t i d artery 32 of the donor to perfuse the sinus of the recipient, and the perfusate was returned to t^he donor v ia the jugular vein. CSP was maintained constant between 100-110 mmHg throughout the experiment to minimize baroreceptor influences. To stimulate the chemoreceptors of the recipient, the 100% 0^ cylinder supplying the respirator pump to the donor was replaced by another cylinder of a low 0 2 gas mixture (8% 0 2 , 5% C 0 2 , 87% N 2 ) . The ar ter ia l blood gases and pH of the donor perfusing the carotid sinus of the recipient changed from P 0 , , 297+14 mmHg, P C 0 2 25+2 mmHg, pH 7.37+0.02 to P 0 2 41+5 mmHg, PC02 49+1 mmHg, pH 7 . 1 5 + 0 . 0 5 . These changes provided adequate stimuli to evoke the carotid chemoreceptor reflex and cause a pressor effect of more than 20 mmHg in PAP in a l l the recipient animals. norepinephrine Infusion and Change of Blood Volume In 5 animals, a higher basal blood pressure was obtained by HE infusion. 0 . 2 % NE from a v i a l was diluted to 0 . 3 jig/ml with isotonic saline and stored in a 30 ml syringe. A multiple-speed steady infusion pump (Harvard Apparatus, Model 942) was used to infuse 0 . 0 7 - 0 . 1 5 ^ig/kg /min of NE into the cat via the femoral vein, so that a 33 pressure comparable to that obtained by chemoreceptor stimulation was obtained. In another 5 animals, the basal blood pressure was changed by loading with exogenous blood, or by bleeding to y i e l d a higher and a lower systemic blood pressure respectively; which were comparable to those obtained by chemoreceptor and baroreceptor stimulations. 34 EXPERIMENTAL PROTOCOL In a l l the experiments, the duration of one a o r t i c s t r e t c h (AS) was about 1 min, and the i n t e r v a l between any two AS was at l e a s t 5 min. (I) i n the blood flow measurement experiments, any two of the a r t e r i a l flows were measured simultaneously before, during and a f t e r AS. These procedures were repeated i n some of the experiments when the animals were under alpha-adrenergic, beta-adrenergic and cholinergic blockade. (II) i n the carotid baroreceptor a c t i v a t i o n experiments, a few AS were performed to show that the response of the TAR was constant. The l a s t of these AS was considered to be the control stretch; then the baro-receptor r e f l e x was evoked, and a f t e r i t had reached a steady state (usually within 1 min), the thoracic aorta was stretched again to the same degree as the control s t r e t c h . In some t r a i l s , t h i s order of a control AS followed by an AS during baroreceptor a c t i v a t i o n was reversed to ensure randomness i n the operation, so that any change i n the TAR response during baroreceptor 35 a c t i v a t i o n compared to the control r e f l e x response would be s o l e l y due to baroreceptor a c t i v a t i o n and not to the order i n which the two AS were performed. ( i l l ) i n the carotid chemoreceptor a c t i v a t i o n experiments, the procedures were s i m i l a r to those described f o r the c a r o t i d baroreceptor a c t i v a t i o n experiments. (IV) i n the p r e v a i l i n g a r t e r i a l pressure experiments, AS were performed at low, control and high basal blood pressures. The order of operation was again randomized. 36 DATA ANALYSIS A l l calculations were done on an e l e c t r o n i c desk computer ( O l i v e t t i Programma 101) . The control values and the maximum changes induced i n each v a r i a b l e by AS were averaged f o r each animal. The r e s u l t i n g values were pooled from the same group of experiments to give the mean and standard deviation f o r that p a r t i c u l a r group. The paired student t - t e s t was used to determine the l e v e l of significance regarding any change within one experimental group, while the unpaired t-te s t was applied when the re s u l t s between two experimental groups were compared (Snedecor & Cochran, 1973) • P value greater than 0.05 was considered not s i g n i f i c a n t . In some of the graphs that are presented, l i n e a r regression was used to f i n d the b e s t - f i t t i n g l i n e , and the c o r r e l a t i o n c o e f f i c i e n t between the two plotted variables was I determined. 37 PART I I I . RESULTS REFLEX CHANGES IN ARTERIAL PRESSURE  Control In a control group of 22 vagotomized cats with both common carotid a r t e r i e s occluded, stretch of the walls of the thoracic aorta (AS) induced marked increases i n a r t e r i a l pressure measured i n the common carotid artery (CAP) and the femoral artery (FAP). A l l these changes were* s t a t i s t i c a l l y s i g n i f i c a n t (Table I ) . F i g . 3 shows one of these experiments: p u l s a t i l e CAP and mean FAP rose soon a f t e r i n f l a t i o n of the a o r t i c b a l l o n and remained elevated throughout the period of AS. The hemodynamic ef f e c t s were not due to obstruction of the thoracic a o r t i c blood flow because a p a r a l l e l pressure increase was observed both above and below the a o r t i c cannula. In most experiments, as i n the one presented i n F i g . 3» a second l a t e increase i n blood pressure was recorded about 50 s©c a f t e r the i n i t i a l r i s e . When the ball o n was deflated, the a r t e r i a l pressure f e l l towards 38 the control l e v e l , and sometimes f e l l "below i t , but rose again and eventually reached the control l e v e l a f t e r d i f f e r e n t periods of time (30-200 sec). These e f f e c t s of AS on a r t e r i a l pressure observed i n the present study were consistent with those described by Lioy et a l (1974). I min. C A P r - 2 0 0 - 150 A.S . FAP 1 5 0 -100 -5 0 -h 100 5 0 Figure 3. Reflex changes i n a r t e r i a l pressure induced by stretching the walls of the thoracic aorta. Abbreviations used i n th i s and subsequent figures: A.S.=aortic stretc h , CAP=common carotid artery pressure, FAP=feraoral artery pressure. Table I Effects of Phenoxybenzamine, Propranolol, and Atropine on the Reflex Pressure Changes induced by Stretching the Walls of the Thoracic Aorta. Carotid artery systolic pressure Group Bo. cats Ho. t r i a l s Control (mmHg) AAS Control 22 89 142+4 +35+2* Phenoxy-benzamine 7 30 128+3 + 8+2 P £ 0.0005 Propran-olo l 7 24 120+3 + 3+2 P ... N.S. Atropine 7 26 122+4 +5+3 P N.S. Both common carotid arteries occluded and the vagi cut. Phenoxybenzamine (5 mg/kg), propranolol (1 mg/kg) and atropine (0.5 mg/kg) were administered i . v . in sequential order (see METHODS). A l l values are mean+S.E. 40 Carotid artery Femoral artery d i a s t o l i c pressure mean pressure (mmHg) (mmHg) Control A AS Control AAS 105+3 +26+1* 115+3 +29+2* 87+4 -11+3 99+3 - 1+2 < 0.0005 4 0.0005 86+3 - 5+2 95+2 + 1+2 N.S. N.S. 85+3 + 2+3 95+3 + 3+2 N.S. N.S. AAS=change with a o r t i c s t r e t c h , 3t=P<0,01 or hetter (paired student t - t e s t ) . P values between successive experimental groups were determined by the unpaired student t - t e s t . 41 Pharmacological Blockade Sequential administrations of PhB and Prop caused immediate and transient decreases i n a r t e r i a l pressure i n seven animals, successive administration of atropine did not cause any further e f f e c t . The blood pressure of these animals s t a b i l i z e d at a l e v e l s i g n i f i c a n t l y (P40.01, paired student t-test) lower than that of the control group (Table I). Ef f e c t s on the TAR Response When AS was performed a f t e r PhB treatment, the TAR response was d r a s t i c a l l y reduced ( F i g . 4B,5B), an observation which had also been described by Lioy et a l ( 1 9 7 4 ) . The remaining r e f l e x response was abolished a f t e r administration of Prop ( F i g . 4C,5C). The blockade was s t i l l e f f e c t i v e a f t e r atropine treatment ( F i g . 4 D ) , although i n some cases, the response started to reappear as i f the adrenergic blockade was waning ( F i g . 5*0 . Cholinergic blockade did not af f e c t the TAR much, except that the small decreases i n d i a s t o l i c and mean a r t e r i a l pressure obtained during AS under adrenergic blockade 42 were no longer present i n the atropine treated animals (Table l ) . A l l these pressure changes induced by AS under adrenergic and/or cholinergic blockade were not s t a t i s t i c a l l y s i g n i f i c a n t (Table I ) . Figure 4. Reflex changes i n blood flows i n the external i l i a c a r t e r i e s of the i n t a c t and skinned legs induced by a o r t i c s t r e t c h . A:Control. B:After phenoxybenzamine (5 mg/kg). CrAfter propranolol (1 mg/kg). D:After atropine (0.5 mg/kg). The drugs were administered i n sequential order as l i s t e d . The cat was vagotomized and car o t i d occluded b i l a t e r a l l y . . A.S.=period of a o r t i c stretch. CAP=carotid artery pressure (mmHg). IF=external i l i a c artery flow (ml/min). IMP=inferior mesenteric artery pressure (mmHg). 4 4 45 CAP RF ® I min. ® I AS. I I A.S. I 1 A.5. I I A.S. I Figure 5» E f f e c t s of a o r t i c stretch on blood flows i n the renal and the superior mesenteric a r t e r i e s . A:Control. B:After phenoxybenzamine. C:After propranolol. D:After atropine. RF=renal artery flow (ml/min), SMF=superior mesenteric artery flow (ml/min). Other abbreviations, preparations of animal and dosages of drugs are the same as i n Figure 4« 46 REFLEX CHANGES IN ARTERIAL FLOWS  Control In 22 animals, renal flow (RF) was higher than external i l i a c flow (IF) and superior mesenteric flow (SMF). The calculated vascular resistance was accordingly-lowest i n the renal a r t e r y , s l i g h t l y higher i n the i l i a c a r t e r y and highest i n the superior mesenteric a r t e r y (Tables 1L-JZ). When AS was performed, a r t e r i a l pressure rose as already described, IF and SMF f e l l ( F i g . 4A,5A), while RF might r i s e or f a l l , or d i d not change. The peak of the r i s e i n a r t e r i a l pressure frequently corresponded to the nadir of the f a l l i n a r t e r i a l flows. Both para-meters returned to control l e v e l s when the s t r e t c h was released. The pooled r e s u l t s show that during AS, there was a s l i g h t but not s i g n i f i c a n t reduction i n I F (Table JL) and SMF (Table 3U»), but there was no change i n RF (Table I t ) . A s i g n i f i c a n t increment i n vascular resistance was evident i n a l l three beds measured i n d i c a t i n g v a s o c o n s t r i c t i o n ( F i g . 6); the extent of the v a s o c o n s t r i c t i o n was l a r g e r 47 i n the superior mesenteric "bed, A d i r e c t c o r r e l a t i o n e x i s t s between the magnitude of the TAB, as expressed by the increase i n a r t e r i a l pressure, and the extent of the v a s o c o n s t r i c t i o n i n the vascular beds of the three v e s s e l s ( F i g . 7). The points i n t h i s figure were p l o t t e d by c a l c u l a t i n g the change i n a r t e r i a l pressure and the change i n vascular resistance as a percent of t h e i r r e s pective pre—stimulation values. IF i n the i n t a c t and the skinned legs were compared i n f i v e animals. There was no s i g n i f i c a n t d i f f e r e n c e between the responses from the two legs (Table 3 £ ) when TAR was evoked i n control s i t u a t i o n and under pharmacological blockade ( F i g . 4). Pharmacological Blockade Administrations of PhB and Prop increased IF, SMF and RF with a drop i n resistance i n these vascular beds. The change i n SMF was l a r g e r and was s t a t i s t i c a l l y s i g n i -f i c a n t ( P < 0 . 0 2 5 , paired student t - t e s t ) . Atropine d i d not. induce f u r t h e r changes i n SMF and RF, but caused a s l i g h t and n o n - s i g n i f i c a n t decrease i n IF, which returned to control values (Tables "JSL\ Table II E f f e c t s of Phenoxybenzamine, Propranolol, and Atropine on the r e f l e x changes i n Blood Flow and Vascular Resistance i n the External I l i a c Artery induced by Ao r t i c Stretch Group No. cats No. t r i a l s? Femoral artery mean pressure (mmHg) Control AAS Control 13 77 114+7 +26+3* 1 PhB 5 21 96+3 + 9+2 P < 0.0025 Prop 5 18 92+3 + 4+3 P N.S. Atropine 5 19 95+2 + 7 i 2 P N.S. Preparations and abbreviations are the same as i n Table I. 49 External i l i a c artery flow (ml/min) Vascular resistance (mmHg/ml/min) Control AAS Control A AS 21.0+2.7 -0 .8+0.3 5.6+1.2 +1.3+0.3* 22.4+2.4 +0.7+0.2 < 0.005 4.3+0.5 +0.3+0.2 < 0.05 22.4+3.8 +2.4+0.4 <T0.005 4.1+1.0 - 0 . 9 + 0 . 3 * £ 0.01 20.4+4.2 +1.0+0.3 < 0.0125 4 .6+1.3 +0.1+0.1 < 0.01 Table III Eff e c t s of Phenoxybenzamine, Propranolol, and Atropine on the r e f l e x changes i n Blood Plow and Vascular Resistance i n the Superior Mesenteric Artery induced by A o r t i c Stretch Group No. cats No. t r i a l s Femoral artery mean pressure (mmHg) Control A AS Control 8 50 115+5 +24+3* PhB P 4 13 100+2 + 6+1 4 0.0025 Prop / P 4 12 98+3 + 3+1 < 0.05 Atropine P 4 16 101+3 + 9+2 < 0.025 Preparations and abbreviations are the same as i n Table I. 51 Superior mesenteric artery flow Vascular resistance (ml/min) (mmHg/ml/min) Control ZiAS Control A AS 16.7+2.4 -1.1+0.1 6.8+1.3 +2.0+0.3* 27.0+3.1 +0.7+0.2 3.7+0.6 +0.3+0.1 0.0005 4 0.0025 27.6+1.9 +1.5+0.3 3.6 +0.6 +0.2+0.1 < 0.05 N.S. 27.7+2.5 +1.0+0.2 3.6+0.7 +0.2+0.1 N.S. N.S. Table IV Ef f e c t s of Phenoxybenzamine, Propranolol, and Atropine on the r e f l e x changes i n Blood Plow and Vascular Resistance Stretch i n the Renal Artery induced by Ao r t i c No. No. Femoral artery mean pressure (mmHg) Group cats t r i a l s Control A A S Control 8 49 118+6 +28+4* PhB 5 18 102+1 + 5+2 P 0.0025 Prop 5 15 98+3 + 3+1 P N . S . Atropine 5 14 103+2 +11+2 P < 0.005 Preparations and abbreviations are the same as i n Table I. 53 Renal artery flow Vascular resistance (ml/rain) (nunHg/ml/min) Control A AS Control A AS 23.0+1.1 0.0+0.1 5.5+0.6 +1.1+0.2* 25.2+2.4 -1.0+0.1 4.0+0.5 +0.3+0.0 £ 0.0005 4. 0.005 26.2+1.8 -2.8+0.2 3.7+0.9 +0.9+0.3* < 0.0005 < 0.05 26.1+2.3 -1.0+0.2 4.0+1.0 +0.7+0.4 < 0.0005 N . S . Figure 6 . The time-related changes i n vascular resistance during the period of a o r t i c stretch under normal conditions and under pharmacological blockade. PhB = phenoxybenzamine. Prop = propranolol. 55 Figure 7. Correlations "between changes i n vascular resistance and a r t e r i a l pressure induced by a o r t i c s t r e t c h . 57 58 Table V E f f e c t s of A o r t i c Stretch on Blood Plows and Vascular Resistance i n the Intact and Skinned Legs I n f e r i o r mesenteric artery mean pressure vr >T (mmHg) No. No. v J  Group cats t r i a l s Control AvAS Intact Leg 5 32 109+4 +25+3* Skinned Leg P Preparations and abbreviations are the same as i n Table I. 59 External i l i a c a rtery flow Vascular resistance (ml/min) . ^s/ml/mxn) Control A A S Control A AS 20.2+2.0 -0.6+0.1 5.2+1.3 +1.2+0.1* 19.1+4.3 -0.5+0.2 5.4+2.0 +1.1+0.2* N .S . ' N .S . N .S . N .S . 6 0 E f f e c t s of Pharmacological Blockade  on Reflex Changes of Blood Plow In a l l but two of the animals, AS under alpha-adrenergic blockade induced small changes i n IF, SMF and Ri' ( F i g . 4F,5B), which were not s t a t i s t i c a l l y s i g n i f i c a n t (Tables X-3SO. In two cats, however, AS a f t e r PhB treatment induced a delayed but marked increase i n IF, i n d i c a t i n g v a s o d i l a t a t i o n i n t h i s vascular bed ( F i g . 8 B J , while some vaso c o n s t r i c t i o n was s t i l l present i n the renal a r t e r y . The average maximum decrease i n i l i a c r e s i s t a n c e was 1.9+0.2 mmFg/ml/min i n these animals, and was s t a t i s t i c a l l y s i g n i f i c a n t (P<0.005» paired student t — t e s t ) . This i l i a c v a s o d i l a t a t i o n was almost completely abolished by admini-s t r a t i o n of atropine ( F i g . 8 C ) . Some a r t e r i a l pressure increase was s t i l l present a f t e r PhB and atropine probably because of beta-adrenergic stimulation of the hear t . Since such d i s t i n c t i v e v a s o d i l a t a t i o n was not observed i n the other animals, the r e s u l t s from these two cats were not grouped with those obtained from the other- f i v e animals. 61 Figure 8. Reflex cholinergic vasodilatation i n the external i l i a c vascular hed induced by a o r t i c s t r e t c h . A:Control. B:After phenoxybenzamine. C:After atropine. 62 In the other animals, AS performed a f t e r Phb and Prop treatments induced a s l i g h t increase i n I F ( F i g . 4G) with a s i g n i f i c a n t decrease i n vascular re s i s t a n c e ( F i g . 6A), which disappeared a f t e r c h o l i n e r g i c blockade ( F i g . 4D.6A). This i 3 the same phenomenon that was more dramatically evident i n the experiment presented i n F i g . 8. AS under complete adrenergic blockade induced a s l i g h t increase i n SMF and a considerable decrease i n RF ( F i g . 5C) . The change i n renal resistance was l a r g e r and was s i g n i f i c a n t ( F i g . 6B,C). Cholinergic blockade d i d not change the pattern of the r e f l e x response i n the superior mesenteric and renal vascular beds (Tables J&.,JZ)m 63 THE BARORSCSPTOR REFLEX & THE TAR Ef f e c t s of Carotid Occlusion and Sinus Denervation F i f t e e n vagotomized cats were divided i n t o three groups of 5 animals each. In the group I animals, the ca r o t i d sinus nerves (CSN) on both sides were cut and blood flows i n the common ca r o t i d a r t e r i e s were normal. In the group I I animals, the l e f t c a r o t i d sinus was perfused at a pressure (CSP) of 103+3 mmHg-, and the co n t r a l a t e r a l CSN was sectioned. In the group I I I animals, the l e f t sinus was also perfused at a con t r o l CSP of 101+3 mmHg, and the c o n t r a l a t e r a l common c a r o t i d a r t e r y was occluded. A f t e r the conditions of the animals had become steady>*,the control blood pressure was lower i n group I I I than i n group I and I I (Table ' s ! ) When AS was performed, the c h a r a c t e r i s t i c p r e s s o r response of the TAR already described occured i n a l l three groups of animals. The magnitude of the response was somewhat smaller i n group I I I than i n the other two groups (Table TB). Table VI Effects of Carotid Baroreceptor Stimulation on the reflex pressor response induced by Aortic Stretch Group No. cats No. t r i a l s Baro-receptor stim. Carotid artery systolic pressure (mmHg; Control A AS I 5 37 CSN stim. Control 153+8 +42+2* Stim. 124+7 +21+2* P < 0.0005 II 5 33 CSP(mmHg) 103+3 152+2 +37+2* 212+9 127+5 +18+1* P <C 0.0005 I I I 5 32 CSP(mmHg). 101+3 136+9 +28+3* 203+8 114+12 +18+2* P < 0.0125 Vagi cut in a l l cats. Group I: both CSN cut. Group I I : contralateral CSN cut. Group I I I : contra-la tera l carotid occluded. CSN = carotid sinus nerve(s). 65 c a r o t i d artery d i a s t o l i c pressure (mmHg) Control A AS Femoral artery mean pressure (mmHg) Control A AS 112+7 82+4 +28+3 +14+2* £ 0.0025 127+8 95+5 +32+3 +16+2* < 0.0025 114+2 84+5 +27+2 +18+1* < 0.0025 120+2 92+5 +29+2 +17+1* < 0.0005 103+7 81+ +22+2 +14+2* <0.0125 112+7 88+7 +24+2 +18+1* < 0.025 CSP = ca r o t i d sinus perfusion pressure. Other abbreviations and s t a t i s t i c a l treatments are the same as i n Table I. 66 Carotid Baroreceptor St jural at ion Stimulation of tho centx'al end of one CSN (1•5—3 V, 3 msec, 5-15 Hz) i n group I animals induced s i g n i f i c a n t (P<0.005 or better, paired student t — t e s t ) depressor responses c h a r a c t e r i s t i c of the baroreceptor r e f l e x . Increasing the CSP i n group I I and I I I animals also caused an immediate f a l l i n a r t e r i a l pressure. Although the increment i n CSP was the same i n the two groups, the f a l l i n blood pressure was l e s s marked i n group I I I than i n group II (Table!!,). Throughout the period of baroreceptor stimulation, the new and lower blood pressure u s u a l l y remained steady. I f i t 'escaped' i . e . the blood pressure started to re t u r n towards the pre—stimulation l e v e l even though the s t i m u l a t i o n was s t i l l continuing, the s h i f t i n baseline was taken i n t o account i n determining the magnitude of the TAB response E f f e c t s on the TAR Response When AS was performed during the a c t i v a t i o n of the baroreceptor r e f l e x e i t h e r by s t i m u l a t i n g the CSN (Pig . 9) or by inc r e a s i n g the CSP ( F i g . 10), the r e f l e x response was s i g n i f i c a n t l y suppressed (Table2 E ) . 67 Figure 9. E f f e c t s of stimulating the central end of the carotid sinus nerve on the pressor response induced by a o r t i c s t r e t c h . The cat was vagotomized and both sinus nerves cut. SN=carotid sinus nerve stimulation. 68 © ® C A P I min. Figure 1 0 . E f f e c t s of increasing the carotid sinus perfusion pressure on the pressor response induced "by a o r t i c s t r e t c h . The co n t r a l a t e r a l sinus nerve and the vagus nerves were cut. CSP=carotid sinus pressure (mmHg). 5>, 69 The degree of suppression was rel a t e d to the stimulation frequency applied to the CSN ( F i g . 9 ) , and to the l e v e l of pressure i n the sinus ( F i g . 10). This i s also i l l u s t r a t e d i n F i g 11 and 12, where the extent of the TAR response during baroreceptor stimulation, expressed as a percent of the response obtained i n the control s i t u a t i o n , i s plotted against the frequency of stimulation ( F i g . 11) and against the CSP ( F i g . 12). The suppression of the TAR response by baroreceptor a c t i v a t i o n appeared to be less pronounced i n the group I I I (c o n t r a l a t e r a l c a r o t i d occluded) animals than i n the group II ( c o n t r a l a t e r a l sinus denervated) animals ( F i g . 12A,B). 70 Figure 11. Stimulus frequency-Response curve ( © ) f o r the baroreceptor f i b r e s i n the carotid sinus nerve (CSN) and the graded suppression on the pressor response ( ® ) induced by a o r t i c s t r e t c h . Vagi and both CSN were cut i n a l l 5 animals. 71 @ Contralateral Common Carotid Occluded 95-II0 I40-I55 I95-2I5 Carotid Sinus Perfusion Pressure ( mmHg) Figure 12. Stimulus-Response relationships (O) for the carotid sinus in two groups of animals and their graded suppression on the pressor response (< )^ induced by aortic stretch. 72 THE CHEMORECEPTOR REFLEX & THE TAR Carotid Chemoreceptor Stimulation In s i x vagotomized cats, stimulation of one of the sectioned CSN with short duration pulses (1.5-4 v , 0.07 msec, 8-40 Hz) evoked pressor responses c h a r a c t e r i s t i c of the chemoreceptor r e f l e x . In another six animals with the contralateral CSN cut, perfusion of the l e f t sinus with hypoxic, hypercapnic and a c i d o t i c "blood from a donor at a constant CSP (104+3 mmHg) induced s i m i l a r s i g n i f i c a n t (P<0.005 or better, paired student t-test) increases i n a r t e r i a l pressure. Increased 1. respiratory movements of the diaphragm were observed i n some experiments i n which the effects of gallamine t r i e t h i o d i d e were apparently wearing o f f . In most of the chemoreceptor stimulation experiments, the pressor response remained at a stable l e v e l throughout the period of stimulation. In those cases where a r t e r i a l pressure •escaped', the baseline s h i f t was taken into account i n determining the magnitude of the TAR response. 73 E f f e c t s on the TAR Response When AS was performed during- chemoreceptor a c t i v a t i o n e i t h e r by CSN stimulation (.Fig. 13) or by sinus perfusion ( F i g . 14)» the TAR response was s i g n i f i c a n t l y reduced i n comparison to the control . s i t u a t i o n s (Table 22). The degree of suppression of the TAR also appeared to be r e l a t e d to the stimulation frequency of the CSN ( F i g . 13B,C); but to p l o t a stimulus-response curve s i m i l a r to F i g . 11 was not f e a s i b l e i n t h i s case because there was a great v a r i a t i o n of stimulation frequency necessary to induce comparable chemoreceptor responses i n d i f f e r e n t animals. Instead, the extent of the TAR i s p l o t t e d against the a c t i v i t y of the chemoreceptors ( F i g . 1 5 A , 3 ) , no d i r e c t c o r r e l a t i o n between them i s evident. In t h i s f i g u r e , the extent of the TAR i s expressed as a percent of the response obtained i n control s i t u a t i o n , and the a c t i v i t y of the chemoreceptors i s assessed as the percent increase i n FAP from control during chemoreceptor stimulation. 74 ® C A P 2 0 0 I min FAP , c m I S N - 0 . 0 7 ms 10 Hz I C A P 2 0 0 • 150-100-J _ F A P n i s i S N - 0.07 ms 20 Hz | Figure 13. E f f e c t s of stimulating the central end of the carotid sinus nerve with short duration ( 0 . 0 7 msec) pulses on the pressor response induced "by a o r t i c s t r e t c h . 75 Figure 14. Effe c t s of carotid chemoreceptor stimulation by cross-perfusion of the sinus on the pressor response induced by a o r t i c s t r e t c h . Blood gases and pH of the donor are given i n the f i g u r e , and of the r e c i p i e n t : P 0 2 213 mmHg, P C 0 2 25 mmHg and pH 7.39. DFAP=femoral artery pressure (mmHg) of the donor. CAP, FAP and CSP are of the r e c i p i e n t . 76 Table VII Effects of Carotid Chemoreceptor Stimulation on the reflex pressor response induced by Aortic Stretch Chemo-Carotid artery systolic pressure (mmHg) Group cats t r i a l s stim. Control A AS I 6 35 CSN stim. Control 143+7 +34+3* Stim. 163+10 +25+3* P < 0.05 II 6 38 CS perfusion P0 2 PC02 pH 297 " 2 5 7.37 144+7 +39+4* 41 49 7.15 ,171+10 +26+4* P < 0.025 Both CSN cut in group I, and contralateral CSN cut in group II . 7 7 a Femoral artery mean pressure (mmHg) Control A AS 1 1 6 + 8 + 3 0 + 3 1 3 7 + 1 0 + 2 1 + 3 * < 0 . 0 5 Carotid artery d i a s t o l i c pressure (mmHg) Control A AS 1 0 7 + 9 +28+3 1 2 7 + 1 0 + 1 8 +3* < 0 . 0 2 5 1 0 4 + 6 128+7 + 3 1 + 4 * +18+4* < 0 . 0 2 5 1 1 7 + 4 1 4 0 + 5 + 3 2 + 4 * + 2 0 + 4 < 0 . 0 5 77b 100 75 -c 50 ' + 25 o < 0 o @ Chemoreceptor Stimulation ' ( Cross-Perfusion ) N=6 x - A * -i r Chemoreceptor CSN Stim. i r g © NE Infusion g, IOO-i 75-50 -| + 25H o-1 r N = 5 1 1 1 1 1 +10 20 30 40 50 © Blood Volume Expansion N = 5 — I 1 1 1 + 10 20 30 40 Change in FAP (%) Figure 15. Suppression of the TAR pressor response by chemoreceptor stimulations (A and B) , and by higher control blood pressures (C and D). 78 EFFECTS OP PREVAILING ARTERIAL PRESSURE Since the e f f e c t s of pharmacological blockade, and of the baroreceptor and chemoreceptor r e f l e x e s on the TAR were assessed when the blood pressure had been changed as a r e s u l t of drug treatment and baroreceptor or chemoreceptor a c t i v a t i o n , i t i s necessary to assess* the e f f e c t s of these s h i f t s i n baseline pressure. Therefore the TAR was induced at a lower c o n t r o l pressure comparable to those obtained a f t e r alpha-adrenergic blockade and during baroreceptor s t i m u l a t i o n , and at a higher control pressure comparable to the l e v e l obtained during chemoreceptor stimulation. Lower Blood Pressure The blood pressure lower than c o n t r o l was acquired by bleeding. There was no s i g n i f i c a n t changes i n the r e f l e x response at a systemic a r t e r i a l pressure of 9 2 + 3 mmHg as compared to a control pressure of 1 1 9 + 6 mmHg (Table However, the AS induced response was abolished i f the systemic a r t e r i a l pressure was lowered below 7 5 mmHg. 79 H i g h e r Blood P r e s s u r e The blood pressure higher than c o n t r o l was obtained e i t h e r by NE i n f u s i o n (.Pig. 16) o r by blood volume expansion ( F i g . 17)« The r e f l e x response was s t i l l present but s i g n i f i c a n t l y smaller than when AS was performed at the control pressure. (Table .Vlil.). However, the reduction on the TAS response was l a r g e r during chemoreceptor sti m u l a t i o n than during NE i n f u s i o n or blood volume expansion (Fig.. 15 and Tables 3ZI, 32EO. 80 ® C A P 2 0 0 - . I 5 0 H 100 5 0 min F A P 4-150 100 50 MMW C A P FAP NE Infusion (0.1 / j g / k g / m i n ) A.S. | A.S. Figure 16. Ef f e c t s of a higher basal blood pressure on the pressor response induced by a o r t i c stretch. Norepinephrine (NE) was infused intravenously. 81 Figure 17. E f f e c t s of a higher basal blood pressure on the pressor response induced by a o r t i c s t r e t c h . Table V I I I E f f e c t s of Low and High Basal Blood Pressure on the r e f l e x pressor response induced by A o r t i c Stretch No. No. Carotid artery s y s t o l i c pressure (mmHg) Group cats t r i a l s Control A AS I 5 21 A B V Control 1 4 3 + 5 + 3 5 + 3 * Low 1 2 0 + 5 + 3 3 + 4 * N.S. High 162+6 +2 4 + 3 * P < 0 . 0 2 5 I I 5 22 NE Control 1 4 6 + 9 + 2 7 + 2 * High 178+11 +2 3 + 3 * P N.S. Both c a r o t i d sinus nerves and vagi were cut i n a l l animals. .ABV = change i n blood volume by bleeding or loading. NE = norepinephrine ( 0 . 0 7 - 0 . 1 5 jxg/kg/min) i n f u s i o n . 83 Carotid artery d i a s t o l i c pressure Control A AS Femoral artery mean pressure (mmHg) Control A AS 106+5 +23+2" 119+6 +26+2 83+5 +22+3 N.S." 92+3 +25+3 N.S. 128+7 +16+2 < 0.025 134+6 +17+1~ < 0.0025 105+9 134+4 +22+2* +16+2* < 0.05 111+7 143+10 +23+2 +17+2* < 0.05 84 PART IV. DISCUSSION In t h e i r report on the hemodynamic changes induced by a mechanical stretch of the walls of the thoracic aorta, Lioy et a l (1974) have demonstrated r e f l e x increases i n a r t e r i a l pressure, heart rate and c o n t r a c t i l i t y (LV dP/dt max). In the present study only the a r t e r i a l pressure was measured as an index of the r e f l e x response, and no data were obtained concerning the inotropic and chronotropic e f f e c t s . Since the aim of some of the experiments was to investigate the extent and c h a r a c t e r i s t i c s of supraspinal control on the s p i n a l r e f l e x , the central nervous system was l e f t i n t a c t i n a l l animals. REFLEX CHANGES IN ARTERIAL PRESSURE AND FLOWS The Doppler Flowmeter Method The possible sources of error of the Doppler technique f o r measuring a r t e r i a l flows have been discussed i n Appendix A. Since i n the present experiments the period 85 of blood flow measurement i n each t r i a l (before, during and a f t e r an AS) was about two minutes, and i t i s very u n l i k e l y that the conditions of the probes and of the animal could have changed d r a s t i c a l l y within t h i s period of time, errors due to p o s i t i o n i n g of the probes and/or changes i n hematocrit would be minimal. I t i s our opinion that the Doppler flowmeters used i n the present study are adequate:to show r e a l t i v e changes i n flow induced by the TAR. The External I l i a c Flow (IF) I t i s conventional to report blood flow i n ml/min/g t i s s u e . This u n i t i s not used i n the present report because we have not measured the weights of the organs i n which blood flows have been assessed. The external i l i a c flow (IF) i n anesthetized cats measured by the Doppler flowmeters was about 23 ml/min. This i s comparable to measurements by electromagnetic flowmeters i n conscious animals (Mancia, B a c c e l l i , Zanchetti, 1972). Under control s i t u a t i o n s , AS induced marked increases i n a r t e r i a l pressure and a small but consistent 86 reduction i n IF, which was caused by a s i g n i f i c a n t potentiation of vascular resistance, as i l l u s t r a t e d i n F i g . bA. Moreover, there was a close c o r r e l a t i o n between the changes i n a r t e r i a l pressure and i l i a c resistance ( F i g . 7A,B). I t i s concluded that vasoconstriction of the i l i a c bed contributes to the pressor response of the TAR. The external i l i a c artery supplies the muscle, skin and other tissues i n the hindlimb ( F i e l d & Taylor, 1 9 5 6 ) . A l a r g e r portion of the blood flow i s probably directed to the muscle, because the skin of the l e g receives comparatively less vessels than the muscle. The c a p i l l a r y density of the skin has been estimated to 2 be 16-65 capillaries/mm as compared to 1000-2000 capillaries/mm i n s k e l e t a l muscle (Korner, 1 9 7 4 ) , but the basal tone of the skin vessels i s considerably lower than that of the muscle's (Celander & Folkow, 1 9 5 3 ) • We can therefore conclude that vasoconstriction d e f i n i t e l y occured i n the muscular bed and perhaps also i n the cutaneous bed, since both the muscle and the skin i n the cat's l e g are innervated by sympathetic vasoconstrictor f i b r e s (Green & Kepchar, 1 9 5 9 ) . However, t h e i r r e l a t i v e 87 c o n t r i b u t i o n to the r e f l e x response can not be assessed because blood flow to the skin alone has not been measured. The i n d i v i d u a l e f f e c t s of P h 3 and Prop on the r e f l e x have already been studied (Lioy et a l , 1 9 7 4 ) » Therefore we administered the same blockers s e q u e n t i a l l y to eliminate the prominent adrenergic component of the r e f l e x , so that some other components may become unmasked. This procedure may raise some objections because the time l a g between PhB and Prop administrations was 1.5—2 hours, therefore the effe c t s of PhB might have faded away and f a i l e d to provide complete adrenergic blockade. While t h i s was c e r t a i n l y observed i n some experiments, the alpha-adrenergic blocker appeared to be s t i l l e f f e c t i v e 2 - 5 hours a f t e r i t s administration i n most cases, support-i n g the notion that PhB blockade i s slow—acting and pe r s i s t e n t (Nickerson, 1 9 6 2 ) . A f t e r PhB treatment, the r e f l e x response was greatly suppressed and the d i a s t o l i c pressure decreased. IF increased s u b s t a n t i a l l y i n two young male animals and l e s s dramatically i n another group c o n s i s t i n g one male and four female animals. These changes i n d i a s t o l i c 88 pressure and IP were reversed a f t e r the administration of atropine. The suppression of the r e f l e x pressor response was not due to the lower control blood pressure r e s u l t i n g from PhB treatment, since the response was normal at a comparable blood pressure when the animal was not subjected to pharmacological interventions. Moreover, Aars ( 1 9 7 2 ) has demonstrated that PhB i n h i b i t s sympathetic nervous a c t i v i t y independently of p r e v a i l i n g blood pressure. With respect to IP, the causes of the difference i n response between the two groups of animals are not known. Differences due to sex i n the changes i n sympathetic nervous a c t i v i t y have been demonstrated i n cat and dog when the baroreceptor r e f l e x i s evoked by i n f u s i o n of angiotensin and norepinephrine (Morrison & Pickford, 1 9 7 1 ) . We have not investigated the a p p l i c a b i l i t y of t h i s to the TAR. The vasodilatation i n the group with one male and four female animals appeared to be enhanced when AS was performed a f t e r the sequential administration of Prop (Table I I ) . This i s probably due to the slow-acting e f f e c t of PhB, which takes about 30-60 min to reach i t s 89 maximum blocking capacity (Nickerson, 1962). Furthermore, Prop does not possess any sympathomimetic e f f e c t on the i l i a c vasculature (Ahlquist, 19&8). Since IF increased instead of decreased during AS a f t e r Prop, t h i s i n d i r e c t l y indicates that the i l i a c v a s o d i l a t a t i o n was not mediated by the beta-adrenergic system. As f a r as the mechanism responsible f o r the i l i a c v a s o d i l a t a t i o n i s concerned, i t i s u n l i k e l y that i t was caused by further neural i n h i b i t i o n of sympathetic a c t i v i t y , even though AS has been shown to cause a decrease i n a c t i v i t y i n some sympathetic efferent f i b r e s (Pagani et a l , 1974). The v a s o d i l a t a t i o n was d e f i n i t e l y not caused by sudden opening up of arterio-venous anastomoses which are primarly located i n the skin and paw , and r a r e l y within the muscle tissue (Eriksson & Myrhage, 1972), because i t was also observed i n skinned legs. Furthermore, the smooth muscle of the cutaneous anastomoses i s under-alpha-adrenergic control (Spence, Rhodes &. Wagner-, 1972) which had been pharmacologically i n h i b i t e d . I t i s also u n l i k e l y that l o c a l 'autoregulation* was responsible f o r the d i l a t a t i o n , since the change i n i l i a c perfusion pressure, 90 i . e . systemic a r t e r i a l pressure, induced by AS under alpha-adrenergic blockade was n e g l i g i b l e , and should not cause any s i g n i f i c a n t changes i n the diameter of the i l i a c v e s s e l s . Also, i t should not be caused by the accumulation of met-ab o l i c v a s o d i l a t o r s , since no vasoconstriction was observed before the d i l a t a t i o n and therefore presumably no tissue ischemia. F i n a l l y , parasympathetic and dorsal root v a s o d i l a t o r p a r t i c i p a t i o n i n the r e f l e x control of blood flow to s k e l e t a l muscle can be eliminated since there i s no evidence that para-sympathetic f i b r e s innervate the vessels i n t h i s tissue (Folkow, 1 9 5 5 ) , and dorsal root f i b r e s do not convey c e n t r a l l y induced vasodilator impulses (Folkow, Strom & Uvnas, 1 9 5 0 ) . Therefore, i t i s concluded that the i l i a c v a s o dilatation observed was the r e s u l t of a r e f l e x mechanism, probably due to a c t i v a t i o n by AS of sympathetic cholinergic v asodilator f i b r e s since the response was abolished by intravenous i n j e c t i o n of atropine. Other vasodilator mediators such as histamine (Beck, P o l l a r d , Kayaalp & Weiner, 1 9 6 6 ) are not completely ruled out, but apparently they are not involved. 91 Sympathetic cholinergic innervation i n s k e l e t a l muscle of the hindlimb has been demonstrated i n dog (Bulbring & Burn, 1935. Uvnas, i960), and sequently i n several species including cat, fox, sheep and goat; but not i n monkey, r a t and other species (Bolme, Novotny, Uvnas & Wright, 1970). The same innervation i s apparently lacking i n the cutaneous bed of the cat's l e g (Polkow & TJvnas, 1948; Zimmerman, 19^8). In most studies, the cholinergic vasodilator f i b r e s are activated by central nervous stimulation of the s p e c i f i c pathway connecting the cerebral cortex, the hypothalmus and the spina l cord (Korner, 1971), or by peripheral sympathetic nerve stim-u l a t i o n a f t e r alpha-adrenergic blockade (Mauskopf, Gray & Renkln, 19^9)• Sympathetic cholinergic v a s o d i l a t i o n has been e l i c i t e d by emotional s t i m u l i capable of evoking the so-called 'defence reaction' (Lisander, 1970; Mancia et a l , 1972) and by conditioned movements of the legs ( E l l i s o n & Zanchetti, 1971). Evidence of r e f l e x cholinergic v a s o d i l a t a t i o n presumably evoked by intr a t h o r a c i c receptors has been presented by several workers. Jones & Berne (1963) 9 2 have demonstrated that at constant i l i a c perfusion pressure elevation of a o r t i c pressure produced b y blood transfusion or intravenous epinephrine e l i c i t s a three- to f o u r - f o l d increase i n blood flow to the i s o l a t e d and skinned l e g of the dog. This va s o d i l a t a t i o n could be prevented by maintaining a o r t i c pressure at a control l e v e l s by means of a pressure compensator. However, i n determining the neural pathway f o r t h i s r e f l e x d i l a t a t i o n , the sympathetic cholinergic pathway was not seriously considered by these workers because *no evidence could be found that t h i s pathway took part i n vasodilator reflexes e l i c i t e d by stimulation of the depressor nerves, the sinus nerves, or various other afferent nerves' (Jones & Berne, 1 9&3)• Bergel & Makin (1967) have shown i n the dog that ap p l i c a t i o n of nicotine to the surface of the l e f t v e n t r i c l e would i n i t i a t e an increase i n mean femoral flow. Since th i s femoral va s o d i l a t a t i o n i s prevented by atropine, they concluded that the cholinergic v a s odilator system had been r e f l e x l y activated. Another report we are aware of but have no access to i s by Bevan ( 1 9 6 8 ) , who has published 93 an a r t i c l e e n t i t l e d "Reflex v a s o d i l a t a t i o n i n s k e l e t a l muscle following stimulation of i n t r a t h o r a c i c sensory endings". The experimental d e t a i l s of t h i s study are not a v a i l a b l e f o r comparison. There are some important differences between the r e s u l t s of Bergel & Makin (19^7) and the r e s u l t s reported here. The vagi were i n t a c t i n t h e i r preparations and the femoral vasodilatation was accompanied by decreases i n blood pressure and heart r a t e . Moreover, these workers have shown that cooling of the vagi abolishes the r e f l e x , i n d i c a t i n g that the afferent pathway f o r the ' e p i c a r d i a l chemoreflex' i s i n the vagus nerves. In contrast, the present i l i a c v a s o dilatation d e f i n i t e l y does not involve a vagal afferent since a l l experiments are performed i n vagotomized preparations. Since AS induced a decrease i n d i a s t o l i c pressure s i m i l a r to the one observed i n the present experiments i n spinal animals (Lioy et a l , 1974), i t i s possible that the sympathetic cholinergic pathway i s r e f l e x l y activated at the spinal l e v e l . However, we do not have any d i r e c t evidence support t h i s hypothesis. 94 The simultaneous a c t i v a t i o n by AS of sympathetic vasoconstrictor and d i l a t o r pathways i n the cat's l e g appears to be antagonistic i n action. A scrutiny of the d i s t r i b u t i o n of vasoconstrictor and d i l a t o r nerve terminals i n s k e l e t a l muscle vessels indicates that the antagonism i s not as severe as i t may seem. Folkow, Oberg & Rubinstein (19°4) have speculated that the vasodilator nerves innervate predominantly the i n t e r n a l sheath of the muscle wall of the vessel while the vaso-c o n s t r i c t o r nerves innervate i t s external sheath. This i s probably not so because i n cat and dog, both cholinergic and adrenergic nerve terminals have been i d e n t i f i e d i n the same layer, i . e . the a d v e n t i t i a , surrounding the muscular media of the vessels (Bolme & Fuxe, 1970). However, cholinergic nerve terminals, as revealed by acetylcholinesterase s t a i n i n g , are found much les s frequently than adrenergic terminals i n the a d v e n t i t i a . Thus, the observation that a high p r e v a i l i n g vasoconstrictor tone reduces the response on vasodilator nerve stimulation may be explained by t h i s quantitative difference rather than by the r e l a t i v e r a d i a l d i s t r i b u t i o n of adrenergic 95 and cholinergic nerve terminals. Furthermore, the adrenergic terminals are spread over a considerably wide spectrum of vessels both on the a r t e r i a l as well as the venous side, while cholinergic innervation i s confined only to a r t e r i o l a r vessels $0 to 100 micron i n diameter (Bolme & Fuxe, 1970). Therefore, a c o n f l i c t between the action of the two transmitters would probably occur only at t h i s short vascular section where both are present; but because cholinergic innervation does not reach the p r e c a p i l l a r y sphincter section or the capacitance vessels, c a p i l l a r y d i f f u s i o n capacity, flow d i s t r i b u t i o n and resistance are a l l dominantly regulated by the adrenergic system (Mellander & Johansson, 1968). 96 The Superior Mesenteric Flow (SMF) In cats, the organs supplied by the superior mesenteric artery include the pancreas, duodenum, transverse and descending colons, lower part of the ileum and the caecum ( F i e l d & Taylor, 1 9 5 6 ) . A l l of them except the colons and caecum are not drained d i r e c t l y into the venous system as are most organs, but into the p o r t a l vein from which the blood passes through a second set of c a p i l l a r i e s i n the l i v e r before entering the i n f e r i o r vena cava; therefore blood flow i n the superior mesenteric artery may be a l t e r e d by a change i n resistance e i t h e r i n the superior mesenteric or i n the hepatic vascular bed-(Grim, 1 9 6 3 ) . We have made no attempt to separate these two mechanisms i n presenting our r e s u l t s . I t i s generally agreed that the sympathetic adrenergic system has a profound influence on blood flows i n the mesenteric beds (Grayson, 1 9 7 4 ) . The present study provides evidence supporting t h i s view. F i r s t , SMF i n control conditions was about 17 ml/min, which i s considerably lower than IF ( 2 3 ml/min) ; but the organs supplied by the superior mesenteric artery are a l l much 97 more vascularized than the sk e l e t a l muscles i n the l e g (Folkow, Ludgren & Wallentin, 1963; Hulten, 1 9 6 9 ) , therefore i t indicates that the resistance vessels of the superior mesenteric bed maintain a considerable •basal tone*. This becomes clear when a f t e r the admini-s t r a t i o n of PhB, SMP increased s i g n i f i c a n t l y and surpassed IP as i f i t had been 'released* from the vigorous adrenergic vasoconstrictor tone. Secondly, AS i n control conditions induced a marked vasoconstriction i n the superior mesenteric bed. AS a matter of f a c t , the vasoconstriction i n terms of eithe r a decrease i n flow or an increase i n vascular r e s i s t -ance i s larger than those of the i l i a c and renal beds. This suggests an extensive range of sympathetic adrenergic control i n the mesenteric vessels. Furness & Marshall (1974) have observed reduction Of up to 50-70% of the control i n t e r n a l diameter i n p r i n c i p a l a r t e r i e s , and 4 0 - 6 5 % i n small a r t e r i e s and- terminal a r t e r i o l e s , when paravascular nerves i n the rat's mesentery are stimulated at frequency of 4-6 Hz, and by t o p i c a l a p p l i c a t i o n of norepinephrine at concentrations as low as 1 0 ~ ^ g/ml. We conclude that vasoconstriction i n this vascular bed accounts s i g n i f i c a n t l y f o r the pressor response of the TAR. 98 The AS induced resistance change i n the mesenteric "bed i s s i m i l a r to that e l i c i t e d by d i r e c t sympathetic vaso-c o n s t r i c t o r f i b r e stimulation, i n that the phenomenon of "autoregulatory escape" occurs i n both instances. According to previous reports on the i n t e s t i n e (Folkow, Lewis, Lundgren, Mallander & Wallentin, 1964a) and on the colon (Hulten, 1969)» neurogenic c o n s t r i c t i o n i n the mesenteric resistance vessels i s c h a r a c t e r i s t i c a l l y biphasic, with an i n i t i a l peak response followed by a decline to a new steady state of moderately increased resistance, despite continued stimulation. This "auto-regulatory escape" from the vasoconstrictor influence involves a neurogenic (Folkow et a l , 1964b) r e d i s t r i b u t i o n of blood flow from the mucosa towards the low-resistance submucosal structures causing what i s known as 'mucosal blanching' (Dresel & Wallentin, 1966), and i s not the same as the l o c a l l y mediated 'autoregulation' of blood flow observed i n the e x t r i n s i c a l l y denervated i n t e s t i n e (Richardson &. Johnson, 1969). This 'escape' phenomenon was observed i n most of the present experiments, i n which SMF started to return to control l e v e l s before the stretch on the a o r t i c walls was released (e.g. F i g . 5A) . 99 Since the resistance changes during AS do not follow the same temporal pattern i n the d i f f e r e n t experiments, th i s "autoregulatory escape" i s not evident i n F i g . 6B, which shows the average r e s u l t s obtained i n the whole group of animals. The mesenteric vasoconstriction induced by AS was completely abolished by PhB, i n d i c a t i n g that t h i s process was mediated v i a alpha-adrenergic receptors. Since the sequential administrations of Prop and atropine did not cause any further changes i n SMF, AS probably did not induce any beta-adrenergic or cholinergic e f f e c t s . The resu l t s of atropine treatment are compatible with our knowledge of the mesenteric c i r c u l a t i o n . There i s no evidence of sympathetic cholinergic innervation to the cat's mesentery (Folkow, i 9 6 0 ) . Opinions concerning the r o l e of the parasympathetic cholinergic system on the regulation of mesenteric blood flows are not unanimous, but most workers seem to agree that t h i s system probably has l i t t l e d i r e c t influence other than that secondary to augmentation of m o t i l i t y i n the i n t e s t i n e (Grim, 1963; Grayson, 1974). This conclusion i s supported by the r e s u l t s of vagal stimulation (e.g. Kewenter, 19&5) a : n^ L of t o p i c a l acetylcholine a p p l i c a t i o n (Bean & Sidky, 1 9 5 9 ) . 1 0 0 The Renal Flow (RF) Under control situations, RF was about 25 ml/min. This flowrate i s comparable to measurements by cannulating the renal vein and diverting the outflow through an o p t i c a l drop-counter (Kendrick, Oberg & Wennergren, 1972). RF was greater that IF and SMF, and therefore the calculated resistance of the renal bed was lower. This indicates a low 'basal tone', which becomes obvious when a f t e r the administration of PhB, RF increased only s l i g h t l y . The renal vascular tone i s maintained mainly by the sympathetic adrenergic system, since the vessels i n t h i s bed are r i c h l y innervated by these vasoconstrictor f i b r e s . Vasodilator f i b r e s and cholinergic vasotomor fi b r e s do not e x i s t i n the renal vascular bed (Christensen, Lewis & Kuntz, 1951), therefore the administrations of Prop and atropine did not provoke any s i g n i f i c a n t changes i n RF. The two conditions, namely the low 'basal tone' and the r i c h sympathetic innervation, make the kidney an i d e a l organ for increasing vascular x-esistance during r e f l e x a c t i v i t y (Korner, 1974). As shown i n Table IV and F i g . 70, AS induced an increment of renal vascular 101 resistance, since RF changed very l i t t l e i n the experiments while renal perfusion pressure (aortic) consistently increased. Pomeranz, B i r t c h & Barger (196"8) have demon-strated i n conscious and i n anesthetized dogs that mild renal nerve stimulation decreases perfusion of the outer c o r t i c a l peritubular c a p i l l a r i e s and increases that of the outer medulla without a l t e r i n g t o t a l RF. We do not know i f t h i s i s the mechanism responsible f o r constant RF during AS. However, i t i s obvious that the renal vessels constricted and maintained a constant flow i n the face of a higher renal perfusion pressure. This vasoconstriction could be due to ei t h e r nervous a c t i v i t y or to autoregulation. In the f i r s t case AS would have induced an increased discharge i n the renal sympathetic nerves, so that: a vasoconstriction would occur regardless of the p r e v a i l i n g perfusion pressure i n the renal artery. However, even though Pagani et a l (1974) have showed that AS indeed e l i c i t s an increased a c t i v i t y i n some sympathetic e f f e r e n t f i b r e s , t h i s has not yet been demonstrated i n sympathetic efferents to the kidney. 102 Since the change i n renal resistance occurs when the renal perfusion pressure has "been elevated by the TAR from 120 to 150 mmHg, thi s leads to the poss-i b i l i t y that the c o n s t r i c t i o n i s not caused by the pre— mentioned neural r e f l e x mechanism, but rather by a l o c a l autoregulatory mechanism,counteracting the elevated perfusion pressure. The renal vasculature i s known to be capable of adjusting i t s resistance i n response to change i n a r t e r i a l pressure, so that blood flow and glomerular f i l t r a t i o n rate are kept e s s e n t i a l l y constant (Selkurt, 1963; Gelsko & Nissen, 1 9 7 2 ) . These two variables are maintained over somewhat d i f f e r e n t ranges of a r t e r i a l pressure, about 75 to 180 mmHg fo r R F and 100 to 240 mmHg f o r glomerular f i l t r a t i o n rate (Abe, Dixon & McKay, 1 9 7 0 ) . This autoregulation of flow and f i l t r a t i o n i s independent of renal nerves and e x t r i n s i c hormones, but a generally accepted explanation of the phenomenon i s s t i l l l acking. Thus, two mechanisms are available to explain the AS induced renal vasoconstriction. It could be caused by re f l e x sympathetic vasoconstriction as a primary 103 e f f e c t of AS; or i t could be a secondary e f f e c t r e s u l t i n g from the hypertension produced by the TAR. Further experiments with perfusion of the kidney at constant pressure and/or renal denervation are required to e s t a b l i s h the r e l a t i v e contribution of these two mechanisms. Under complete adrenergic blockade, AS again e l i c i t s vasoconstriction i n the renal bed, but i t s magnitude i s much smaller. -The response i s not al t e r e d by atropine (Table I V ) . This can not be s a t i s f a c t o r i l y explained by inadequacy of the alpha-adrenergic blockade, because PhB, Prop and atropine are administered i n sequential \ orders, and the vasoconstriction i s more marked when AS i s performed a f t e r Prop than when a f t e r atropine. No reports of s i m i l a r studies are available f o r comparison, therefore the mechanism responsible remains obscure. 1 0 4 THE CAROTID BARORECEPTOR REFLEX & THE TAR Ef f e c t s of Carotid Occlusion  and Sinus Denervation The generally accepted procedure to prevent influences from the a o r t i c baroreceptors i s to cut t h e i r afferent f i b r e s i n the vagus or the depressor nerves. For the car o t i d baroreceptors, the equivalent procedure i s to section the carotid sinus nerves (CSN); but another much simpler and rountinely used method i s to occlude the common ca r o t i d a r t e r i e s . This l a t t e r procedure probably does not completely eliminate baroreceptor . influences since 'back pressure' from the external and i n t e r n a l c a r o t i d a r t e r i e s may s t i l l exert some s t r a i n on the sinuses. This i s supported by the observations made i n group II (sinus denervated) and group I I I (c a r o t i d occluded) animals of the baroreceptor experiments (Table V I ) . F i r s t , the blood pressure i n group II animals s t a b i l i z e s at a l e v e l much higher than i n group I I I , even though the sinus perfusion pressure (CSP) i s about the same i n both groups. Secondly, the changes i n a r t e r i a l 105 pressure evoked eit h e r by carotid baroreceptor stimulation or by AS are always greater i n magnitude i n group II than group III animals. With the understanding that the main function of the baroreceptors i s to 'buffer* pressure changes i n the c i r c u l a t o r y system, and that the two carotid baroreceptor stations are mutually 'buffering' pressure changes caused by t h e i r a c t i v a t i o n (Heymans & N e i l , 1958)» i t i s conceivable that the baroreceptors on the side of the carotid occlusion r e t a i n some pressure 'buffering' capacity. Thus, the lower control blood pressure and the l e s s marked depressor or pressor responses i n group II I (carotid occluded) animals are probably caused by the influence from these p a r t i a l l y deactivated receptors. Inhibitory E f f e c t s of the Carotid Baroreceptor Reflex  on the TAR The f a c t that the TAR pressor response i s smaller i n the carotid occluded (group III) than i n the sinus denervated (group II) animals gives the f i r s t i n d i c a t i o n i n the present study that the baroreceptor r e f l e x has an 106 i n h i b i t o r y e f f e c t on the TAR. Previously, Lioy et a l (1974) have noticed that the r e f l e x responses are generally moderate i n the presence of normally functioning c a r o t i d baroreceptors but are more pronounced a f t e r carotid occlusion. They too, have suggested that supraspinal baroreceptive mechanisms may exert an i n h i b i t o r y influence on the AS induced r e f l e x changes i n a r t e r i a l pressure, heart rate and LV dP/dt max. The present experiments provide d i r e c t evidence of thi s i n h i b i t i o n . In one series of experiments, the c a r o t i d baro-receptor r e f l e x was activated by di r e c t stimulation of the CSN which carries afferent f i b r e s to the glosso— pharygeal nerve (IXth c r a n i a l nerve), and then projects to supraspinal areas (Korner, 1971). In agreement with the r e s u l t s of Douglas & Schaumann (195&) i n the cat, there was a stimulation frequency (between 5 and 10 Hz) beyond which very l i t t l e or further increase i n response was obtained. The baroreceptor r e f l e x was also activated i n another series of experiments by varying the pressure i n the i s o l a t e d perfused carotid sinus (so-called open-loop experiments). P u l s a t i l e instead of constant flow was 107 employed because the "open-loop gain" G, defined as the r a t i o of change i n femoral artery pressure (FAP, the output pressure) to change i n carotid sinus perfusion pressure (CSP, the input pressure), i s greater when the blood flow to the sinus, i . e . the input pressure, i s p u l s a t i l e (Ead, Green & N e i l , 1952; S p i c k l e r , Kezdi & Ge l l e r , 1967). We have not co l l e c t e d enough data to construct a pressure-characteristic curve (Scher & Young, 1963) r e l a t i n g the output pressure and the input pressure but, i n the CSP range between 100-150 mmHg, G was estimat to be 0.7 i n group II (co n t r a l a t e r a l sinus denervated) and 0.5 i n group I I I (c o n t r a l a t e r a l carotid occluded) animals. These G values are comparable to those reported by Kumada, Nogami & Sagawa (1975) i n the cat under the same anesthesia, but apparently much lower than those found i n the dog (Donald & E d i s , 1971)• This difference due to species may be explained by the observations by N e i l , Hedwood & Schweitzer (1949)* that i n the cat but not the dog chloralose anesthesia diminishes the s e n s i t i v i t y of the vasomotor centre to baroreceptor stimulation , and of the car o t i d sinus to changes of sinus pressure. 1 0 3 The 'escape' phenomenon i n which a r t e r i a l pressure st a r t s returning to control l e v e l despite _p»f continued baroreceptor stimulation was observed i n some experiments ( F i g . 9 ) . The process was usually gradual and slow, and i t s time course resembles that described by Diamond (1955) who has observed that the large increase i n f i r i n g frequency i n the CSN produced by a sudden r i s e i n CSP star t s to subside to a steady l e v e l a f t e r about 1.5 min i n spite of a continued elevation of the sinus pressure. This i s probably due to the r e l a t i v e l y slow adaptation of the a r t e r i a l baroreceptors (Heymans & N e i l , 1 9 5 8 ) . The suppression of the TAR response by the baro-receptor r e f l e x i s not a secondary e f f e c t due to the hypotension produced by baroreceptor stimulation, since the r e f l e x response i s normal at comparable blood pressure achieved by bleeding. Furthermore, the extent of i n h i b i t i o n i s r elated to the CSN stimulation frequency or to the CSP. At maximum baroreceptor stimulation (15 Hz or CSP=205 mmHg), the AS induced pressure r i s e i n group I (CSN stimulation) animals i s about 55% of the control value, and i n the 109 group II and I I I animals, i t i s about 60% and 75% respectively ( F i g . 11 and 1 2 ) . Thus, two points become cl e a r : F i r s t , the TAR response i s suppressed but i s not completely abolished by the baroreceptor r e f l e x ; Secondly, there i s a trend of graded suppression of the AS r e f l e x response corresponds to the number of active or p a r t i a l l y active carotid baroreceptor stations i n the animal groups. Thus, the baroreceptor r e f l e x i n the group I animal i s f u l l y activated, and because i t s ef f e c t s are not •buffered' by any a r t e r i a l baroreceptors, a f u l l strength i n h i b i t i o n on the TAR response i s ensured. On the other hand, the baroreceptor r e f l e x i n the group I I I animals i s hampered by the p a r t i a l l y active baroreceptors on the carotid-occluded side, and therefore i t s i n h i b i t o r y action on the AS response i s r e l a t i v e l y weaker. The i n t e r a c t i o n between the baroreceptor r e f l e x and the TAR could take place at the supraspinal, medullary l e v e l , or at the spinal l e v e l . Compatible with the f i r s t suggestion i s the f a c t that, although the TAR i s a spi n a l r e f l e x , t h i s does not rule out the p o s s i b i l i t y that an afferent component of t h i s r e f l e x projects to the supraspinal 110 l e v e l . This i s known to be the case f o r the spinal component of somatosympathetic reflexes (Sato, 1 9 7 1 ) . Furthermore, Koizumi, S e l l e r , Kaufman & Brooks ( 1 9 7 1 ) have shown that the early spinal phase of the somato-sympathetic r e f l e x i s not influenced by c a r o t i d baro-receptor stimulation, but the l a t e supraspinal phase i s i n h i b i t e d j therefore they concluded that the sympathetic i n h i b i t i o n takes places at the supraspinal medullary l e v e l . This could be the case also f o r the baroreceptor i n h i b i t i o n on the TAR. On the other hand, there i s also some evidence supporting the second suggestion, i . e . the i n h i b i t i o n occurs at the sp i n a l l e v e l . Thus, S e l l e r (1973) has indicated that the preganglionic sympathetic neurons which discharge i n r e l a t i o n to cardiovascular events, are composed of d i f f e r e n t pools, one of which i s influenced mainly through descending pathways from the bra i n stem while another one i s connected mainly with the spinal dorsal afferents. Subsequently, Coote & MacLeod ( 1 9 7 4 b ) have found descending monoaminergic pathways i n the dorso-lateral spinal cord, mediating baroreceptor-induced i n h i b i t i o n on sympathetic a c t i v i t y . The presence 111 of these descending pathways has been confirmed (Trzebski, Lipske, Majcherczyk, Szulczyk & Chruscielewski, 1975); and by means of latency studies, Trzebski et a l suggested that a rather complex multisynaptic organization within the spinal cord neurons i s involved i n the i n h i b i t i o n . Furthermore, contrary to the r e s u l t s of Koizumi et a l (1971), Coote & MacLeod (1974a) were able to p a r t i a l l y reduce the spinal component of the somatosympathetic r e f l e x by CSN e l e c t r i c a l stimulation and by carotid sinus d i s t e n t s i o n . Therefore, i t i s equally possible that the TAR i s modulated by baroreceptor stimulation at the spinal l e v e l . 112 THE CAROTID CHEMORECEPTOR REFLEX & THE TAR Methods of Chemoreceptor Activation I t i s known that the CSN i s a mixed afferent nerve containing afferents from the baroreceptors of the c a r o t i d sinu3 and from the chemoreceptors of the carotid body (Heymans & N e i l , 1958). N e i l , Redwood & Schweitzer (1949) were the f i r s t to demonstrate i n cats anesthetized with choralose that i t i s possible to evoke a depressor response i f the CSN i s stimulated with long duration (0.5-1.0 msec) pulses, and a pressor response i f short duration (0.05-0.1 msec) st i m u l i are applied. These responses presumably r e s u l t from d i f f e r e n t i a l a c t i v a t i o n of baroreceptor and chemoreceptor f i b r e s . Our result s obtained by CSN stimulation i n chloralose-urethan anesthetized cats confirm these observations. There are two factors which make t h i s d i f f e r e n t i a l a c t i v a t i o n of baroreceptor and chemoreceptor f i b r e s possible. The f i r s t i s the i n h i b i t o r y effects of chloralose on the baroreceptor r e f l e x both c e n t r a l l y and p e r i p h e r a l l y . As mentioned b r i e f l y before, N e i l et a l (1949) have provided 113 experimental evidence that chloralose reduces the e f f e c t s of baroreceptor stimulation on the vasomotor centre, and decreases the s e n s i t i v i t y of the baroreceptors to changes of sinus pressure. Also according to these workers, only cats but not dogs or rabbits are affected i n t h i s way by th i s anesthetic. Thus, chloralose anesthesia i n cats would 'unmask' the chemoreceptor r e f l e x from the predominant baroreceptor r e f l e x induced by CSN stimulation. The second factor i s the f i b r e - s i z e of the baroreceptor and chemo-receptor f i b r e s , which determine t h e i r r e l a t i v e threshold f o r a c t i v a t i o n , and account f o r t h e i r difference i n response to the duration of the s t i m u l i . Fidone & Sato ( 1 9 6 9 ) have estimated that i n the cat's CSN, the A-fibre population i s comprised of approximately 2/3 chemore,ceptors and 1/3 baroreceptors f i b r e s , while the reverse i s true 'for the C-fibre population, i . e . 2/3 baroreceptors and 1/3 chemoreceptors f i b r e s . Thus, long duration pulses presum-ably activate the high threshold C-fibres, the majority of which comes from the barore'ceptors, therefore causing a depressor response; while short duration s t i m u l i probably 114 activate only the A - f i b r e s , and because most of these are chemoreceptor afferents, a pressor response i s induced. Douglas & Ritchie (1956) have demonstrated that stimulation of the CSN at i n t e n s i t i e s above the threshold f o r e x c i t i n g C-fibres causes a reduction i n sympathetic vasoconstrictor outflow accompanied by depressor responses. S i m i l a r l y , de Groat & L a l l e y (1974) have shown that stimulation of the A-fibres e l i c i t s r e f l e x discharge i n the thoracic and lumbar sympathetic nerves, and also increases r e s p i r a t i o n and a r t e r i a l blood pressure. The s t i m u l i used to activate the carotid chemo-receptor r e f l e x i n the perfusion experiments are considered to be more physiological since the chemoreceptors respond to changes i n POg* F C ^ 2 3 X 1 ( 1 p H °^ a r - t e r i a l blood. The technique i s c l a s s i c a l and the perfusates used frequently are venous blood, hypoxic saline or other solutions, or hypoxic a r t e r i a l blood from a donor animal (Heymans & N e i l , 1958 ) * A warming device was included i n our perfusion system because i t has been shown that low -temperature perfusate reduces chemoreceptor a c t i v i t y (Bernthal & Weeks, 1 9 3 9 ) . 115 We have e a r l i e r demonstrated that a c t i v a t i o n of the ca r o t i d baroreceptors has an i n h i b i t o r y e f f e c t s on the TAR pressor response. Therefore, i n order to study the v e f f e c t s of chemoreceptor a c t i v a t i o n on thi s spinal r e f l e x , influences from the baroreceptors should be eliminated or maintained constant. Thus, both CSN were cut close to the sinuses i n the nerve stimulation experiments. This procedure eliminated baroreceptor influences from the c o n t r a l a t e r a l baroreceptors when the CSN on one side was stimulated, and also abolished simultaneous e x c i t a t i o n of efferent sympathetic f i b r e s i n the i p s i l a t e r a l CSN going to the baroreceptors and chemoreceptors (Goodman, 1973). For the perfusion experiments, the sinus was perfused at a constant pressure (100-110 mmHg) and the co n t r a l a t e r a l CSN was cut i n a l l animals, therefore the baroreceptor influence must be minimal, since Mancia (1975) has demonstrated i n dogs that the vascular responses to chemoreceptor stimulation are i n h i b i t e d only by high (215 mmHg) pressure i n the sinus. By CSN stimulation or by hypoxic blood perfusion, we obtained an a r t e r i a l mean pressure increase of about 20-45% from the pre-stimulation l e v e l of about 120 mmHg. 116 These r e s u l t s are comparable to those of K i l l i p (1963) who stimulated the CSN i n chloralosed cats with short duration pulses, and to those of L i t t l e & Oberg (1975) who perfused both sinuses simultaneously with venous blood also i n chloralosed cats. However, none of these workers has described the 'escape* of the pressor response during continous chemoreceptor a c t i v a t i o n , probably because the stimulation period i n t h e i r experiments i s r e l a t i v e l y shorter (0.5-1.0 min) as compared to about 2.0-3.0 min i n our experiments. Since the resul t s from the CSN stimulation and the hypoxic blood perfusion experiments were s i m i l a r , we conclude that both methods are equally e f f e c t i v e i n a c t i v a t i n g the chemoreceptor r e f l e x . Inhibitory E f f e c t s of the Carotid Chemoreceptor Reflex  on the TAR Activ a t i o n of the chemoreceptor r e f l e x reduces the TAR response (Table VII). However, as described i n the RESULTS section, there was no di r e c t c o r r e l a t i o n between the i n h i b i t i o n and the extent of stimulation. The r e s u l t s obtained from the two groups of animals (CSN stimulation 117 and hypoxic perfusion) are s i m i l a r , and the average reduction of the TAR pressor response was about 35% i n both groups, supporting the conclusion that both methods of chemoreceptor a c t i v a t i o n are equally e f f e c t i v e . As compared to the baroreceptor r e f l e x which reduces the TAR by as much as 45% during maximum a c t i v a t i o n , the chemoreceptor r e f l e x seems to be s l i g h t l y l ess e f f e c t i v e . This may be due to the f a c t that both chemoreceptors and the TAR activate while the baroreceptors i n h i b i t the sympathetic vasoconstriction system. Another explanation could be the basic functional difference between the baroreceptor and the chemoreceptor reflexes. The most prominent e f f e c t s produced by e x c i t a t i o n of the chemore-ceptors under normal conditions are changes i n r e s p i r a t i o n , while they are not as competent as the baroreceptors i n regulating blood pressure changes (Heymans & N e i l , 1958). During chemoreceptor a c t i v a t i o n , AS was performed at a blood pressure l e v e l higher than that present i n the control s i t u a t i o n . We were able to provide evidence that at l e a s t part of the chemoreceptor " i n h i b i t i o n ^ of the TAR i s caused by this f a c t . Thus, AS evoked smaller than control responses when the systemic a r t e r i a l pressure 118 had been r a i s e d by norepinephrine or by blood tra n s f u s i o n to l e v e l s comparable with those obtained by chemoreceptor stimulation. The reduction of the TAR by both methods of increasing- the blood pressure was about 25% ( F i g . 15C,D). Norepinephrine mimics the pheripheral. e f f e c t s of the chemoreceptor r e f l e x i n causing an increased vascular tone, and so probably l i m i t s the extent of any further v a s o c o n s t r i c t i o n inducible by AS. However, an increase i n blood volume probably does not a f f e c t the vascular tone, therefore the mechanism by which blood volume expansion reduces the TAR response i s not c l e a r . A l a n i s , D e F i l l o & Gordon (1968) have studied the interactions between the chemoreceptor r e f l e x and the somato sympathetic r e f l e x i n dogs. They stimulated the saphenous nerve and observed an increase i n cardiac nerve a c t i v i t y and i n blood pressure. When they repeated the stimulation during a c t i v a t i o n of the c a r o t i d chemoreceptors obtained by i n j e c t i o n of KCN (50 >ig/kg) into the c a r o t i d a r t e r y , the somatosympathetic r e f l e x appeared to be augmented; howerer, no s t a t i s t i c a l treatment of 119 the r e s u l t s was provided. These data would appear to be i n c o n f l i c t with our r e s u l t s , but differences i n animal species and i n experimental technique make the comparsion of the experiments quite d i f f i c u l t . We are not aware of any other reports on the ef f e c t s of the c a r o t i d chemo-receptor r e f l e x on spinal sympathetic r e f l e x e s . 120 CONCLUSIONS We conclude from the present study that the pressor response induced by stretching the walls of the thoracic aorta without obstructing a o r t i c blood flow i s caused, i n part, by active adrenergic vasoconstrictions i n the external i l i a c , the superior mesenteric and the renal vascular beds. The increase i n resistance during AS i s most remarkable i n the superior mesenteric bed, i n d i c a t i n g a s i g n i f i c a n t contribution from t h i s vascular area. There i s also an increase i n the renal resistance, because renal blood flow i s maintained constant i n spite of the increase i n a r t e r i a l pressure. We suggest that the mechanism of renal autoregulation of blood flow may be involved. As to the external i l i a c bed, AS induces a vaso-c o n s t r i c t i o n which i s reversed to vasodilatation i f the alpha-adrenergic response of the TAR i s blocked pharma-c o l o g i c a l l y . Since this i l i a c v asodilatation i 3 abolished by atropine, the p o s s i b i l i t y exists that AS may r e f l e x l y activate the sympathetic cholinergic vasodilator f i b r e s innervating the s k e l e t a l muscles of the cat'c hindlimb. 121 Ve also conclude that the AS induced response i s i n h i b i t e d but i s not abolished by both the carotid baroreceptor and the chemoreceptor reflexes. The TAR response, although smaller, i s s t i l l present at a high l e v e l of baroreceptor and chemoreceptor a c t i v i t y . In comparison with the baroreceptor i n h i b i t i o n , the chemoreceptor i n h i b i t i o n on t h i s spinal r e f l e x appears to be less e f f e c t i v e , and part of this i n h i b i t i o n can be accounted f o r by the increase of the pre-stretch l e v e l of blood pressure produced by the chemoreceptors. The ph y s i o l o g i c a l situations i n which the TAR and the carotid baroreceptor and/or chemoreceptor reflexes can be simultaneously activated are not clear, therefore the significance of the TAR i s not known at present. However, spinal CV reflexes e l i c i t e d from various cardiac and vascular areas have been demonstrated ( f o r reference, see INTRODUCTION), and now we demonstrate that one spi n a l r e f l e x (the TAR) i s i n h i b i t e d but not abolished by supra-spin a l mechanisms. This lends support to the suggestion that nervous control of c i r c u l a t i o n i s obtained through a v a r i e t y of neural c i r c u i t s and that spinal reflexes may represent d i f f u s e , elementary mechanisms(Lioy et a l , 1974). 122 LITERATURE CITED Aars H (1972) Reflex i n h i b i t i o n of sympathetic nerve a c t i v i t y by phenoxybenzamine. Acta phy s i o l . Scand. 85_: 433-437. 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Physiol. 205: 758-744. 61. Korner PI (1971) Integrative neural cardiovascular c o n t r o l . P h y s i o l . Rev. $Vi 312-367. 62. Kumada M, Nogami K & Sagawa (1975) Modulation of carotid sinus baroreceptor r e f l e x by s c i a t i c nerve stimulation. Am. J . P h y s i o l . 228: 1535-1541. 63. Kuntz A (1945) Anatomic and physiologic properties of cutaneo-visceral vasomotor r e f l e x arcs. J . Neuropjysiol. 8: 421-430. 64. Kuntz A & Hazelwood LA (1940) Circulatory reactions i n the g a s t r o - i n t e s t i n a l t r a c t e l i c i t e d by l o c a l i z e d cutaneous stimulation. Am. Heart J . 20: 743-749. 65. Langley JN (1924) Vaso-motor centers. Part I I I . Spinal vascular (and other autonomic) reflexes and the ef f e c t of strychnine on them. J . P h y s i o l . 9^_: 231-258. 132 66. Leitner JM & Pe r l ER (1964) Receptors supplied by-spinal nerves which respond to cardiovascular changes and adrenaline. J . P h y s i o l . 175: 254-274. 67. Lioy F, M a l l i a n i A, Pagani M, Recordati G & Schwartz PJ (1974) Reflex hemodynamic responses i n i t i a t e d from the thoracic aorta. C i r c . Res. _3_4.: 78-84. 68. Lisander B (1970) Factors i n f l u e n c i n g the autonomic component of the defence reaction. Acta p h y s i o l . Scand. Supple. 55 ! » 1-42. 69. L i t t l e R & Oberg B (1975) Circulatory response to stimulation of the car o t i d body chemoreceptors i n the cat. Acta p h y s i o l . Scand. _9_3_: 34-51. 70. M a l l i a n i A, Lombardi F, Pagani M, Recordati G & Schwartz PJ (1975) Spinal cardiovascular reflexes, Brain Res. 87: 259-246. 71. M a l l i a n i A, Pagani M, Recordati G & Schwartz PJ (1971a) Spinal sympathetic reflexes e l i c i t e d by increases i n a r t e r i a l blood pressure. Am J . P h y s i o l . 220: 128-134. 72. M a l l i a n i A, Pagani M, Recordati G & Schwartz PJ (197.1b) Sympathetic reflexes from the heart. In "Cardiovascular regulation i n health and disease" (ed. B a r t o r e l l i C & Zanchetti A), pp. 119-135, Cardiovascular Research I n s t i t u t e , Milan. 133 73. Malliani A, Parks M, Tuckett RP & Brown AM (1973) Reflex increases in heart rate e l i c i t e d by-stimulation of afferent cardiac sympathetic nerve fibres in the cat. Circ. Res. j52: 9-14. 74. Malliani A, Peterson DF, Bishop VS & Brown AM (1972) Spinal sympathetic cardiocardiac reflexes. Circ. Res. _3J): 158-166. 75. Malliani A, Recordati G & Schwartz PJ (1973) Nervous activity of afferent cardiac fibres with a t r i a l and ventricular endings. J. Physiol. 229: 457-469. 76. Malliani A, Recordati G, Schwartz PJ & Pagani M ("1972) Tonic afferent sympathetic activity from the heart, Experientia (Basel) 28: 269-270. 77. Malliani A, Schwartz PJ & Zanchetti A (19&9) A sympathetic reflex e l i c i t e d by experimental coronary occlusion. Am. J, Physiol. 217 ; 703-709. 78 Mancia G (1975) Influence of carotid baroreceptors on vascular responses to carotid chemoreceptor stimulation i n the dog. Circ. Res. _3j6; 270-276. 134 79. Mancia G, B a c c e l l i G & Zanchetti A (1972) Hemodynamic responses to di f f e r e n t emotional stim u l i i n the cat: patterns and mechanisms. Am. J . P h y s i o l . 223.: 925-933. 80. Mauskopf JM, Gray SD & Renkin EM (1969) Transient and persistent components of sympathetic cholinergic v a s o d i l a t a t i o n . Am. J . Phys i o l . 216: 92-97. 81. Mellander S & Johansson B (1968) Control of resistance, exchange, and capacitance functions i n the peripheral c i r c u l a t i o n . Pharmacol. Rev. 20: 117-196. 82. Morrison JFB & Pickford M (1971) Sex differences i n the changes i n sympathetic nerve a c t i v i t y when a r t e r i a l pressure i s raise d by in f u s i o n of angiotensin and noradrenaline. J . P h y s i o l . 216: 69-85. 83. 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Peterson DP & Brown AM (1971) Pressor reflexes produced by stimulation of afferent fibres in the cardiac sympathetic nerves of the cat. Giro. Res. 28: 605-610. Pomeranz BH, Birtch AG & Barger AC (1968) Neural control of intrarenal blood flow. Am. J. Physiol. 21J5_: 1067-1081. 136 91. Richardson DR & Johnson PC (1969) Comparison of autoregulatory escape and autoregulation i n the intest inal vascular "bed. Am. J . Physiol . 217: 586-590. 92. Richins CA & Brizzee K (1949) Effect of local ized cutaneous stimulation on circulat ion in duodenal arterioles and capillary beds. J . Neurophysiol. J_2: 131-136. 93. Sahs AL & Fulton JF (1940) Somatic and autonomic reflexes in spinal monkeys. J . Neurophysiol. 1 : 258-268. 94. Sato A (1971) The spinal and supraspinal somato-sympathetic reflexes. In "Reseach in Physiology" (ed. Kao FF, Vassalle M & Koizumi K) , pp. 507-516. Bologna, Aulo Gaggi. 95. Sato A & Schmidt RF (1973) Somatosympathetic reflexes: Afferent f ibres , central pathways, discharge characterist ic. Physiol . Rev. j5_3_5 916-947. 96. Schaeffer H (1960) Central control of cardiac function. Physiol . Rev. 40 (supple. 4): 213-249. 97. Seher AM & Young AC (1963) Servoanaly3is of carotid sinus reflex effects on peripheral resistance. C i rc . Res. 12: 152-162. 137 98. Selkurt EE (1963) The renal c i r c u l a t i o n . In Hand-book of physiology, section 2, v o l . 2, C i r c u l a t i o n , (ed. Hamilton WF & Dow P) , Am. Ph y s i o l . Society, Washington, D.C, pp. 1457-1516. 99» S e l l e r H (1973) The discharge pattern of singl e units i n thoracic and lumbar white rami i n r e l a t i o n to cardiovascular events. Pflugers Arch. ges. Physi o l . 334: 317-330. 100. Sherrington CS (1906) The integrative action of the nervous system, (pp.241-243)• Archibald Constable & CO., Ltd. London. 101. Snedecor GW & Cochran WG (1973) S t a t i s t i c a l methods. 6th ed., The Iowa State University Press, Iowa. 102. Spence RJ, Rhodes BA & Wagnes EN J r (1972) Regulation of arteriovenous anastomotic and c a p i l l a r y blood flow i n the dog l e g . Am.J. Phy s i o l . 222: 326-329. 103. Spickler JW, Kezdi P & Ge l l e r E (1967) Transfer c h a r a c t e r i s t i c s of the carotid sinus pressure control system. In "Baroreceptors and Hyper-tension" (ed. Kezdi P), Pergamon press, pp. 31-40. 138 a 104. Trzebski A, Lipski J , Majcherczyk S, Szulczyk P & Chruscielewski L (1975) Central organization . and chemoreceptor sympathetic reflex. Brain Res. 87_: 227-237. 105. TJeda H, Uchida Y & Kamisaka K (1969) Distribution and responses of the cardiac sympathetic receptors to mechanically induced circulatory changes. Jap. Heart J . 10;70-81. 106. Uvnas B (1960) Sympathetic vasodilator system and blood flow. Physiol . Rev. 40 (Supple. 4): 69-76. 107. Zimmerman BG- (1968) Comparison of sympathetic vasodilator innervation of lindlimb of the dog and cat. Am. J . Physiol . 214: 62-66. 138 b 22b. Diamond J (1955) Observations on the e x c i t a t i o n by acetylcholine and by pressure of sensory receptors i n the cat's c a r o t i d sinus. J . P h y s i o l . 150; 513-532. 61b. Korner PI (1974) Control of blood flow to spec i a l vascular areas: Brain, kidney, muscle, skin, l i v e r & i n t e s t i n e . In MTP international review of science, Physiology series one, v o l , _1_ (Cardio-vascular Physiology), (eds. Guyton AC • & Horrobin D), pp. 123-162. Univ e r s i t y Park Press, Baltimore. 87b. Pagani M, Schwartz PJ, Bishop VS & M a l l i a n i A (1974) Reflex sympathetic changes i n ao r t i c d i a s t o l i c pressure-diameter r e l a t i o n s h i p curves i n the cat. In "Proceedings of the i n t e r n a t i o n a l union of physiological sciences", v o l . XI, 26th in t e r n a t i o n a l congress, New D e l h i , No. 579, PP. 193. 106b. Wesolowski SA, Pries CC, Sabini AM & Sawyer PN (1965) Significance of turbulence i n hemic systems and i n the d i s t r i b u t i o n of the at h e r o s c l e r o t i c l e s i o n . Surgery sfljt 155-162. 139 APPENDIX A Measurement of A r t e r i a l Blood Flow by the Ult r a s o n i c Doppler Flowmeter: The P r i n c i p l e and the P i t f a l l s The p r i n c i p l e of operation i s the Doppler 1 2 e f f e c t . * Ultrasound energy at a r e p e t i t i o n rate of about 10 m i l l i o n times a second i s focused i n t o the moving blood stream. The wavelength of the ultrasound energy i s so small that a portion of the transmitted energy i s r e f l e c t e d back from the i n d i v i d u a l red c e l l s . When there i s no blood movement, the frequency of the energy r e f l e c t e d back to the re c e i v i n g c r y s t a l i s the same as that which was sent from the transmitting c r y s t a l and there i s no sound. But with blood movement the frequency of the r e f l e c t e d waves i s s l i g h t l y d i f f e r e n t from that transmitted, so a sound i s heard. The p i t c h of the sound depends upon the difference i n frequency between the transmitted and r e f l e c t e d waves, and t h i s frequency ( c a l l e d the Doppler-shift frequency) i s proport-io n a l to the v e l o c i t y of the blood according to the 3 equation: -^ 1 4 0 2 F t (V) cos a whereas F^ = Doppler-shift frequency (Hz) = frequency transmitted - frequency received F, = frequency transmitted (Hz) x V = v e l o c i t y of hlood flow (cm/sec) a = angle between the acoustical axis and the flow of blood 5 C = v e l o c i t y of sound i n blood (1.5x10 cm/sec) The Doppler-shift frequency i s then modulated, amplified and displayed i n an appropriate way. There are c e r t a i n inherent sources of e r r o r i n the u l t r a s o n i c technique f o r measuring blood flow. Inaccuracies may be-introduced by: a) P o s i t i o n i n g of the probe.^ Although the probes used are c a r e f u l l y selected to match the external circum-ference of the vessels to avoid c o n s t r i c t i o n on the one hand and to insure s t a b i l i t y on the other, any motion of the probes or marked changes i n vessel diameter within the transducers ( c r y s t a l s ) would cause inadequate coupling 141 • a of the vessel to the transducers, and a change i n the angle between the acoustical axis and blood flow, such that an a l t e r a t i o n of the Doppler-shift frequency may r e s u l t . \ 2.4 b) V e l o c i t y p r o f i l e of the blood flow. , H Blood flow i n a vessel (or tube) does not have a single value of v e l o c i t y . The v e l o c i t y of blood corpuscles varies with the r a d i a l p o s i t i o n of the corpuscle i n the vessel so that a mixture of Doppler-shift frequencies i s obtained. Some of these frequencies are ' f i l t e r e d * out and i n general, the measurement i s somewhat higher than the true v e l o c i t y of blood flow. c) Changes i n hematocrit. Since the operation of the Doppler system i s dependent upon the presence of suspended p a r t i c u l a t e matter within the f l u i d , i t i s conceivable that a l t e r a t i o n i n the p a r t i c u l a t e concentrate of an inhomogenous solution might a l t e r the frequency of the r e f l e c t e d wave and thus the Doppler-shift frequency. Contradictory r e s u l t s have been reported concerning t h i s point. d) Reversal of blood flow.^ The Doppler flowmeter i s not capable of sensing the d i r e c t i o n of flow. Accordingly, 1 4 2 errors would be introduced whenever reverse flow i s s i g n i f i c a n t . An example i s the measurement of small changes i n flow i n the i l i a c or femoral a r t e r i e s of r e s t i n g dog. e) Method of volume c a l i b r a t i o n . ^ Since c a l i b r a t i o n i s done i n v i t r o , the normal blood v e l o c i t y p r o f i l e has been disrupted. This and other changes i n experimental conditions would c e r t a i n l y induce inaccuracy. In summary, there are l i m i t a t i o n s i n the method, therefore the exact magnitude of the change i n blood flow can not be measured, but any r e l a t i v e change i n flow within a short period of time i s adequately r e f l e c t e d by the Doppler flowmeter. References 1 . Parks E l e c t r o n i c s Laboratory. Mannual f o r Doppler Flowmeter Model 803, 1974. 2. Trotman RE ( 1 9 7 0 ) Ultrasonic Doppler methods i n blood flow studies. Biomed. Engineering j>: 4 5 3 - 4 5 4 » 3. Michie DD & Cain CP ( 1 9 7 1 ) E f f e c t s of hematocrit upon the s h i f t i n Doppler frequency. Proc. Soc. Exp. Bio & Med. H 8 : 7 6 8 - 7 7 2 . 4 . Vatner SF, Franklin D & VanCitter RL ( 1 9 7 0 ) Simul-taneous comparison and c a l i b r a t i o n of the Doppler and electromagnetic flowmeters. J . Appl. P h y s i o l . 2^1 9 0 7 - 9 1 0 . 143 Drugs Acetylcholine bromide Atropine sulphate o-Chloralose F l a x e d i l (gallamine t r i e t h i o d i d e ) Heparin sodium Norepinephrine ,. b i t a r t r a t e Phenoxybenzamine hydrochloride Propranolol " S w hydrochloride Urethan (Ethylcarbamate) APPENDIX B Supplier Eastman Kodak Co., New York, N.Y. BDH Pharmaceuticals, Toronto, Ontario BHH, Poole, England Poulenc Ltd. Montreal, P.Q. N u t r i t i o n a l Biochemicals Corp., Cleveland, Ohio Winthrop, Aurora, Ontario Smith, K l i n e & French, Montreal, P.Q. Ayerst Laboratories, Montreat, P.Q. Matheson Coleman & B e l l , Norwood, Ohio 

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