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The role of vasopressin in the diuretic response to left atrial distension Mason, James Melvin 1971

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THE ROLF, OF VASOPRESSIN IN THE DIURETIC RESPONSE TO LEFT ATRIAL DISTENSION by JAMES MELVIN MASON B.Sc.(Hons.), University of British Columbia, 1969 A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in the Department of Physiology We accept this thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA April 1971 In presenting this thesis in partial fulfilment of the requirements for an advanced degree at the University of British Columbia, I agree that the Library shall make it freely available for reference and study. I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the Head of my Department or by his representatives. It is understood that copying or publication of this thesis for financial gain shall not be allowed without my written permission. Department of The University of British Columbia Vancouver 8, Canada i i "Following our instinct for i n t e l l e c t and knowledge, we acquire pieces of knowledge; and presently, i n the generality of men, there arises the desire to relate these pieces of knowledge ". Matthew Arnold, Discourses i n America (1885). ABSTRACT. The concept t h a t the r e n a l e x c r e t i o n of water and e l e c t r o -l y t e s i s p a r t l y governed by the volume i n some f l u i d compart-ment of the body i s one which has r e c e i v e d some e x p e r i m e n t a l v e r i f i c a t i o n . Attempts t o d e f i n e a mechanism s e n s i t i v e t o changes i n some f l u i d compartment of the body have p r o v i d e d sup-p o r t both f o r and a g a i n s t the theory t h a t s t i m u l a t i o n of sensory nerve endings i n the i n t r a t h o r a c i c c i r c u l a t i o n s e t s up a f f e r e n t impulses i n the vagus nerves which d i m i n i s h the r e l e a s e o f a n t i d i u r e t i c hormone from the neurohypophysis and so cause d i u r e s i s . E vidence which supports the theory comes from e x p e r i -ments i n which a t r i a l d i s t e n s i o n has been a s s o c i a t e d w i t h a r e d u c t i o n of a n t i d i u r e t i c a c t i v i t y i n the c i r c u l a t i n g b l o o d . Evidence which does not support the theory comes from e x p e r i -ments i n which the d i u r e t i c response to l e f t a t r i a l d i s t e n s i o n has been demonstrated d u r i n g i n f u s i o n o f v a s o p r e s s i n a t r a t e s adequate to completely i n h i b i t water d i u r e s i s i n conscious dogs (0.025 m-u./kg./min.). A s e r i e s o f experiments has been c a r r i e d out i n an attempt to d e f i n e the r o l e of a n t i d i u r e t i c hormone i n the d i u r e t i c r e -sponse to l e f t a t r i a l d i s t e n s i o n . In one s e r i e s , experiments were designed to t e s t the e f f e c t s of d i f f e r e n t doses of vaso-p r e s s i n upon the d i u r e t i c response t o l e f t a t r i a l d i s t e n s i o n . The r e s u l t s o f t h i s s e r i e s showed t h a t the d i u r e t i c response to l e f t a t r i a l d i s t e n s i o n was composed of an i n c r e a s e i n s o l u t e e x c r e t i o n and an i n c r e a s e i n water e x c r e t i o n . At a r a t e of i n -f u s i o n o f v a s o p r e s s i n of 0.4 m-u./kg./min. or above the i v i n c r e a s e i n water e x c r e t i o n was a b o l i s h e d w h i l e the i n c r e a s e i n s o l u t e e x c r e t i o n was u n a f f e c t e d . In another s e r i e s , e x p e r i -ments were designed to t e s t the r e n a l response t o l a r g e changes i n v a s o p r e s s i n c o n c e n t r a t i o n . The c o n c e n t r a t i o n s of vas o p r e s -s i n used (0.4 m-u./kg./min. and 0.04 m-u./kg./min.) completely i n h i b i t e d water d i u r e s i s i n con s c i o u s dogs. The r e s u l t s o f t h i s s e r i e s i n d i c a t e d t h a t i n the hydrated a n a e s t h e t i z e d dog, a change from the h i g h c o n c e n t r a t i o n o f v a s o p r e s s i n t o the lower c o n c e n t r a t i o n may cause a t r a n s i e n t d i l u t e d i u r e s i s . These r e s u l t s support the view t h a t a decrease i n the c i r -c u l a t i n g c o n c e n t r a t i o n of a n t i d i u r e t i c hormone i s one mechanism which may produce a d i u r e t i c response t o l e f t a t r i a l d i s t e n s i o n . The r e s u l t s w i l l be r e p o r t e d i n a condensed form (Mason and Ledsome, 1971; Ledsome and Mason, 1971). V TABLE OF CONTENTS. Page. ABSTRACT i i i LIST OF TABLES. v i i i LIST OF FIGURES. i x ACKNOWLEDGEMENTS. x PART I . INTRODUCTION. 1 - 6 . PART II. METHODS. 7 - 23. a. Introduction. 7. b. Anaesthetic and general management. 9. c. Surgical procedures. 10. d. Chemical analysis. 13. e. Calculations. 17. f. Experimental protocol. 17. 1. A t r i a l distension experiments. 17. 2. Half hour infusion experiments. 20. 3. Two hour infusion experiments. 21. g. S t a t i s t i c a l treatment of results. 22. h. Format of presentation of results. 23. PART I I I . RESULTS. 2 4 - 6 5 . a. Effect of l e f t a t r i a l distension. 25. 1. Cardiovascular effects. 25. 2. Effect on urine flow. 34. 3. Effect on urine osmolality. 35. 4. Effect on solute excretion. 36. 5. Effect on free water clearance. 40. 6. Effect of control urine osmolality 42. ) I I I v i Page. on urine composition and flow. 4 2 . b. Effect of changing the rate of vasopressin infusion from 0.4 m-u./kg/irriru to 0.04 m-u./kg/min. for a 30 min. period. 4 6 . 1. Cardiovascular effects. 2. Effect on urine flow. 4 6 . 3. Effect on urine osmol<ality. ^g. 4 . Effect on solute excretion. ^g. 5 . Effect on free water clearance. 5 0 . 6. Effect on glomerular f i l t r a t i o n rate. 5 1 . c. Effect of changing the rate of vasopressin infusion from 0.4 m-u./kg/min. to 0 . 04 m-u./kg/min. for a period of 2 hours. 5 6 . 1. Cardiovascular effects. 5 6 . 2. Effect on urine flow. 5 6 . 3. Effect on urine osmolality. 57. 4. Effect on solute excretion. 57. 5. Effect on free water clearance. 5 8 . 6l Effect on glomerular f i l t r a t i o n rate. 58. d. Urine flow and composition in animals 5 8 . under chloralose anaesthesia. e. Effect of the hydration procedures on'the experimental animal. 6 4 . PART IV. DISCUSSION. 66.- 8 8 . a. Introduction. 6 6 . b. Cardiovascular effects of vasopressin infusion and l e f t a t r i a l distension. 6 6 . 1. A t r i a l distension experiments. 6 6 . 2. Infusion experiments. 6 7 , c. Urinary effects of a t r i a l distension experiments. 6 8 . V l l Page. 1. Assessment of the maximum effective dose of vasopressin i n the a t r i a l distension experiments. 68. 2. Changes i n urinary composition during the a t r i a l distension experiments. 69. d. Urinary effects of vasopressin infusion experiments. 75. 1. 30 min. infusion experiments. 75. 2. 2 hour infusion experiments. 76. e. Evidence against antidiuretic hormone as an agent producing the diuresis to l e f t a t r i a l distension. 77. f. Evidence for antidiuretic hormone as an agent producing the diuresis to l e f t a t r i a l distension. 81. g. The role of vasopressin as controlled by l e f t a t r i a l distension i n the control of body f l u i d volume. 85. 1. Normal function. 85. 2. Abnormal function. 86. h. Conclusions concerning the role of anti-diuretic hormone i n the diuretic response to l e f t a t r i a l distension. 87. PART V. BIBLIOGRAPHY. 89. - 94. / v i i i LIST OF TABLES. Table. Page. I. The number of dogs used and the experiment performed on each dog. 8. II. Effects of l e f t a t r i a l distension upon heart rate, a r t e r i a l pressure, l e f t a t r i a l pressure. 24. III. Effects of l e f t a t r i a l distension upon urinary-excretion. 27. - 29. IV. Effects of vasopressin infusion on urinary excretion. 33. V. Regression analysis. 39/ VI. Effect of control osmolality on urinary excretion. .VO. Effects of changing the rate of vasopressin infusion upon heart rate and blood pressure. 52. VIII. Effects of changing the rate of vasopressin infusion upon urinary excretion. 53 m _ 5/^ IX. Average plasma values for sodium concentration, potassium concentration, and osmolality in the control and experimental periods during the experimental procedures. 61, X. Assessment of the volume load to individual dogs used in the experiments. 62. - 63. i x LIST OF FIGURES. Figure. Page 1. Effects of l e f t a t r i a l distension on urinary-excretion during the infusion of saline and vasopressin in doses of 0.025 m-u./kg./min., 0.1 m-u./kg./min., 0 .4 m-u./kg./min and 1.0 m-u./kg./min. 30. 2. Effects of l e f t a t r i a l distension on urinary excretion in dog number 7 of series one. 31. 3. Effects of l e f t a t r i a l distension on urinary excretion in dog number 4 of series one. 32. 4. Regression analysis of free water clearance vs. osmolal clearance. 38. 5. Effect of an infusion of vasopressin at a rate of 0.025 m-u./kg./min. on urinary excretion. 43. 6. Effects of changing the rate of vasopressin infusion for a 30 min. period. 45. 7. Effects of changing the rate of vasopressin infusion for a 30 min. period in dog number 19. 47. 8. Effects of changing the rate of vasopressin infusion for a 30 min. period in dog number 20. 49. 9. Effects of changing the rate of vasopressin infusion for a 2 hour period. 55. X ACKNOWLEDGEMENTS. Foremost I must thank my supervisor, Professor J.R. Ledsome, for his encouragement, helpful criticism, invaluable advice, and participation i n the experiments. I should also l i k e to thank Mrs. Glenda Bennion for her expert assistance during the experiments and i n the s t a t i s t i c a l treatment of the results. I am grateful to Mr. Kurt Henze for his preparation of the photographic reproductions. Lastly, I should l i k e to thank my wife, Saskia, without whose co-operation and patient understanding I could not have written this thesis. P A R T I. I N T R 0 D U C T I O N . 1. In 1878 Claude Bernard (1878) wrote "La f i x i t e du milieu interieur est l a condition de l a vie l i b r e , inde'pendante: le mecanisme qui l a permet est celui qui assure dans l e milieu interieur l e maintien de toutes les conditions necessaires a l a vie des elements". Starling and Cannon later added to this concept of a constancy to the internal environment. Starling (1909) pointed out that the organism must be provided with distinct mechanisms for the regulation of the amount, the composition ?nd the molecular concentration of the fluids i n the body. Cannon (1932) developed the concept of homeostasis, the existence of steady states in an organism maintained by complex co-ordinated physiological processes. Cannon considered the prime assurance against extensive shifts in a steady state was the provision of "sensitive automatic indicators" which would set in motion corrective processes at the very beginning of a disturbance. That such a mechanism exists to maintain the molecular concentration of the fluids in the body has been demonstrated by Verney (1947). The work of Verney (1947) has established the stringency with which the osmotic pressure of the blood i s maintained by a balance between water drunk, food ingested, urine lost, and sweat evaporated. The existence therefore of a mechanism to maintain the amount of f l u i d in the body has also been sought. The existence of two mechanisms, one controlling osmotic pressure, the other controlling volume should not, imply that regulation of volume must be independent of regulation of osmotic pressure, as these two factors are bound together i n determining the "milieu interieur". However, there are conditions such as excess water intake during enforced sodium loss (McCance, 1936) i n which the homeostasis of osmotic pressure and that of blood volume mutually interfere, and there is now evidence which indicates that in' such conditions constancy of composition may be 2. sacrificed i n favor of constancy of volume (Arndt, 1965; Zehr, Johnson and Moore, 1969). The principle of control of the circulatory blood volume in i t s most general form was f i r s t expressed by J.P. Peters (1935). He stated that "the fullness of the blood stream may provoke the diuretic response on the part of the kidney". His concept received experimental verification when i t was shown that an isosmotic expansion of blood volume w i l l indeed increase urine flow (Welt and Orloff, 1951* Zuidema, Clarke, Reeves, Gauer and Henry, 1956). Gauer and Henry (1963) explored the mechanisms through which a change in kidney function may be provoked i f the "fullness of the blood stream" i s altered. They argued that a sensory limb to register volume would be expected to exist on the low pressure side of the circulation. The low pressure side of the circulation, which included the l e f t ventricle i n diastole, had mechanical properties which established a predictable pressure - volume relationship and therefore had the capability of responding to changes i n blood volume. Biological receptors registering vascular wall tension at any region throughout the low - pressure system could from a physical point of view record the "fullness of the blood stream". The presence of nerve endings in the posterior a t r i a l walls, in the junctional region between the venae cavae and pulmonary veins and the at r i a had been described by Nonidez (1937). Electrophysiological recordings of the rhythmic a c t i v i t y in vagal afferents from the heart have established these nerve endings on the low pressure side of the circulation as receptors (Paintal, 1953; Whitteridge, 194-8). Knowledge with respect to the normal stimulus for these cardiac receptors i s s t i l l limited, however in the case of some l e f t a t r i a l receptors the adequate stimulus appears related to the degree of stretch of the a t r i a l muscle fiber (Paintal, 1953). 3. Thus there existed for the cardiac receptors in the l e f t atrium mechanical and neurophysiological evidence condusive to a role as part of a sensory limb to register blood volume. Gauer, Henry and Sieker (l96l) observed that a wide variety of procedures which changed total blood volume or the distribution of blood had i n common the effect that an increase i n intrathoracic blood volume occurred together with a diuresis while a decrease was always associated with oliguria. Experiments were designed to localize more precisely the area within the intrathoracic circulation responsible for the diuresis. Henry, Gauer and Reeves (1956) achieved this localization by comparing the effects of mitral obstruction with the effects of snaring the extra-pericardial pulmonary veins or with the effects of blocking the pulmonary circulation with multiple emboli. It was found that an effective diuresis was produced only i f the l e f t atrium was included in the congested area. Unfortunately their experimental arrangement did not provide any evidence on right a t r i a l sensitivity nor did i t permit separate distension of the l e f t atrium. Henry et aL (1956) concluded that a sensitive region would appear to be somewhere between the pulmonary vein - l e f t a t r i a l junction and the atrioventricular junction. This area and the corresponding region of the right atrium was the location of the subendocardial stretch receptors of Nonidez (1937) and Paintal (1953). Based on the diuretic response to l e f t a t r i a l distension Gauer and Henry (1963) have proposed that cardiac receptors have a role i n volume regulation. However, the three manoeuvres used by Henry et a l (1956) were not s t r i c t l y comparable since mitral obstruction did not limit the stimulus to receptors i n the l e f t atrium and also obstructed blood flow. It could be argued therefore that stimulation of l e f t a t r i a l receptors by mitral obstruction was not responsible for the diuresis. This argument 4. has been resolved with the demonstration by Ledsome and Linden (1968) that mechanical stimulation of l e f t a t r i a l receptors i s involved in the diuretic response to l e f t a t r i a l distension. These authors showed that inflation of small balloons in the pulmonary vein - l e f t a t r i a l junction which had been reported to vigorously stimulate l e f t a t r i a l receptors to discharge and did not obstruct blood flow through the atrium (Kidd, Ledsome and Linden, 1966) caused a diuresis with characteristics similar in a l l respects, except i t s magnitude, to the diuretic response to mitral obstruction. Furthermore, the diuretic response to mitral obstruction in their exper-iments was abolished by vagotomy. Thus the results of Ledsome and Linden (1968) supported the view that mechanical stimulation of l e f t a t r i a l receptors can cause diuresis and that such stimulation i s a major factor in the production of the diuretic response to mitral obstruction. The fact that distension of the l e f t atrium does produce an unequivocal diuretic response has been confirmed i n the case of a t r i a l balloon i n f l a t i o n by Shu'ayb, Moran and Zimmermann (1965), Johnson, Moore and Segar (1969) and by Ledsome, Linden and O'Connor (1961) and i n the case of a t r i a l distension by a mitral snare by Lydtin and Hamilton (1964) and by Shu'ayb et al. (1965) . The nature of the efferent mechanism (s) acting on the kidney to produce the diuresis to l e f t a t r i a l distension i s uncertain. Gauer and Henry (1963) argued that the time course and dilution of the urine characteristic of the diuretic response to l e f t a t r i a l distension was compatible with a decrease in the rate of release of antidiuretic hormone from the neurohypophysis. They supported the theory that stimulation of sensory nerve endings i n the intrathoracic circulation sets up afferent impulses i n the vagus nerves which diminish the release of antidiuretic hormone from the neurohypophysis and so cause diuresis (Gauer and Henry, 1963; Gauer, Henry and Behn,-1970). Indirect evidence 5. i n support of this view has come from the investigation by Carswell, Hainsworth and Ledsome (1970). The results of Carswell et a l . (1970) indicated that the diuretic response to l e f t a t r i a l distension was unlikely to be due to changes in pH, P ™ , P Q , or haematocrit of the a r t e r i a l blood ^ 2 but that a blood borne agent was involved. Share (1965) has reported evidence linking l e f t a t r i a l distension with the circulating level of antidiuretic activity. He demonstrated that a marked elevation of plasma antidiuretic a c t i v i t y on occlusion of the common carotid arteries was blocked by simultaneous i n f l a t i o n of a balloon in the l e f t atrium which raised l e f t a t r i a l pressure approximately 20 cm. H2O. His balloon inf l a t i o n was ineffective i n the vagotomized dog. Several other workers have shown a relationship between distension of the l e f t atrium, a decrease i n the circulating level of antidiuretic a c t i v i t y and a diuresis (Zehr et a l . 1969; Shu'ayb et a l . , 1965; Johnson et a l . , 1969). Zehr et a l . (1969) observed this relationship i n the unanesthetized ewe while Johnson et a l . (1969) and Shu'ayb et a l . (1965) observed i t i n anesthetized dogs. Johnson et a l . (1969) reported results that indicates the level of plasma antidiuretic a c t i v i t y decreased linearly with increasing l e f t a t r i a l pressure up to 7 cm. H2O. They did not report increases i n l e f t a t r i a l pressure beyond 7 cm. H2O. Thus there existed evidence for antidiuretic hormone as an agent producing the diuretic response to l e f t a t r i a l distension. However, there also existed evidence against antidiuretic hormone acting as an agent producing the diuretic response to l e f t a t r i a l distension. Ledsome et a l . (1961) and Lydtin and Hamilton (1964) demonstrated a dilute diuresis in response to l e f t a t r i a l distension during an infusion of vaso-pressin at a rate of 0.025 m-u./kg./min. This rate of infusion of vaso-pressin had been demonstrated to be at least two times greater than that necessary to completely inhibit water diuresis i n a conscious dog (Ledsome 6. et al., 1961) and the results were interpreted (Ledsome et al., 196l) as making unlikely the argument that the diuretic response to l e f t a t r i a l distension was due to a decrease i n the rate of release of antidiuretic hormone from the neurohypophysis. Further evidence against antidiuretic hormone acting as an efferent agent has been the demonstration that the diuretic response i s transient despite continued distension of the l e f t atrium (Henry et al . , 1 9 5 6 ; Ledsome et a l . , 1961; Lydtin and Hamilton, 1964; Shu'ayb et a l . , 196$) and despite a continued reduction i n blood anti-diuretic activity during the distension (Shm'ayb et a l . , 1965; Figs. 2 and 3 ) . Thus the role of antidiuretic hormone i n the diuretic response to l e f t a t r i a l distension was uncertain. Available evidence established a relationship whereby l e f t a t r i a l pressure may control the circulating level of antidiuretic hormone while other evidence appeared to have established that the circulating level of antidiuretic hormone did not affect the diuresis but that a blood borne agent was involved. Further uncertainty was placed on the function of a reduced circulating level of antidiuretic hormone by the demonstration of Arndt, Reineck and Gauer (1963) of an increase i n osmolal clearance i n response to l e f t a t r i a l distension which suggested that a renal hemodynamic factor may be operative. It was i n the hope of elucidating the true, role of antidiuretic hormone i n the diuretic response to l e f t a t r i a l distension that the following experiments were undertaken. P A R T II. M E T H O D S . 7. A. INTRODUCTION. The experiments to be described are an extension of those performed by Ledsome, Linden and O'Connor (1961). In the present experiments three procedures were used. F i r s t l y , the mitral o r i f i c e was partially blocked by a balloon during an infusion of either saline or vasopressin in doses of 0.025 m-u./kg./min., 0.1 m-u./kg./min., 0.4 m-u./kg./min., or 1.0 m-u./kg./ min. ( a t r i a l distension experiments). Secondly, dogs were infused with vasopressin at a rate of 0.4 m-u./kg./min. and this rate was reduced to 0.04 m-u./kg./min. for periods of 30 minutes (half hour infusion experiments). Thirdly, dogs received each dose of vasopressin, 0.4 m-u./kg./min. and 0.04 m-u./kg./min. for two hours at a time (two hour infusion experiments). Measurements of urinary and cardiovascular variables were made during these procedures. 8. Dog No. Experiment. 1. Series one a t r i a l distension. 2. it ti n II 3. II n II II 4. II II II II 5. ti it I I II 6. it n I I it 7. II tt it n 8. it II II it 9. it I I ti ti 10. 30 min. infusion experiment. 11. Series two a t r i a l distension. 12. 30 min. infusion experiment. 13. II it ti it 14. Series two a t r i a l distension. 15. 30 min. infusion experiment. 16. it tt ti ti 17. Series two a t r i a l distension. 18. 30 min. infusion experiment. 19. II it ti it 20. ti I I it ti 21. 2 hour infusion experiment. 22. ti it tt it 23. it it it ti 24. ti II II tt 25. Series two a t r i a l distension. 26. 2 hour infusion experiment. 27. Series two a t r i a l distension. 28. it tt it u 29. it tt tt tt 30. it it n ti 31. I I it I I it 32. it it tt it 33. n ti it it Table I. The number of dogs used and the experiment performed on each dog. 9. B. ANAESTHETIC AND GENERAL MANAGEMENT. Mongrel dogs of 12 to 26 kg. were used. The dogs were fed a diet of ground beef supplemented with canned dog food (Dr. Ballards') for two to eight days prior to the experiment. The dogs were deprived of food 24 hours prior to the experiment but were allowed water ad libetura. Dogs numbered 1 - 16, 18, 25, 32, 33 were premedicated with morphine sulphate (0.5 mg/kg.) by subcutaneous injection one hour before the anesthetic was given. The remaining dogs (17, 19-24, 26-31) received no premedication. The o animals were anesthetized by the slow intravenous infusion of a warm (65 C) solution of 1$ chloralose (British Drug Houses; dose 10 ml. = 0.1 g/kg.) in sodium chloride solution 0.6 g/100 ml. The anesthetic was delivered through a catheter inserted under local anesthesia (Carbocaine 1%) into the saphenous vein to the level of the in f e r i o r vena cava. Usually, no additional anesthetic was needed during the surgical procedures which were normally completed i n 1 to 2 hours. Occasionally some dogs were restless and developed pronounced reflex muscular twitching i n which case a supplementary dose of anesthetic was given. Subsequently, during the experimental procedure a steady state of light anesthesia was maintained by the infusion every ten minutes of either 1$,cfeloralose or 0.6$ saline, approximately 1 ml/kg. The anesthetic was usually just enough to prevent muscular movements. However, the animal would show a reflex jerk when the table was tapped and i t was the magnitude of this reflex jerk which formed the criterion for the level of anesthesia. The chloralose solution was prepared by dissolving 5 g. of chloralose i n 500 ml. of 0.6% sodium chloride solution and placing this in a water o bath maintained at 65 C. The chloralose was kept at this temperature throughout the experiment and the bottle stoppered to prevent evaporation. Even at this temperature there was a tendency for the chloralose to come 10. out of solution and each dose was f i l t e r e d immediately before use. Dogs numbered 12 - 15, 17 - 20, 22 - 26, 29 were given intramuscular injections of 5 mg. desoxycorticosterone acetate (Ciba Co.) every day for at least one day and not more than three days prior to the experiment. C. SURGICAL PROCEDURES. When the animal had been anesthetized hair was shaved from the flanks, back, chest and neck. A l l experiments were acute and therefore no s t e r i l e precautions were taken. No antibiotics were given either before or during the experiment. The animal was placed on his back on the operating table and the trachea was cannulated.All i n i t i a l chest, neck, and flank surgery was performed using a Birtcher electro-surgical unit model 755. Each ureter was approached through a muscle-splitting flank incision 1 cm. above and parallel to the i l i a c crest. By opening the peritoneal cavity the ureters could be immediately identified in most experiments and were catheterized without significant blood loss. Polyethylene tubing of 1 mm. bore was used to catheterize the ureters; the end of the tubing was blunted to avoid damaging the ureteric epithelium. Each catheter was inserted approximately 4 inches up the ureter. The dead space i n each catheter was usually 1 ml. or less. The incisions were closed in layers and the catheters brought out through the incisions. In the series of experiments i n which the chest was opened ( a t r i a l distension experiments) intermittent positive pressure ventilation was provided with a mixture of 40$ oxygen in a i r from a Harvard Respiratory pump. The stroke volume and rate of respiration were adjusted to about equal that of the animal's spontaneous respiration. A resistance to expiration of 3 cm. water was provided to ensure adequate inf l a t i o n of the 11. lungs. Thoracotomy was performed in the l e f t f i f t h intercostal space. Bleeding was carefully controlled using the electro-surgical unit. The ribs were retracted and the lobes of the l e f t lung carefully pushed to one side so that the pericardium was exposed. The pericardium was incised in the area covering the t i p of the l e f t a t r i a l appendage. The l e f t a t r i a l appendage was gripped with forceps and pulled up through the pericardial incision. A soft string snare was placed around the base of the appendage and tightened. The t i p of the appendage was removed, the cut edges secured with forceps and a balloon inserted into the appendage, the snare was then loosened and the balloon pushed on into the atrium. A ligature around the t i p of the appendage held the balloon i n position. The a t r i a l balloon was made from a finger cot tied over the end of a polyethylene tube of 2 mm. bore. A second polyethylene tube of 1 mm. bore was tied alongside the balloon for the measurement of l e f t a t r i a l pressure. The balloon was judged to be sati s f a c t o r i l y placed when i t lay with i t s base included i n the ligature around the t i p of the a t r i a l appendage. The chest was not closed but the incision was covered over with a cloth which was kept moist for the duration of the experiment. After completion of thoracotomy approximately one hour was allowed to elapse before the f i r s t experimental tests. o Esophageal temperature was maintained between 36 - 38 C by a temperature control unit, (Yellow Springs Inc.). A wide bore metal cannula was placed i n the l e f t femoral artery for the recording of a r t e r i a l blood pressure and obtaining blood samples. A poly-ethylene tube of 1 mm. bore was inserted into the l e f t femoral vein and the experimental infusions of vasopressin or saline were given through this cannula. In experiments i n which glomerular f i l t r a t i o n rate was measured in u l i n was infused through a cannula of 1 mm. bore inserted into the l e f t external jugular vein. In the experiments in which glomerular f i l t r a t i o n rate was measured i t was sometimes necessary to cannulate the right femoral artery for obtaining blood samples when d i f f i c u l t y was experienced in collecting blood samples from the l e f t femoral a r t e r i a l cannula. In some experiments a polyethylene cannula of 2 mm. bore was inserted via the right external jugular vein into the superior vena cava for a record of central venous pressure. Femoral a r t e r i a l pressure, mean l e f t a t r i a l pressure and central venous pressure were measured using Statham P 23 Gb strain gauge manometers connected to a D.C. amplifier with the record displayed on an ultra-violet recorder (Honeywell visicorder oscillograph model 1508). The frequency response of the system recording femoral a r t e r i a l pressure, obtained by the method of Hansen (1949) was f l a t (+ $%) to better than 35 c/s. Mean pressures v/ere obtained e l e c t r i c a l l y . The manometers were calibrated in a stopwise manner using mercury and saline manometers. Zero pressure for the a t r i a l and superior vena caval manometers was recorded post-mortem as pressure at the cannula t i p with the t i p free i n a i r at the same level in which i t lay in the animal. Heart rate was measured from the E.C.G. The E.C.G. was recorded from two chest leads with the signal amplified (Grass P15 A.C. preamp.) displayed on an oscilloscope (Textronix dual-beam type RM 5 6 5 ) and recorded on an ultra-violet recorder. Immediately after completion of surgery and at intervals during the procedures samples of a r t e r i a l blood were taken and pH, PQQJ and P Q w e r e measured. The measurements were made using an Instrumentation Laboratory Inc. pH-blood gas analyzer model 113 - 51 and displayed on Instrumentation Laboratory Inc. Delta-matic pH/mV electrometer's model 245. The electrodes were calibrated before and after each measurement using a pH standard of 13. 7.382 and standard gas mixtures of 02 12.25$ and CO2 5.03$. Throughout the experiment the CO2 concentration of expired a i r was monitored using a Beckman Medical Gas Analyzer model LB-1. Appropriate adjustments were made to the respiratory pump or small infusions (10 - 15 m-equiv.) of sodium bicarbonate solution ( IN.) were given to maintain i f a c O j between 35 and 40 mm Hg. and a r t e r i a l pH within the range 7.3-7.4; no adjustments were made during the control and experimental periods. During surgery Dextran (Travenol, 6% Gentran 75 i n 0.9$ sodium chloride) was infused intravenously in an amount equal to 10$ of the animals estimated blood volume. D. CHEMICAL ANALYSIS. Urine and plasma were analyzed for sodium and potassium using an Instrumentation Laboratory Inc. flame photometer model 143. This flame photometer u t i l i z e d an internal lithium standard (15 mEq.. Li/l»). As an index of both sodium and potassium concentration, the emission intensities of sodium and potassium were electronically ratioed against the emission intensity of the internal lithium standard. A b u i l t - i n analog computer provided the ratioed internal standard operation with direct readout via d i g i t a l servo-counters. As calibrated i n the present experiments the readout was displayed directly i n milliequivalents of sodium and potassium per l i t e r i n the original sample. A l l dilutions of samples for sodium and potassium analysis were made using an Automatic Fisher Diluter model 240. A l l samples and standards were diluted . 1/200 using -25 u l . of sample in 5 ml. of lithium standard. The automatic diluter was calibrated photometrically using Ferroin ( l , 10-Phenanthroline ferrous sulphate complex, Fisher Chemicals) and a Spectromic "20". The calibration was repeated u n t i l diluter-14. dispensed colorimetric dilutions agreed to within 0.025 absorbance units on the Spectromic "20" scale. The flame photometer was recalibrated after every 8 to 10 sample readings using the 100 mEq./L.Na and 100 mEq./L.K standard for urine and the 140 mEq./L.Na and 5 mEq./L.K standard for plasma. When recalibrating i f the standard reading did not agree to within - 5% of i t s known con-centration the preceding sample measurements were redetermined. Generally, the readjustment necessary when recalibrating with the standards was 1 or 2 mEq./L. Duplicate determinations were made on a l l samples and the value was recorded i f the determinations agreed to within + 5$. The average difference between duplicate determinations was found to be 2.26 mEq./L. Plasma and urine osmolality was measured using the method of freezing point depression, (Osmette, precision osmometer). The Osmette u t i l i z e s the techniques of supercooling and then freezing the sample through vibration. A thermistor probe was used to measure the temperature of the sample during the entire process. By proper calibration of the instrument a direct d i g i t a l readout in milliosmols per kilogram was obtained. The calibration of the Osmette was checked twice during the a t r i a l distension experiments using the company supplied standards of 100 m.osm./kg. and 500 m.osm./kg. Using these standards the calibration had not changed for this period of time which extended from May 1969 to July 1969. The next series of experiments began i n September and the Osmette was recalibrated the f i r s t week of October using prepared standard osmolalities of 100 m.osm./ kg., 500 m.osm./kg., 700 m.osm./kg. and 1100 m.osm./kg. and 1300 m.osm./kg. The calibration of the Osmette was then rechecked before every experiment and was found not to have changed. The Osmette was calibrated using successive determinations on the f u l l range of standards. The Osmette was accepted as calibrated i f duplicate 1 5 . readings agreed to within t 5 m.osm./kg. A sample reading was recorded i f duplicate determinations agreed to with-i n - 10 m.osm./kg. The average difference between duplicate determinations on 80 randomly selected values was found to be 4.44 m.osm./kg. Urine volumes were measured in graduated cylinders at the time the urine was collected. If the urine volume for the 10 min. collection period was less than 10 ml. the urine was measured i n 10 ml. graduated cylinders the accuracy of which i s ± 0.05 ml. If the urine volume exceeded 10 ml. i n 10 min. the urine was measured i n 25 ml. graduated cylinders the accuracy of which i s - 0.1 ml. Inulin was measured colorimetrically using the method of Roe, Epstein, and Goldstein, (1949). The basis of the method is the hydrolysis of inulin to fructose which forms a colored compound with resorcinol. The colorimetric reagent used was resorcinol-thiourea reagent (0.1 gm.$ resorcinol plus 0.25 gm.% thiourea i n gl a c i a l acetic acid; Fisher Chemicals). Hydrolysis was accomplished using 30% HC1 (Fisher Chemicals). The plasma and urine deproteinizing reagents were 10% ZnSO^. 7H2O (Fisher Chemicals) and 0.5 N.NaOH (Fisher Chemicals). The deproteinizing reagents neutralized each other volume for volume when t i t r a t i o n was performed with phenol-phthalein as indicator. Standard inulin solutions were prepared of 0.4 mg.$, 0.6 mg.%, 0.8 mg.%, 1 mg.%, 1.25 mg.%, 2.0 mg.% and 3-0 mg.%. The procedure used was to take 0.5 ml. of plasma or urine to which was added 7.5 ml. d i s t i l l e d water, 1 ml. of 10% ZnS0^.7H20 and 1 ml. of 0.5 N.NaOH. This mixture was centrifuged and 2 ml. of the supernatant was decanted and treated i n the case of plasma and standards with 1 ml. of resorcinol-thiourea reagent and 7 ml. of 30$ HC1. In the case of urine i t was necessary to dilute the supernatant to contain 2.0 mg.$ to 10.0 mg.% of i n u l i n and then treat as for plasma above. The mixtures were then placed in a water bath at 80°c. for exactly 10 minutes. They were then cooled by immersing in tap water in the dark for 5 minutes and were read at 520 mu. within 30 minutes on a Spectromic "20". Error of measurement of inulin clearance arises from two main sources: error arising from the experimental protocol such as timing, washout, changing plasma levels, plasma sampling and changing renal function; and errors i n chemical quantitation. Errors i n chemical quantitation were minimized by incorporating standards, reagent blanks, and plasma and urine blanks into each analysis. Measurements were also made on duplicate samples with agreement between duplicate determinations being approximately one-half a division on the absorbance scale of the Spectronic "20" This agreement between duplicate determinations represents an error of approximately - 5% i n determining inulin concentrations. Such an error can magnify i t s e l f to an error of t 10% when computing urine/plasma ratios. The probability of this happening however i s very low. Wesson (1957) found that the sigma of the errors involved i n measuring successive U/P ratios on the same urine and plasma samples was approximately 4.3 F e E' cent for inu l i n . Davies and Shock (1950) also report a sigma of 5 to 7 per cent for the variation of a single inulin clearance about the mean of a series. The main source of error due to experimental protocol arose from a changing plasma concentration of inul i n . The standard deviation from the mean i n four experiments was 19%. Although this figure i s rather high a changing level of plasma inulin concentration i s also recognized by Smith (1956) with his statement; "Fairly constant plasma concentrations are generally obtained ". The significance of a changing level of plasma inulin concentration results from the extrapolation which must be made to determine the plasma concentration which i s representative of the mean during the collection period. This l a t t e r fact combined with the errors due to chemical quantitation result i n changes in G.F.R. of only 10% or more as being significant. This i s in agreement with the stated limits of significance as quoted by Wesson (1957) of from 6% to 20%. E. CALCULATIONS. In expressing the results the following calculations were used. 1) . Osmolal clearance (Cosm). Cosm (ml./min.) = V (urine flow, ml./min.) x U (Urine osmolality.mosm./kg.) P (plasma osmolalityjiriosm./kg.). 2) . Free water clearance (C ^ Q ) . CH2O (ml./min.) = V (urine flow,ml./min.) - Cosm. (osmolal clearance,ml./min.) 3) . Inulin clearance ( C i n) C j n = V (urine flow.ml./min.) x U (urine cone, mg./ml.). P (plasma cone.,mg./ml.). 4) . % of Na i n total osmolal excretion = Na (mEq./min). x 100 V (urine flow,ml./min.) x U (urine cone,mosm. F. EXPERIMENTAL PROTOCOL. 1. A t r i a l Distension Experiments. The a t r i a l distension experiments were performed in two series. In the f i r s t series (dogs 1 - 9 ) saline solution (0.6 gm.$) or vasopressin in doses of 0.1 m-u./kg./min. or 1.0 m-u./kg./min. was infused at a rate of 1 ml./min. In the second series (dogs 11, 14, 17, 25, 27 - 33) saline solution (0.6 gm.%) or vasopressin in doses of 0.025 m-u./kg./min. or 0.4 m-u./kg./min. was infused, also at a rate of 1 ml./min. The order of infusion was randomized in each series. In dogs 7, 8 and 9 a priming dose of inulin (50 mg./kg) was given and an infusion of inulin ( l ml./miri.) was begun in an amount calculated from Smith (1956) to maintain a plasma level of approximately 25 mg.%. Urine volumes, femoral a r t e r i a l pressure, and mean l e f t a t r i a l pressure were measured at 10 minute intervals. The urine samples from the two kidneys were pooled after the 10 minute collection i f their volume were similar (approximately + 25$). It was found from the f i r s t two experiments that such pooling reflected the quantitative characteristics of either sample. 4 ml. blood samples were withdrawn every 20 minutes and replaced with dextran. The a t r i a l balloon was inflated during each infusion with enough 0.6$ sodium chloride solution (usually about 1 ml./kg. body weight) to cause a rise of l e f t a t r i a l pressure of about 20 cm. water. The f i r s t i n f l a t i o n came after a period of steady urine flow had been obtained. This period was usually 40 minutes after the start of the f i r s t infusion. Left a t r i a l distension was maintained for 30 minutes and the infusion continued for 40 minutes after release of the distension. The rate of infusion of vasopressin was then changed and after a further 50 minutes a second distension was performed. The third distension was performed after similar time intervals. A 90 minute interval elapsed between successive balloon distensions. The experimental, protocol is diagrammed below: 19. cont. exp. cont. i 1 i 1 1 t 1 f 1 \ t balloon change change inflated infusion infusion Each ver t i c a l division marks one 10 minute period. After surgery the experiment was completed in approximately 6 hours. The f i r s t control period was taken as the three 10 minute periods immediately prior to balloon distension. The experimental period was taken as the second and third 10 minute periods after balloon in f l a t i o n and the f i r s t 10 minute period after balloon deflation. The second control period was taken as the three 10 minute periods immediately following the experimental period. 20. 2. Half Hour Infusion Experiments . The protocol for the half hour infusion experiments paralleled the a t r i a l distension experiments with the following exceptions - the rates of vasopressin infusion were 0 . 4 m-u./kg./min. and 0.04 m-u./kg-/min., a balloon was not employed nor was the surgery associated with balloon emplacement performed. Dogs 10, 12, 13, 15, 16 received inulin i n an amount previously described for the a t r i a l distension experiments. The eight dogs received an infusion of vasopressin at a rate of 0 . 4 m-u./kg./min. for 90 min. The concentration of vasopressin being infused was then reduced decreasing the rate of infusion to 0 .04 m-u./kg./min. for a period of 30 min. after which infusion at the previous concentration was restored. This procedure was repeated at least twice and not more than 3 times in each dog. The protocol i s diagrammed below. cont. exp. cont. i—J r — i J L _ 1 1 I 0.4 0.04 0.4 Rate of Vasopressin Infusion in m-u./kg./min. Each v e r t i c a l division marks one 10 minute period. 21. The experiment was completed in approximately 6§ hours after surgery. The f i r s t control period was taken as the last three 10 minute periods of the 0.4 m-u./kg./min. infusion of vasopressin. The experimental period was taken as the 2nd and 3rd periods a.fter the beginning of the 0.04 m-u./kg./min. infusion of vasopressin and the subsequent f i r s t period after the resumption of the 0.4 m-u./kg./min. infusion of vasopressin. The second control period was taken as the three 10 minute periods immediately following the experimental period. In a l l infusions 5 minutes or less elapsed between the end of one infusion and the beginning of the next. 3. Two Hour Infusion Experiments. The two hour infusion experiments paralleled the half hour infusion experiment with the exception that both doses of vasopressin (0.4 m-u./kg./ min. and 0.04 m-u./kg./min. ) were infused for a 2 hour period. The five dogs received alternating infusions of 0.4 m-u./kg./min. and 0.04 m-u./kg./ min. for 2 hours at each rate. In these experiments the f i r s t infusion of vasopressin at a rate of 0.4 m-u./kg./min. was started 30 min. after the surgical procedures - were completed. The experimental protocol i s diagrammed below. cont. exp. cont. r 1 i 1 i \ 1 t t t 0.4 0.04 0.4 Rate of Vasopressin Infusion in m-u./kg./min. Each ver t i c a l division marks one 10 minute period. 22. The f i r s t and second control periods were taken as the last six 10 min. periods of the f i r s t and second infusions of 0 .4 m-u./kg./min of vaso-pressin. The experimental period was taken as the 4th to 9th 10 minute period from the beginning of the 0.04 m-u./kg./min. infusion of vasopressin. In the 2 hour infusion experiments dogs numbered 22, 23, 24, 26 received inulin i n an amount previously described for the a t r i a l distension experiments. G. STATISTICAL TREATMENT OF RESULTS. The paired results were analyzed using the t - test for paired data. The mean of the experimental period was compared with the mean of the two control periods. Differences were considered significant i f 2P< 0 . 1 0 . The choice of a two tailed significance was made because experimental changes in either direction from the control values would be considered significant. Furthermore a 2PX0.10 level of significance was chosen since i n the biological system under investigation a 90$ or greater probability that the effects were due to the present experimental design and not chance seemed reasonable. The unpaired results were analyzed using the t - test for unpaired data. Hovever, only those results v/ere compared in which the variances were shown not to be significantly different by the F - test. In the a t r i a l distension experiments results from the two series were compared only i f the results during infusion of saline could be pooled and the results during infusion of saline were pooled only i f the control values during the infusion of saline i n both series of experiments were shown using the t - test for unpaired date not to be significantly different. In Table V and Fig. 4 the results are analyzed as a linear regression. The calculation for the linear regression uti l i z e d the least squares method while significance tests and 95 per cent confidence limits u t i l i z e d the t -test. The complete regression analysis used i n the present investigation is described i n detail i n Sokal and Rohlf (1969): H. PRESENTATION OF RESULTS. In the report of the effects of l e f t a t r i a l distension to follow those experiments in which saline was infused may serve as a control with which to compare the effects of the vasopressin infusions. Changes in the measured variables w i l l be presented by comparing the average experimental value (E) with the average of the two control values (C). In preparing graphs the scale for urine flow in the a t r i a l distension experiments and the 30 min. infusion experiments i s the same as that given in Fig. 1, throughout. The other scales of osmolal clearance (Cosm.), free water clearance ( C J^Q) and urine osmolality were adjusted out of necessity to accomodate the variety of changes observed i n the individual experiments. P A R T III. R E S U L T S . TABLE II. Rate of Infusion of Vasopressin (m-u./kg./min). 1 Heart ± S ™ d Rate n 2P beats/min. SALINE C E 137 180 10.1 9.8 43 17 <0.001 C E 134 191 15.0 9.6 56.5 10 <0.005 C E 139 194 12.9 16 55.6 9 <0.001 C E 124 173 12.6 8.8 49.3 10 <0.C®5 C E 109 168 14.7 12.9 59 9 <0.001 Arterial X +SEM Pressure d n mm. Hg. 2P 135 123 4.6 5.2 -11.8 17 <0.001 139 128 2.8 2.9 -10.8 10 <0.005 129 120 6.3 7.C -9.6 9 <0.10 143 130 4.2 4.4 -12.8 10 <0.001 134 126 4.2 7.0 -8.2 9 NS X Left ±SEM a A t r i a l n Pressure cm. HjO 11 34 0.8 1.9 22.1 17 13 35 1.1 1.4 21.8 10 12 30 0.6 2.C 17.9 9 15 35 1.0 1.6 19.8 10 13 35 1.5 1-5 21.2 9 Table II. Effects of l e f t a t r i a l distension upon heart rate, a r t e r i a l pressure, l e f t a t r i a l pressure. X i s mean SEM i s standard error of the mean; d i s the difference between control and experimental: n = number of tests; 2P i s the level of significance. A. EFFECT OF LEFT ATRIAL DISTENSION. 25. 1. Cardiovascular effects. Inflation of the balloon in the l e f t atrium caused an immediate rise in mean l e f t a t r i a l pressure (Table II). It was necessary to record mean l e f t a t r i a l pressure rather than pulsatile l e f t a t r i a l pressure since oscillations in the pressure record produced from the flexible catheter i n the beating heart obscured the pulsatile pressure recording. In practice there was an increase in mean l e f t a t r i a l pressure of between 5 cm. HjO and 43 cm. H20, from a mean of 14 cm. H2O before in f l a t i o n to 34 cm. H20 during in f l a t i o n . The infusions of vasopressin had no significant effect on the control mean l e f t a t r i a l pressure. Once the increased pressure in the l e f t atrium was established i t tended to remain constant; adjustments in balloon distension of a few mis. during the 30 min. period were necessary on a few occasions only. Upon deflation of the balloon mean l e f t a t r i a l pressure f e l l promptly to previous control levels. When a balloon was inflated i n the l e f t atrium a r t e r i a l pressure usually f e l l rapidly by between 20 mm. Hg. to 90 mm. Hg. However, i t recovered within one to two minutes to reach a level somewhat lower than the pressure before i n f l a t i o n . The a r t e r i a l pressure remained at this lowered level for the duration of the balloon in f l a t i o n (Table II). Individual changes i n a r t e r i a l pressure during balloon i n f l a t i o n ranged from a f a l l of 29 mm. Hg. to a rise of 22 mm. Hg. After deflation of the balloon the a r t e r i a l pressure rose to about i t s pre-inflation level. The infusions of vasopressin had no significant effect on the a r t e r i a l pressure either before, during or after balloon inflat i o n . The heart rate in these experiments was between 54 and 234 beats/min. during the control periods and increased to about 181 beats/min. when the 26. balloon was inflated in the l e f t atrium. The mean increase in heart rate for a l l the experiments was 53 beats/min. (Table II). When the balloon was deflated heart rate did not immediately return to i t s former level but remained elevated about 20 beats/min. over the mean of the f i r s t control period. Infusion of vasopressin had no significant effect on heart rate during either the control or experimental periods. Inflation of a balloon in the l e f t atrium therefore caused a rise in mean l e f t a t r i a l pressure, a f a l l in a r t e r i a l pressure and an increase in heart rate. TABLE III. Rate of Infusion of Vasopressin (m-u./kg./min.). Urine X Flow SEM ml./min. . 2P C E 1.25 1 . 9 1 0.21 0.29 0.65 17 CO.02 <J . \J< j C E 1.03 1.39 0.20 0.24 0.37 10 <0.02 C E 0;79 0.97 0.19 0.24 0.18 9 <0.10 C ' E 0.92 1.14 0.13 0.19 0.22 10 NS C E 0.63 0.71 0.17 0.18 0.08 9 <0.025 Urine X Osmolality S E ^ m.osm./kg n 2P 616 437 88 78 -177 17 <0.005 717 536 115 93 -180 10 <0.01 819 757 130 129 -62 9 <0.05 835 786 101 84 -48 10 NS 896 865 110 108 -31 9 NS Osmolal x Clearance S ^ ml./min. n 2P 1.58 1.81 0.13 0.18 0.23 17 <0.05 1.73 2.02 0.24 0.29 0.29 10 <0.05 1.73 1.86 0.31 0.34 0.13 9 NS 2.26 2.72 0.30 0.44 0.46 10 <0.10 1.69 1.86 0.40 0.38 0:17 9 <0.05 Free X Water SEM Clearance °* ml./min. 2 p -0.32 +0.10 0.24 0.34 0.42 17 <0.05 -0.70 -0.63 •0.24 C0.35 0.08 10 NS -0.94 -0.88 0.22 0.78 0.10 9 NS -1.36 -1.58 0.21 0.27 -0.23 10 <0.05 -1.06 -1.14 0.25 0.22 -0.08 9 NS TABLE III. Sodium Excretion * juEq/mirr SEM d Series 1. n 2P; C E 134 '149 :27 41 15 8 NS * *s ~- S C E C E 131 151 43 44 20 9 < 0.10 C E C E 138 154 54 55 16 9 < 0.10 1 SEM d Series 2. n 2P 63 68 14 16 5 9 NS 107 127 26 29 21 10 NS 176 218 30 50 43 10 NS Potassium ^ SEM Excretion °-n jiEq./min. 2P 56 59 7 8 3 17 NS 63 67 8 8 4 10 <0.05 54 55 6 7 2 9 NS 91 94 12 11 3 10 NS 56 60 7 7 4 o < 0.10 Na./K- x SEM a Series 1. n 2P 2.42 2.46 0.32 0.4£ 0.04 8 NS 2.33 2.59 0.56 0.57 0.26 9 <0.05 2.17 2.36 0.68 0.68 0.18 9 NS TABLE III. SALINE , 0.025 , 0^1 0.4 , 1^0 X SEM Series 2. ^ 2P C E 1.47 1.68 0.43 0.52 0.21 9 NS C E 1.77 1.92 0.37 0.36 0.15 10 NS C E C E 2.03 2.33 0.44 0.51 0.30 10 <0.05 C E Percentage X of Sodium SEM in Total d Osmolal n Excretion 2P Series 1. 25.2 23.5 2.33 3.00 -1.9 8 NS 22.8 25.2 3.07 3.37 2.5 9 <0.05 25.2 25.2 2.92 3.65 0.00 9 NS X SEM d Series 2. n 2P 13.7 13.9 2.80 2.99 0.2 9 NS 18.7 19.7 2.91 2.79 1.0 10 NS 23.2 24.7 2.89 2.88 1.5 10 <0.05 Table III. Effects of l e f t a t r i a l distension upon urinary excretion. Symbols as in Table II. 30. Vasopress in Infusion: m U / k g . / m i n . | SoIine *\(0.0250.1 ->|<- 0.4 ->-|«— I 0 - * | ml. /min. 'H 20 C o s m ml. /min. Osmolal i ty m o s m / k g . 5 0 0 . Urine Flow ml./min. l l ( l m l l l l T i l i l o i 1 ) ^ V ' " 1 10 min. intervals Fig. 1. Effects of l e f t a t r i a l distension on urinary excretion during the infusion of saline and vasopressin in doses of 0.025 m-u./kg./min., 0.1 m-u./kg./min., 0.4 m-u./kg./min. and 1.0 m-u./kg./min. Period of l e f t a t r i a l distension i s indicated by the solid bar. Each horizontal line in a 10 min. interval represents the average value from the number of tests (shown in parenthesis below the abscissa) performed during the infusion. Dotted- lines indicate a change in the rate of vasopressin infusion. From above downwards free water clearance i n ml./min., osmolal clearance in ml./min., urine osmolality in m.osm./kg. and urine flow in ml./min. 31. Vasopressin infusion: mU/kg./min. U H 2 0 ml./min. -2.0 -Cosm ml./min. Osmolality m oem / kg. Urine Flow ml./mfn. 1 2 3 4 5 Time in Hours after End of Surgery Fig. 2. Effects of l e f t a t r i a l distension on urinary excretion in dog number 7 of series one. Conventions as in Fig. 1. 3 2 . Vasopressin Infusion: mil /kg. /min. <—1.0-H< Saline >U—0.1—>\ 4.0 -i o.o i 1 1 1 1 1 r I 2 3 4 5 6 7 Time in Hours after End of Surgery Fig. 3 . Effects of l e f t a t r i a l distension on urinary excretion i n dog number 4 of series one. Conventions as in Fig. 1 . TABLE IV. Jrj 7lc .ne Urine Osmol a l i t y 3smcx 21eai Lai -a.nce ?ree I feter Clearance Sodium ibccretion Series 1 Sodium Excretion Series 2 Potassium Excretion Ma/K Series 1 Ha/K Series 2 Percentage Df Sodium Ln Total Dsmolal Excretion 3eries 1. Percentage Df Sodium Ln Total Dsmolal Excretion 3eries 2. C E C E C E C E C E C E C E C E C E C E C E Saline:0.025 - X -Saline:0.1 - ; XX XX XXX - - X - - - - - - - -Saline:0.4 - — XX XXX XXX XX XXX XXX XXX XX XXX XX X — X XXX Saline:1.0 XX XXX XXXX - - X X - - - - - - - -0.025:0.1* - - - - - - - — — 0.025:0.4 - XX XXX XXX X XXX XXX XXX XXX XXX XXX - X XXX 0.025:1.0* - XX - XX - - - - - -0.1: 0.4* — — — — — — — X XX XX 0.1: 1.0 - - XX - - - - - - - - - - -0.4: 1.0* - - - - - - - XX XX Table IV. Effects of vasopressin infusion on urinary excretion. Control values (C) and experimental values (E) given in table III are compared between the pairs of infusions shown on the l e f t . 0.025, 0.1, 0.4 and 1.0 indicate rate of infusion of vasopressin i n m-u./kg./min. A line (-) indicates no-significant difference while (X), (XX), (XXX), (XXXX) indicate significant differencesof 2P<0.10, 2P<0.Q5, 2P<0.01 and 2P<0.001 respectively. A blank indicates that a comparison was not j u s t i f i e d . An asterisk (*) denotes pairs which were compared using the t - test for unpaired data. A l l other pairs were compared using the t - test for paired data. 2. Effect on urine flow. The results during infusion of saline in the two series of experiments have been pooled since the values obtained during the control periods i n the series were not significantly different. Inflation of a balloon in the l e f t atrium led to an increase in urine flow while deflation of the balloon led to a decrease in urine flow. The increase i n urine flow usually began in the f i r s t ten minute period after a t r i a l distension. The peak rate of urine flow was usually reached i n the last period of inflation or the f i r s t period of deflation (Fig. 1). A significant increase in urine flow in response to a t r i a l distension occured during vasopressin infusion at each dose except 0.4 m-u./kg./min. (Table III). Infusion of vasopressin had no significant effect on the urine flow during the control periods. However, urine flow i n the experimental period during the infusion of saline was significantly greater than that during the infusions of vasopressin at rates of 0.1 m-u./kg./min. and 1.0 m-u. /kg./min. (Table IV). Furthermore, the urine flow i n the experimental period during the infusion of vasopressin at a rate of 0.025 m-u./kg./min. was significantly greater than that during the infusion of vasopressin at a rate of 1.0 m-u./kg./min. Otherwise, there were no further significant effects of infusion of vasopressin upon the urine flow i n the experimental period. The size of the diuretic response varied greatly from dog to dog. Fig. 2 and Fig. 3 are two experiments which show this variation. As shown in the figures the diuretic response to l e f t a t r i a l distension occured whether urine flow during the control period was low (Fig. 2) or high (Fig. 3). Inflation of a. balloon in the l e f t atrium therefore led to an increase in urine flow. The infusions of vasopressin had no significant effect upon the average control urine flow compared with that during the infusion of saline. However, the increase in urine flow in response to l e f t a t r i a l 35. distension was significantly reduced by infusion of vasopressin at a rate of 0,1 m-u./kg./min. or above. 3. Effect on Urine Osmolality. The results during infusion of saline i n the two series of experiments have been pooled since the values obtained during the control periods i n the series were not significantly different. Inflation of a balloon in the l e f t atrium led to a decrease in urine osmolality ^^'hile deflation of the balloon led to an increase in urine osmolality. Urine osmolality usually began to decrease i n the f i r s t ten minute period following balloon in f l a t i o n and usually reached i t s lowest value in the last period of infl a t i o n or the f i r s t period of deflation (Fig. 1). The decrease in urine osmolality i n response to a t r i a l distension i s shown i n Table III. Significant decreases in urine osmolality occurred during the infusion of saline and vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. The urine osmolality recorded in the experimental period and i n the control period during the infusion of saline was significantly less than that recorded during the infusion of vasopressin at a rate of 0.1 m-u./kg./ rain, or above. During the infusion of vasopressin at a rate of 0.4 m-u./kg./ min. the control and the experimental osmolality v/as significantly greater than that observed during the infusion of vasopressin at a rate of 0.025 m-u. /kg./min., Furthermore, during infusion of vasopressin at a rate of.1.0 m-u./ kg./min. the experimental osmolality was significantly greater than that observed during the infusions of vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. (Table IV). The changes in urine osmolality also showed a wide variation i n response to l e f t a t r i a l distension. In Fig. 3 during the infusion of saline when urine osmolality was already very low no change i n urine osmolality was observed during a t r i a l distension. However, a decrease of 1000 m.osm./kg. 3:6. was recorded during infusion of saline in the experiment shown in Fig. 2. Inflation of a balloon i n the l e f t atrium therefore led to a decrease in urine osmolality. This decrease in urine osmolality however was prevented during infusion of vasopressin at a rate of 0.4 m-u./kg./min. or above. The urine osmolality recorded i n the control and/or experimental periods during the infusions of saline and vasopressin at rates of 0.1 m-u./kg./min. or below was significantly less than that recorded during infusions of vaso-pressin at rates of 0.4 m-u./kg./min. or above. 4. Effect on Solute Excretion. The results of measurements of osmolal clearance and potassium excretion during infusion of saline in the two series of experiments have been pooled since the values obtained during the control periods in the series were not significantly different. However, the results of measure-ments of sodium excretion, Na./K %ratio, and the percentage of sodium in the total osmolal excretion have not been pooled since the values obtained during the control periods i n the series were significantly different. Inflation of a balloon i n the l e f t atrium led to an increase in osmolal clearance while deflation of the balloon led to a decrease i n osmolal clearance. Osmolal clearance usually began increasing in the f i r s t ten minute period after balloon inflation and usually reached a peak while the balloon was s t i l l inflated (Fig. 1). The increase i n osmolal clearance was consistently significant with the exception of the value recorded during the vasopressin infusion at a rate of 0.1 m-u./kg./min. (Table III). Although the osmolal clearance increased significantly the individual cations did not necessarily increase significantly (Table III). During the infusion of saline there vere no significant increases in sodium or potassium excretion, the Na./K ratio or the percentage of sodium in the total osmolal excretion. 3'7. During the infusion of vasopressin at a rate of 0.02$ m^u./kg./min. only-potassium excretion increased significantly. During the infusion of vaso-pressin at a rate of 0.1 m-u./kg./min. only sodium excretion increased significantly with a consequent significant increase in the Na./K, ratio and the percentage of sodium in the total osmolal excretion. During the infusion of vasopressin at a rate of 0.4 m-u./kg./min. the percentage of sodium comprising the total osmolal excretion and the Na./K. ratio increased significantly. While during the infusion of vasopressin at a rate of 1.0 m-u./kg./min. there was no significant change i n the percentage of sodium comprising the total osmolal excretion but there was a significant increase i n both sodium and potassium excretion and consequently no change i n the Na./K, ratio. While i t was necessary to determine whether the infusions of vaso-pressin had any effect on solute excretion i t was not j u s t i f i e d , as previously indicated, to make comparisons between series one and series two for sodium excretion, Na./K* ratio, and the percentage of sodium i n the tot a l osmolal excretion. However, when a l l j u s t i f i e d comparisons were made the variables of solute excretion differed significantly between infusions only when vasopressin was infused at a rate of 0.4 m-u./kg./min. (Table IV). Thus in f l a t i o n of a balloon in the l e f t atrium led to an increase in osmolal clearance during infusions of both saline and vasopressin. Furthermore, this increase was not augmented by any specific and consistent significant'increase i n sodium or potassium excretion nor was i t consistently altered by vasopressin. 38. t l . O r 1.0 2.0 3.0 Osmolal Clearance (ml./min.) Fig. 4. Regression analysis of freewater clearance vs. osmolal clearance. The relationship between free water clearance, plotted on the ordinate in ml./min., and osmolal clearance plotted on the abscissa i n ml./min. during the infusions. Each point on the graph represents the value for a 10 min. period of urine collection during the infusions of saline (•), 0.025 m-u./kg./min. (o), 0.1 m-u./kg./min. (&), 0.4 m-u./kg./min. ( a ) and 1.0 m-u./kg./min. ( O ) . The calculated regression line (solid l i n e ) i s drawn through the values recorded during the vasopressin infusions of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. The 95 per cent confidence interval i s shown (dotted line) and the regression line i s extrapolated back to 0.0 osmolal clearance. TABLE V. Regression Analysis. 1.0 m-u./kg./min. 0.4 m-u./kg./min. 0.012 = (-1.070) - (-0.629)(1.721) -0.119 = (-1.436) - (0.545)(2.418) b l " b2 = °- 0 84 n = 20 t = 0.0158 2P = NS Combined. -0.142 = (-1.253) - (-0.537)(2.069) n = 20 t = 23.8 2P =< 0.001 Table V. Calculated regression equations from which the regression line in Fig. 4 i s drawn. The regression equations for the vasopressin infusions of 1.0 m-u./kg./min. and 0.4 m-u./kg./min. are given as well as their combined regression equations. b]_ - b 2 indicates the difference between the two regression slopes and "n" the number of 10 min. urine collection periods on which the t value (t) and significant level (2P) are based. Equation i s the form of a = I - bX where "a" is the intercept on the ordinate, "b" i s the slope and Y and X are the mean values of free water clearance and osmolal clearance respectively. vo *-o ;40. 5* Effect on Free Water Clearance. The results during infusion of saline i n the two series of experiments have been pooled since the values obtained during the control periods in the series were not significantly different. Inflation of a balloon in the l e f t atrium led to an increase i n free water clearance during infusion of saline and vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. and a decrease i n free water clearance during infusion of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. Significant changes however occurred only during the infusions of saline and vasopressin at a rate of 0.4 m-u./kg./min. (Table III). Free water clearance usually decreased in the f i r s t ten minute period following a t r i a l distension. During those infusions i n which free water clearance increased the i n i t i a l decrease was followed by an increase which began i n the second ten minute period after a t r i a l distension whereas during the infusion of vasopressin at a rate 0.4 m-u./kg./min. free water clearance continued to decrease. The increase or decrease reached a peak during the last period of balloon in f l a t i o n or the f i r s t period after balloon deflation (Fig. 1). During infusion of vasopressin at a rate of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. free water clearance varied inversely to osmolal clearance (Fig. 4). No significant relationship could be established between free water clearance and osmolal clearance during infusions of saline and vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. However, during infusion of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. a significant functional relationship was found to exist between free water clearance and osmolal clearance. Fig. 4 shows the linear regression line for the two vasopressin doses of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. One regression lin e for the two infusions was ju s t i f i e d on the basis that there was no significant difference between the 41. individual slopes of the regression lines for the two infusions. The regression line was extrapolated to zero osmolal clearance. The 95$ confidence limits are also shown in Fig. 4. The highly significant regression line within the 95$ confidence interval achieves zero free water clearance at zero osmolal clearance. The regression analysis i s summerized in Table V. Free water clearance during the control and experimental periods was significantly greater during infusion of saline than that observed during infusion of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. Free water clearance during the control and experimental periods was significantly greater during infusion of vasopressin at a rate of 0.025 m-u./ kg./min. than that observed during infusion of vasopressin at a rate of 0.4 m-u./kg./min. Free water clearance during the experimental period was significantly greater during infusion of saline than that observed during infusion of vasopressin at a rate of 0.1 m-u./kg./min. Free water clear-ance during the experimental period was also significantly greater during infusion of vasopressin at a rate of 0.1 m-u./kg./min. than that observed during infusion of vasopressin at a rate of 0.4 m-u./kg./min. (Table IV). While infusion of vasopressin at a rate of 0.025 m-u./kg./min. did not significantly affect the free water clearance recorded i n the control and experimental period during infusion of saline i t was capable in some dogs of producing antidiuresis. One such experiment i s shown i n Fig. 5. In Fig. 5 infusion of saline was followed by a dilute diuresis as indicated by a urine flow of approximately 3 ml./min. and a urine concentration of approximately 130 mosm./kg. Infusion of vasopressin at a rate of 0.025 m-u./kg./min. was followed by an antidiuresis as indicated by a urine flow of approximately 0.5 ml./min. and a urine concentration of approximately 600 mosm./kg. After the infusion of vasopressin was stopped urine flow and concentration returned to previous diuretic levels. Thus inflation of a balloon in the l e f t atrium led to an increase in free water clearance during infusion of saline. Infusion of vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. prevented a significant increase i n free water clearance while infusion of vasopressin at rates of 0.4 m-u./kg./min. and above abolished the increase in free water clearance. The free water clearance recorded in the control and/or experimental periods during infusions of saline and vasopressin at a rate of 0.025 m-u./ kg./min. was significantly greater than that recorded during infusion of vasopressin at rates of 0.1 m-u./kg./min. and above. 6. Effects of Control Urine Osmolality on Urine Composition and Flow. In Table VI each balloon inflation i s classified according to i t s urine osmolality (Hiring the control period. Urine flow increased significantly in a l l classifications with the exception of the range from 750 - 1000 m.osm./ kg. Osmolal clearance during the experimental period was slig h t l y decreased in the range from 0 - 250 m.osm./kg. but increased with higher control osmolalities. The largest increase in osmolal clearance occurred when the control osmolality was over 1000 m.osm./kg. Free water clearance increased in the experimental period in the range from -0 - 750 m.osm./kg. and decreased i n the range over 750 m.osm./kg. A significant increase in free water clearance was recorded in the range from 0 - 250 m.osm./kg. and a significant decrease was recorded i n the range over 1000 miosm./kg. A significant decrease in urine osmolality during the experimental period was recorded i n a l l classifications. A3. Osmolality m osm/kg. Cosm ml./min. ^H20 ml./min. Urine Flow ml./min. Vasopressin Vasopressin 0.025 m-u 0.025 m-u /kg./min. Saline /kg./min. Saline 0.0 3 4 5 6 7 Time in Hours after End of Surgery Fig. 5. Effect of an infusion of vasopressin at a. rate of 0.025 m-u./ kg./min. on urinary excretion. Conventions asin Fie. 1. 44. U. VOL. Cosm. CH20 URINE 0-250 I SEM m.osm./kg. 3 n 2P C E 2.53 3.14 0.29 0.24 0.53 5 <0.02 C E 1.56 1.54 0.12 0.11 -0.02 5 NS C E 0.96 1.60 0.29 0.25 0.63 5 <0.001 C E 213 146 19.4 15.7 67 5 <0.005 250-500 X SEM m.osm./kg. 3 n 2P 1.18 1.67 0.13 0.28 0.49 8 <0.025 1.34 1.46 0.13 0.12 0.13 8 NS -0.20 0.20 0.12 0.23 0.41 8 NS 416 312 23.0 39.6 104 8 <0.10 500 - 750 X SEM m.osm./kg. d n 2P 1.07 1.47 0.08 0.16 0.40 20 <0.02 2.25 2.52 0.22 0.24 0.27 20 <0.005 -1.18 -1.05 0.15 0.24 0.13 20 NS 643 564 19.3 42.7 79 20 <0.05 750 - 1000 X SEM m.osm./kg. 3 n 2P 0.62 0.74 0.09 0.17 0.12 10 NS 1.76 2.00 0.27 0.45 0.24 10 NS -1.14 -1.26 0.18 0.28 -0.12 10 NS 843 774 25.0 38.7 69 10 <0.05 1000 -SEM m.osm./kg. 3 n 2P 0.32 0.58 0.05 0.11 0.26 12 <0.01 1.36 1.81 0.17 0.24 0.45 12 <0.01 -1.04 -1.23 0.13 0.15 -0.18 12 <0.1 1311 1087 428 88.1 224 12 <0.005 Table VI. Effect of control osmolality on urinary excretion. The variables of urine flow (ml./min.), osmolal clearance (ml./min), free water clearance (ml./min.) and urine osmolality (m.osm./kg.) are grouped according to the range of osmolality shown on the l e f t within which the control osmolality was measured. Conventions as in Table II. Vasopressin Infusion : mU/kg./min. I I i< o.4 H*-0.04-»| mm Hg 1 6 0 " Arterial Pr. I40H Heart Rate beats/min °H 20 rnl /min C osm ml/min. Osmolality m osm/kg 1.6 -Urine 1.4 -Volume 1.2 -ml./min. 1.0 -0.8 10 min. intervals Fig. 6. Effects of changing the rate of vasopressin infusion for a thirty minute period. Each horizontal lin e i n a 10 min. interval represents the average value from twenty-one tests in eight dogs. From above downwards femoral a r t e r i a l pressure (mm.Hg.), heart rate (beats/min), free water clearance (ml./min.), osmolal clearance (ml./min.), urine osmolality (m.osm./kg.) urine flow (ml./min.), Dotted lines indicate a change in the rate of vasopressin infusion. 46. B. EFFECTS OF CHANGING THE RATE OF  VASOPRESSIN INFUSION FROM 0.4  m-u./kg./min. (HIGH RATE) T0"0.04  m-u./kg./min. (LOW RATE) FOR A  PERIOD OF 30 MINUTES. 1. Cardiovascular Effects. Changing the rate of vasopressin infusion from the high rate to the low rate led to an increase i n heart rate and a decrease in a r t e r i a l pressure (Table VII). Following the change to the low rate heart rate increased i n the f i r s t 10 minute period and continued to increase reaching a peak in the second or third 10 minute period. Heart rate decreased in the f i r s t 10 minute period following resumption of the high rate but did not return to previous control levels (Fig. 6). Arterial pressure decreased in the f i r s t 10 minute period following the change to the low rate and remained at this decreased level u n t i l resumption of the high rate, at which time a r t e r i a l pressure immediately increased to approximately the previous control level (Fig. 6). Thus decreasing the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. led to an increase i n heart rate and a decrease i n a r t e r i a l pressure. 2. Effects on Urine Flow. Decreasing the rate of vasopressin infusion from the high rate to the low rate led to an increase i n urine flow (Table III). There was an increase i n urine flow i n the f i r s t 10 minute period following the change to the low rate. This increase usually reached a peak in the f i r s t 10 minute period following resumption of the high rate and then declined to previous control levels (Fig. 6). Although urine flow increased from an average value during the control periods of 1.26 ml./min to an average value during the experimental periods of 1.48 Ul. i I i i i Vasopressin Infusion: mU/kg. /mln. 0-0 I i i i i i i i i i i i i i i i i i i i i i i i i i i i l i i i i i i i 10 min. Intervals Fig. 7. Effects of changing the rate of vasopressin infusion for a 30 min. period in dog number 19. Conventions as in Fig. 6. 48. ml./min. i n three of the dogs urine flow during an experimental period reached a peak greater than 3 ml./min. One such experiment i s shown i n Fig. 7. In six tests i n two dogs (Dogs numbered 15 and 16) urine flow consistently decreased during the infusion of vasopressin at the lov; rate. A possible reason for this reduction w i l l be introduced in Section 4. Thus a decrease i n the rate of vasopressin infusion from 0.4 m-u./kg./ min. to 0.04 m-u./kg./min. led to an increase i n urine flow. 3. Effect on Urine Osmolality. Decreasing the rate of vasopressin infusion led to a decrease i n urine osmolality (Table VIII). The f i r s t noticable decrease occurred during the second ten minute period after the change to the low rate. Urine osmolality proceeded to decrease usually reaching i t s lowest value during the f i r s t ten minute period after resumption of the high rate (Fig. 6). A return to the high rate of infusion usually caused a rapid return i n urine osmolality to previous control levels. A decrease i n urine osmolality during the experimental period was recorded when the osmolality during the control period was either greatly hypertonic (Fig. 8) or significantly hypotonic (Fig. 7). Thus a decrease i n the rate of vasopressin infusion from 0.4 m-u./kg./ min. to 0.04 m-u./kg./min. led to a decrease in urine osmolality. 4. Effect on Solute Excretion. A decrease i n the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. led to a decrease i n osmolal clearance (Table VIII). The osmolal clearance usually began to decrease i n the second ten minute period after the change to the low rate and continued to decrease usually reaching i t s lowest value i n the last ten minute period during the low rate of infusion (Fig. 6). 49. Vasopressin Infusion: mU/kg./mln. 10min. Intervals Fig. 8. Effects of changing the rate of vasopressin infusion for a 30 min. period i n dog number 20. Conventions as in Fig. 6. 50. As mentioned in section 2 there were two dogs (dogs numbered 15 and 16) in which six tests produced a decrease in urine flow during the experimental period. In dogs 15 and 16 the osmolal clearance was decreased by 0.67 ml./ min. during the experimental period compared to the average for the series of 0.19 ml./min. A reduction in the rate of vasopressin infusion from the high rate to the low rate also led to a decreased excretion of sodium and potassium (Table VIII). The time course of the changes in sodium and potassium paralleled the time course of the change in osmolal clearance. Sodium and potassium excretion both decreased approximately 0.020 m.mole./min. However, there was an increase i n the Na./K. ratio from 2.24 during the control period to 2.60 during the experimental period (Table VIII). The increase i n the Na./K. ratio was a result of the approximate 0.020 m.mdle./ min. decrease i n cation excretion which reduced sodium excretion 10$ and potassium excretion 18$. It appeared that in proportion to the decrease i n osmolal clearance the decrease i n sodium excretion was greater. As shown i n Table VIII the percentage of sodium comprising the tot a l osmolal excretion was significantly decreased i n the experimental period. Thus a reduction i n the rate of vasopressin infusion from 0.4 m-u./kg./ min. to 0.04 m-u./kg./min. led to a decrease i n osmolal clearance. The magnitude of the decrease in osmolal clearance was less than the magnitude of the decrease in sodium excretion. The decrease i n sodium and potassium excretion was quantitatively similar and consequently there was an increase in the Na./K. ratio. 5. Effect on Free Water Clearance. Reducing the rate of vasopressin infusion led to an increase in free water clearance (Table VIII). Free water clearance usually began increasing noticably in the second ten minute period following the change to the low rate of infusion and usually reached a peak in the f i r s t ten minute period after beginning the infusion at .the high rate. Upon return to the high rate of vasopressin infusion free water clearance decreased to previous control levels (Fig. 6 ) . The increase in free water clearance occurred when free water clearance during the control period was either positive (Fig. 7) or greatly negative (Fig. 8 ) . Thus changing the rate of vasopressin infusion from 0 . 4 m-u./kg./min. to 0 . 0 4 m-u./kg./min. led to an increase i n free water clearance. 6. Effect on Glomerular F i l t r a t i o n Rate. Decreasing the rate of vasopressin infusion had no effect on glomerular f i l t r a t i o n rate as measured by the method described. The mean glomerular f i l t r a t i o n rate during the control period was 100 ml./min. which did not change during the experimental period (Table VIII). 52. TABLE VII. g hr. Infusion Experiments. 2 hr. Infusion Experiments. Control. Experimental. Control. Experimental. HEART X 120 132 76 80 + SEM 46.2 10.3 13.9 15.3 RATE d 12 4 n 21 5 BEATS/ t 5.4 1.7 MIN. 2P <0.001 <0.20 MEAN X 150 145 140 133 BLOOD - SEM 4.6 4.5 5.1 7 3.7 PRESSURE d 5 mm.Hg'e. n 21 5 t 4.2 1.8 2P <0.001 <0.20 Table VII. Effects of changing the rate of vasopressin infusion upon heart rate and blood pressure TABLE VIII J Hr, Infusion Experiments. 2 Hr. Infusion Experiments. URINE X FLOW ± SEM ml./min. d n t 2P Control. Experimental. 1.26 1.48 0.12 0.20 0.22 21 1.95 <0.10 Control. Experimental. 0.70 1.25 0.27 0.55 0.55 5 1.81 <0.20 X ±: SEM URINE 3 OSMOLALITY n m.osm./kg. ^ 2P 780 712 84 101 68 21 2.25 <0.05 736 482 89 114 254 5 4.97 <0.01 X ± SEM OSMOLAL 3 CLEARANCE n ml./min. t 2P 2.73 2.54 0.23 0.22 0.19 21 2.07 <0.10 1.46 1.18 0.39 0.25 0.28 5 1.05 <0.4 X FREE WATER d CLEARANCE n ml./min. t 2P -1.46 -1.05 0.20 0.24 0.41 21 5.05 <0.001 -0.73 -K).07 0.13 0.32 0.80 5 1.89 <0.20 1 ±SEM SODIUM d EXCRETION n m.mole/ t min. 2P 0.19 0.17 0.00 0.00 0.02 21 3.17 <0.005 0.12 0.09 0.03 0.00 0.03 5 2.23 <0.10 FOTASSIUM X + SEM EXCRETION g m.mole/min. n t 2P 0.09 0.07 0.01 0.01 0.02 21 5.89 <0.001 0.05 0.03 0.01 0.00 0.02 5 1.77 £0.20 54. g Hr. Infusion Experiments. . 2 Hr. Infusion Experiments. GLOMERULAR X FIL- + SEM TRATION ~ d RATE n ml./min. t 2P Control. Experimental. 100 100 10 10 0 21 0.3 NS Control. Experimental. 80 74 26 20 6 5 0.9 NS X +SEM Na./K. " 3 n t 2P 2.24 2.60 0.25 0.36 0.36 21 2.696 <0.02 2.45 3.15 0.37 0.73 0.69 5 1.342 N.S. PERCENTAGE X OF +SEM SODIUM IN 3 TOTAL n OSMOLAL t EXCRETION 2P 23.6 22.2 1.74 1.83 1.43 21 2.615 <0.02 28.0 26.4 1.99 1.65 1.62 5 1.407 N.S. Table VIII. Effects of changing the rate of vasopressin infusion upon urinary excretion. 55. Vasopressin Infusion : m i l / kg / min. 10 min. intervals Fig. 9 . Effects of changing the rate of vasopressin infusion for a 2 hr. period. Each horizontal line i n a 10 min. interval represents the average value from five tests i n five dogs. Variables are plotted as a percentage of the control values from each experiment. Thus the average control values equal 1 0 0 $ and are.equal to the values given i n Table VIII. From above downwards free water clearance with 1 0 0 $ equal to - 0 . 7 3 ml./min. (from Table VIII) and 0 $ equal to zero free water clearance, osmolal clearance, urine osmolality, urine flow. Dotted lines indicate a change in the rate of vasopressin infusion. C. EFFECTS OF CHANGING THE RATE OF  VASOPRESSIN INFUSION FROM 0.4  m-u./kg./min. to 0.04 m-u./kg./  min. FOR A PERIOD OF 2 HOURS. 1. Cardiovascular Effects. Changing the rate of vasopressin infusion for a two hour period led to qualitatively similar effects to those observed with the 30 min. change. Heart rate increased and a r t e r i a l pressure decreased i n the f i r s t ten minute period following the change to the low rate. Furthermore, these changes persisted u n t i l the high rate of infusion was again introduced at which time the heart rate and a r t e r i a l pressure returned to previous control levels (Table VII). Central venous pressure was measured i n four dogs. The central venous pressure during the control period was 11.25 cm. H2O (S.E.M. ±0.9) and during the experimental period no change was recorded. Thus changing the rate of vasopressin infusion for a 2 hr. period from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. led to a reduction i n a r t e r i a l pressure and an increase i n heart rate for the duration of the infusion at the low rate. 2. Effect on Urine Flow. Decreasing the rate of vasopressin infusion for a 2 hr. period led to a transient increase i n urine flow (Table VIII) (Fig. 9 ) . Following the change to the low rate urine flow began increasing i n the f i r s t 5 - 1 5 minutes and continued to increase u n t i l the seventh ten minute period. Urine flow then decreased from the ninth to the twelfth ten minute period at which time the high rate of vasopressin was again infused. The infusion at the high rate sharply reduced the urine flow which continued at this decreased level for the next two hours. It was necessary to plot the variables i n Fig. 9 as a percentage of control values i n the same animal because of the comparatively low urine flows obtained i n two experiments i n 57. which urine was collected from one kidney only. Thus decreasing the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. led to a transient increase in urine flow. 3. Effect on Urine Osmolality. Decreasing the rate of vasopressin infusion led to a transient decrease i n urine osmolality (Table VIII) (Fig. 9). Urine osmolality usually began decreasing in the second ten minute period following the change to the low rate of infusion. It continued to decrease u n t i l the seventh ten minute period following which i t began to increase. With resumption of the high rate of infusion urine osmolality returned to previous control levels. Thus decreasing the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. led to a transient decrease in urine osmolality. 4. Effect on Solute Excretion. The effect of the two hour infusion experiments on solute excretion was qualitatively similar to the effect produced by the half hour infusion experiments (Table VIII). Following an i n i t i a l osmotic diuresis to be described i n section Vi'osmolal clearance usually continued to decrease u n t i l resumption of the high rate of infusion at which time osmolal clearance began to increase (Fig. 9). During the experimental period sodium and potassium excretion both decreased approximately 0.025 m.mole/min. Also similar to the half hour infusion experiments there was a greater decrease i n sodium excretion than in osmolal clearance during the experi-mental period (Table VIII). Thus a decrease i n the rate of vasopressin infusion for a two hour period from a rate of 0;4 m-u./kg./min. to 0.04 m-u./kg./min. led to a decrease in solute excretion which was similar to that reported for the half hour infusion experiments. 58. 5. Effect on Free Water Clearance. A decrease i n the rate of vasopressin infusion for a two hour period led to a transient increase i n free water clearance (Table VIII) (Fig. 9). Following the i n i t i a l osmotic diuresis free water clearance remained at approximately the lowered level obtained during that diuresis. Decreasing the rate of vasopressin infusion init i a t e d i n the second ten minute period an increase in free water clearance which continued to increase reaching a peak approximately seventy minutes after beginning the infusion at the low rate. After reaching the peak increase free water clearance declined over the following f i f t y minutes of the low rate of infusion and continued to decline after the high rate of infusion was begun. This decline con-tinued u n t i l free water clearance reached a plateau which approximated previous control levels. Thus a decrease i n the rate of vasopressin infusion from 0.4 m-u./kg./ min. to 0.04 m-u./kg./min. led to a transient increase i n free water clearance. 6. Effect on Glomerular F i l t r a t i o n Rate. Decreasing the rate of vasopressin infusion had no significant effect on glomerular f i l t r a t i o n rate (Table VIII). D. URINE FLOW AND URINE COMPOSITION IN ANIMALS UNDER CHLORALOSE ANESTHESIA. The significance of the urinary responses to balloon in f l a t i o n i n the l e f t atrium or to changing the rate of vasopressin infusion can only be assessed after an examination has been made of other variations in urine flow and composition which occur i n animals which have been anesthetized with chloralose and subjected to a traumatic surgical operation. Whilst animals were not sp e c i f i c a l l y prepared for this purpose the necessity of waiting one hour after surgery before beginning the experiment plus 59. intervals between experimental periods lasting from 90 minutes to 120 minutes meant that there was ample opportunity during the experiments already described to observe changes i n urine flow and composition occuring outside the test periods. In the a t r i a l distension experiments although the ureters were catheterized at the beginning of the operation i t was not practicable to collect and measure the urine u n t i l a l l the surgical procedures were completed. However, during the operation the urine did flow into test tubes and there was usually less than 10 ml. of urine collected from each kidney by the time the operation was finished; that i s about 60 minutes collection. In a l l experiments in series two and two experiments i n series one, urine composition was not recorded u n t i l one hour after surgery. However, i n seven experiments i n series one urine flow and composition was recorded twenty minutes after surgery. In five of these experiments a small osmotic diuresis occured i n the f i r s t hour following surgery (dogs numbered 3, 4, 5, 6, 9) while i n one experiment (dog No. 8) a much longer osmotic diuresis occured. The experiment with dog No. 8 is the only experiment i n the a t r i a l distension series i n which the osmotic diuresis directly affected the experimental results. This osmotic diuresis following surgery i s more apparent in the vasopressin infusion experiments where urine flow was recorded 10 minutes after completion of ureter catheterization and no thoracotomy was performed. In the half hour infusion experiments an osmotic diuresis occurred in the f i r s t hour after surgery i n four of the eight dogs (dogs numbered 13, 15, 16, 20). One of these experiments i s shown i n Fig. 8. This i s the only experiment i n which the osmotic diuresis affected the experimental period i n the half hour infusion experiments. During the osmotic diuresis in Fig. 8 the free water clearance remained relatively constant in spite of 60. the increase in osmolal clearance. In the two hour infusion experiments the osmotic diuresis occured i n four of the five experiments (dogs numbered 22, 23, 2k, 2 6 ) . A composite picture of the osmotic diuresis can be seen in Fig. 9 . The osmotic diuresis in the 2 hr. experiments reached a peak approximately 40 minutes after completion of surgery. During this osmotic diuresis free water clearance decreased. After the osmotic diuresis reached a peak i t began to decline over the next 80 minutes. The two hour.infusion series i s the only series of experiments i n which the osmotic diuresis had any significant affect on the i n i t i a l control period. The apparent effect of the osmotic diuresis was to raise the mean control urine flow and osmolal clearance and to lower the i n i t i a l mean control osmolality and free water clearance over that which would be expected had the diuresis not occured. Except for the changes i n urine flow and composition mentioned above the urine flow during the control periods was usually decreasing or remain-ing quite constant. In the 20 to 30 minute interval between control periods i n the a t r i a l distension and half hour infusion experiments urine flow was also very constant with three notable exceptions occuring during the a t r i a l distension experiments. The exceptions occured when a change i n the experimental infusion was made from any dose of vasopressin to saline or from the vasopressin infusion at a rate of 1.0 m-u./kg./min. to the vaso-pressin infusion, at a rate of 0.1 m-u./kg./min. or from vasopressin infusion at a rate of 0.4 m-u./kg./min. to the vasopressin infusion at a rate of 0.025 m-u./kg./min. In ten dogs i n which a change from vasopressin to saline was made four of these changes resulted i n a diuresis (Fig. 3). In one out of four instances i n which a change from a rate of vasopressin infusion of 1.0 m-u./kg./min. to 0.1 m-u./kg./min. was made a small diuresis resulted.- In one out of five instances in which a change from a 61. PLASMA. Sodium Concentration(mEq./L Potassium (mEq./L) Concentration. Osmolality (m.osm./kg.) Atr i a l Distension Series 1. Saline. 0.1 m-u./ kg./min. 1.0 m-u./ kg./min. Series 2. Saline. X S.E.M X S.E.M X S.E.MJ X S.E.M 0.025 m-u./ kg./min. X S.E.M 0.4 m-u./ X kg./min. S.E.M h hr. infusion experiments. X • S.E.M 2 hr. infusion experiments. X S.E.M 143 4.01 140 2.99 137 3.69 140 1.37 139 2.34 139 2.25 140 1.41 140 0.58 139 3.50 140 2.97 139 (3.30 141 1.93 140 2.42 140 2.09 L40 L.48 L38 L.72 C2 141 3.70 139 2.57 138 2.06 142 1.94 139 1.98 140 2.29 139 1.37 137 0.79 °1 4 0.16 4 0.20 4 0.15 4 0.17 4 0.11 4 0.13 4 0.12 3 0.19 4 0.23 4 0.11 4 0.00 4 0.17 4 0.15 4 0.10 4 0.11 3 0.19 4 0.22 4 0.11 4 0.00 4 0.17 4 0.15 4 0.13 4 0.09 3 0.19 Cl 282 4.09 280 5.22 282 4.94 297 1.85 294 2.52 295 1.97 291 1.72 296 1.39 278 4.11 279 4.91 278 4.82 298 2.09 294 2.90 297 2.58 292 1.87 299 1.88 Table IX. Average plasma values (X) for sodium concentration (mEq./L.), potassium concentration (mEq./L.) and osmolality (m.osm./kg.) in the f i r s t control period (C-^), the experimental period (E) and the second control period (C2) during the experimental procedures shown on the l e f t . S.E.M. i s standard error of the mean. NO. Kgs. Wt. in 0.6$ NaCl. Sol. mis. Dextran Total TABLE X. No. of 4 ml Bl.Samples taken and replaced, with Dextra . Vol. of urine excre-.ted. Balance Infused -Excreted. Bal-ance, ml./ kg. Length of Experiment < from I i n i t i a l ) anes. to end of last control (min period. iverage rat< >f accum-ilation ml./ :g./min. Initial CEhlor-alose. Exp. Sal. or CHL. Vaso-press in. Inulin Sept. 19. 10 18 180 840 410 430 150 1860 20 ^73 1187 66 540 0.12 Sept. 26. 11 25 300 760 330 360 150 1900 16 355 1045 42 490 0.09 Oct.3 12 24 240 640 280 320 150 1630 14 347 1283 53 420 0.13 Oct. 10. 13 26 310 860 400 430 200 2200 19 511 1689 65 580 0.11 Oct. 16. 14 24 240 1020 400 400 150 2210 19 280 1930 80 510 0.16 Oct. 29. 15 25 250 700 280 D 150 1380 12 L60 1220 49 480 0.10 Oct. 31. 16 18 200 820 400 3 150 1570 20 S88 882 49 500 0.10 Nov. 4-17 22 220 820 410 3 150 1600 18 329 971 44 520 0.08 Nov. 7. 18 18 180 900 370 D 150 1600 18 L97 1403 78 490 0.16 No. Kgs. Wt. in OA I n i t -i a l Chlor-alose >% NaC Exp. Sal or CHL 1. Sol. Vaso-press-in Inulin mis. Dextran Total TABLE X. No. of 4 ml. BI. Samples taken and replaced with Ifext.ran. Vol. of urine excre-ted. 3alance infused -Cxcreted Bal-ance, ml./ kg. Length of Experiment from i n i t i a l anes. to end of last control (min) period Average rate of accum-ulation ml./kg./ min. Nov. 14. 19 18 200 780 370 410 200 I960 19 274 1686 94 540 0.17 Nov. 21. 20 21 240 660 310 3 3 0 150 1690 16 672 1018 48 430 0.11 Nov. 28. 21 23 280 840 370 400 150 2040 18 161 1879 82 440 0.19 Dec. —22 26 260 720 360 3 9 0 150 1880 17 551 1329 51 560 0.09 Table X. Assessment of the volume load to individual dogs used in the experiments. For description see text. 64. rate of vasopressin infusion of 0.4 m-u./kg./min. to 0.025 m-u./kg./min. was made a small diuresis resulted. Thus in these animals anesthetized with chloralose and surgically traumatized, increases in urine flow and changes in urine composition may be expected either within one hour following surgery or following a reduction in the rate of vasopressin infusion. E. THE EFFECTS OF THE HYDRATION  PROCEDURES ON THE EXPERIMENTAL  ANIMAL. The significance of the urinary responses to balloon inflation in the l e f t atrium or to changing the rate of vasopressin infusion can only be assessed after on examination has been made of the volume load to the animal and the changes in plasma sodium, potassium and osmolality. Table X gives an indication of the volume assessment carried out on each animal used in this investigation. The table indicates the volume of 0.6% saline solution infused and the form in which i t was infused - anesthetic, vaso-pressin or inulin. The infusion of 0.6% NaCl and the dextran given during surgery comprise the gross volume load to the animal. Subtracted from the gross volume load i s the volume of urine excreted during the experiment which gives the net volume load. Because of the variety in dog weights and time to complete the experiment the net volume load is expressed as "the average rate of volume accumulation" i n ml./kg./min. The average volume load to a l l the animals was 59 ml./kg. (SEM - 2.64) and the average rate of volume accumulation was 0.12 ml./kg./min. (SEM -0.005). This degree of hydration however, had no significant effect on plasma sodium, potassium and osmolality as measured by the methods described (Table IX). During the control and experimental periods the average plasma osmolality for a l l 33 dogs was 289 - 1.0 (SEM) m.osmole./kg. Furthermore, there were no individual experiments in which any large 65. changes occured in plasma sodium, potassium or osmolality. There was no correllation between the absolute level or the magnitude of a change of any of the measured renal variables and the rate of volume accumulation or the net volume load. Thus hydration procedures which produced a net volume load of 59 ml./kg. and an average rate of volume accumulation of 0.12 ml./kg./min. had no significant effect on plasma sodium, potassium or osmolality. The hydration procedures also had no apparent effect on the measured renal variables. P A R T I V . D I S C U S S I 0 N. 66. A. INTRODUCTION. The effects of l e f t a t r i a l distension during the saline infusion reported in the present experiments are similar to those reported by other authors (Henry et a l . , 1956; Ledsome et a l . , 1961; Arndt et a l . , 1963; Shu'ayb et a l . , 1965; Johnson et a l . , 1969). Distension of the l e f t atrium resulted in an increase in heart rate, a drop in a r t e r i a l pressure, a dilute diuresis with an increase in free water clearance and an increase i n osmolal clearance. The experimental results of Lydtin and Hamilton (1964) and Ledsome et a l . (1961) have been confirmed and extended; a diuresis to l e f t a t r i a l distension can be produced during an infusion of vasopressin at rates more than adequate to completely inhibit water diuresis i n a con-scious dog. The effects of changes in the rate of infusion of vasopressin within the context of the experimental design used in the a t r i a l distension experiments has also been investigated. In discussing the infusion experiments the assumption has been made that the experimental animals were releasing relatively constant amounts of antidiuretic hormone throughout the control and experimental periods. Changes in the measured variables would then be due to changes in the rate of vasopressin infusion. A change in the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. resulted i n an increase in heart rate, a decrease in a r t e r i a l pressure and a transient dilute diuresis. B. CARDIOVASCULAR EFFECTS OF VASOPRESSIN  INFUSION AND LEFT ATRIAL DISTENSION. 1. A t r i a l Distension Experiments. The vasopressin infusions had no effect on the cardiovascular variables measured in the a t r i a l distension experiments. While i t appeared that as the rate of vasopressin infusion increased the heart rate during the control and experimental periods decreased (Table II) this effect was not significant. In response to an approximate 20 cm. H^ O increase in l e f t a t r i a l pressure heart rate increased and a r t e r i a l pressure decreased. The increase i n heart rate and decrease in a r t e r i a l pressure was quantitatively similar to that described previously by Ledsome et a l . (1961) whose experimental design was similar to that reported i n the present investigation. The increase in heart rate occurred immediately after distension of the l e f t atrium. A reflex increase in heart rate i n response to stimulation of l e f t a t r i a l receptors has been reported by Ledsome and Linden (1963, 1967). The decrease in a r t e r i a l pressure l i k e l y resulted from a decreased cardiac output due to the pooling of blood in the intrathoracic circulation which must have occurred to raise l e f t a t r i a l pressure 20 cm. H2O. Henry et a l . (1956) have indicated that the 20 cm. increase i n l e f t a t r i a l pressure which occurred when the mitral o r i f i c e was blocked was not transmitted to the right atrium and measurements of cardiac output during this degree of l e f t a t r i a l dis-tension indicated that cardiac output was reduced approximately 30% (Henry, 1955). 2. Infusion Experiments. Reduction in the rate of vasopressin infusion for either 30 min. or 2 hrs. was associated with an increase in heart rate and a decrease in a r t e r i a l pressure. Changes in heart rate and a r t e r i a l pressure occurred with the change i n the rate of vasopressin infusion and the values obtained during the experimental period were observed for the duration of the infusion of vasopressin at the low rate indicating that these changes i n heart rate and a r t e r i a l pressure v/ere probably mediated by changes in the circulating vasopressin concentration. The time course of the change i n heart rate during the 30 min. infusion experiments (Fig. 4) which occurred over a period of many minutes was in sharp contrast to the immediate increase in heart rate observed in the a t r i a l distension experiments. Significant cardiovascular changes in response to vaso-pressin infusion are usually considered to occur within a dose range higher than that in which urinary effects are observed. Rocha e Silva and Rosenberg (1969) infused vasopressin into anesthetized dogs at rates between 0.8 m-u./kg./min. and 6.0 m-u./kg./min. and reported only a slight i n i t i a l rise i n blood pressure and a decrease in heart rate; these effects lasted for only 2 - 4 mins. C. URINARY EFFECTS OF ATRIAL  DISTENSION EXPERIMENTS. 1. Assessment of the Maximum Effective Dose of Vasopressin in the A t r i a l  Distension Experiments. The results of the a t r i a l distension experiments indicated that a significant linear relationship was established between free water clearance and osmolal clearance during infusions of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. No significant linear relation-ship was established between free water clearance and osmolal clearance during infusion of saline and vasopressin at rates of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. The accepted view of the mechanisms of urine concentration and dilution as outlined by Dicker (1970) and Berliner and Bennett (1967) argued that the action of vasopressin was to promote the passive re-absorption of water along the concentration gradient existing between the collecting duct and d i s t a l tubule and the medullary interstitium. Further-more, studies i n man (Hollander, Williams, Fordham, and Welt, 1957), dog (Orloff, Wagner and Davidson, 1957), and rat (Gauer and Tata, 1966; Tata and Gauer, 1966) indicated that the amount of water reabsorbed i s p a r t i a l l y dependent on the concentration of vasopressin i n the blood and the volume 69. of f l u i d passing through the collecting duct per unit of time. These studies also indicated that at a "maximum" effective vasopressin concentration the amount of water reabsorbed was limited. In the dog Orloff et a l . (1957) found the majcimum reabsorption of water to be approximately 3 ml./min. It i s logical therefore to assume that i n the dog in the presence of maximum antidiuretic a c t i v i t y an increase in flow, represented by an increase i n osmolal clearance, w i l l result in an increase i n the reabsorption of water u n t i l the quantity of water reabsorbed reaches approximately 3 ml./min. Thus water reabsorption i s dependent on osmolal clearance i n the presence of maximum antidiuretic a c t i v i t y . Changes i n the level of antidiuretic a c t i v i t y when less than maximum may result i n changes i n free water f clearance which occur independently of changes i n osmolal clearance. Thus i n Fig. 4 infusion of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. establish identical significant linear relationships between free water clearance and osmolal clearance identifying these infusions as possessing maximum antidiuretic a c t i v i t y . Intuitively i t would be expected that as osmolal clearance approached zero, free water clearance would also approach zero. Fig. 4 indicates that within the 95$ confidence limits of the regression lin e this i s indeed the case. The failure of the infusions of saline and vasopressin i n doses of 0.025 m-u./kg./min. and 0.1 m-u./kg./ min. to establish a significant relationship between free water clearance and osmolal clearance identifies these infusions as possessing less than maximum antidiuretic a c t i v i t y under the present experimental conditions. 2. Changes i n Urinary Composition During the A t r i a l Distension Experiments. The magnitude and time course of the diuretic response to l e f t a t r i a l distension during infusion of saline was comparable with that reported previously by Henry et a l . (1956), Ledsome et a l . (1961) and Lydtin and Hamilton (1964). The infusions of vasopressin while having no significant 70. effect on the urine flow during the control periods significantly reduced the urine flow recorded i n the experimental period during the saline infusion. However, the infusions of vasopressin did not abolish the diuretic response to l e f t a t r i a l distension since a small but significant diuretic response also occurred during the infusion of vasopressin at the maximum anti-diuretic dose of 1.0 m-u./kg./min. Measurement of urine and plasma osmolality had not been previously made on the diuresis to l e f t a t r i a l distension over the range of vaso-pressin infusions investigated i n the present experiments. Evidence has already been presented indicating that vasopressin infusion at a rate of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. achieved maximum antidiuretic a c t i v i t y . This fact also manifests i t s e l f as no significant changes in urine osmolality during l e f t a t r i a l distension over the range of osmolal clearance observed for these two infusions. Furthermore control osmolality did not increase significantly between the infusions of vasopressin at rates of 0.1 m-u./kg./min, 0.4 m-u./kg./min. and 1.0 m-u./kg./min. which may indicate that during the control period of the vasopressin infusion at a rate of 0.1 m-u./kg./min. maximum antidiuretic a c t i v i t y may also have been in effect. During infusion of either saline or vasopressin at a rate less than 0.4 m-u./kg./min. significant decreases i n urine osmolality were observed in response to l e f t a t r i a l distension. The decrease i n urine osmolality was probably not due to increased flow through the tubules since no evidence exists to indicate that such small increases i n flow (see data on osmolal clearance, Table III) as encountered i n the present investigation have a significant effect i n diluting the urine. Rather, the most probable ex-planation i s a change in the concentrating and diluting process of the kidney due to a decreased circulating level of antidiuretic hormone. This explanation receives support from the time course and latency of onset of the urine dilution during the infusions of saline and vasopressin at rates of 0.025 m-u./kg./min. and 0 .1 m-u./kg./min. The urine dilution had a similar time course and latency of onset to that following inhibition of antidiuretic hormone i n conscious dogs (Verney, 1947) and to the time course and latency of onset of the response reported i n the 30 min. infusion experiments. Furthermore, the urine dilution was abolished by the higher rates of vasopressin-infusion. The fact that during the infusions of vasopressin urine osmolality increased significantly i n both the control and experimental periods may lead to the interpretation that the effectiveness of the vasopressin infusions i n increasing the urine osmolality recorded i n the experimental period was not directly dependant on the rate of vasopressin infusion but on the increased urine osmolality i n the control period which also occurred. However, in Table VI i n which the results are grouped according to the urine osmolality recorded i n the control period a urine concentration during the control period as high as 1311 m.osm./kg. did not prevent a significant decrease i n the urine osmolality i n the experimental period from occurring. The variables of osmolal clearance and free water clearance had previously appeared only i n the papers by Arndt et a l . (1963) and Johnson et a l . (1969). Arndt et al j s (1963) investigation indicated that the increase i n osmolal clearance occurred over a wide range of U/P ratios. • The present results extend this view to include an increase i n osmolal clearance which i s independent of vasopressin doses over the range of 0.025 m-u./kg./min. to 1.0 m-u./kg./min. Such a view i s ju s t i f i e d in spite of the significantly greater osmolal clearance observed in the control and experimental period during the vasopressin infusion at a rate of 0 . 4 m-u./ kg./min. The significant difference at this dose appeared to be due to only-two experiments i n which a spontaneous increase i n osmolal clearance co-incided with the test periods. There i s , however, a suggestion from Table VI that an increase in osmolal clearance i s dependent on the urine osmolality recorded during the control period. This suggestion i s based mainly on the observation that osmolal clearance did not increase when urine osmolality was low during the control period. This observation, however, can be disregarded since the population sample was small and of the five dogs, one did respond to l e f t a t r i a l distension with an increased osmolal clearance while two others never responded with an increased osmolal clearance i n response to l e f t a t r i a l distension, even when the urine osmolality i n the control period was much higher. It i s significant to establish i n the present experiments whether the increase i n osmolal clearance in response to l e f t a t r i a l distension i s independent of the concentrating and diluting mechanism i n the kidney. In their report of the effects of l e f t a t r i a l distension Shu'ayb et a l . (1965) argued that the increased excretion of solute they observed might be a result of dead space i n the urinary collecting system. The dead space would not allow changes i n solute concentration to reach the test tube u n t i l sometime after transmitted volume changes with the result that a paradoxical increase i n solute excretion would be recorded. Several observations however, from the present experiments indicate that the increase in osmolal clearance i s independent of the concentrating and diluting mechanisms i n the -kidney. F i r s t l y , during infusion of vasopressin at rates of 0.4 m-u./kg./min. and 1.0 m-u./kg./min. i n which significant increases in osmolal clearance occurred i n response to l e f t a t r i a l distension no change i n the concentrating and diluting mechanism occurred during the experimental period. Furthermore, indirect evidence that the 73. increase in osmolal clearance during the infusions of saline and vasopressin in doses of 0.025 m-u./kg./min. and 0.1 m-u./kg./min. was not due to a change in the concentrating and diluting mechanism i s seen in the vaso-pressin infusion experiments. In the vasopressin infusion experiments despite the fact that urine osmolality decreased 68 m.osm./kg. during the 30 min. infusion experiments and 254 m.osm./kg. during the 2 hr. infusion experiments no increase i n osmolal clearance was observed. In fact a significant decrease in osmolal clearance was recorded during the 30 min. infusion experiments. Thus in establishing a cause and effect relationship for the increase i n osmolal clearance during l e f t a t r i a l distension i t appears that the effect of increased osmolal clearance may be caused by a mechanism independent of the concentrating and diluting mechanism. Although the increase i n osmolal clearance i n the present investigation was accompanied by increases i n sodium and potassium excretion the increases in cation excretion were significant i n only 4 of 11 tests (Table III). Furthermore, to identify the two significant increases in sodium excretion which occurred as a natriuretic response to l e f t a t r i a l distension may be misleading since this may be interpretated as indicating that two mechanisms are operating to increase solute excretion during l e f t a t r i a l distension, one increasing sodium excretion and the other increasing osmolal clearance. Such may not be the case since an increase i n glom-erular f i l t r a t i o n rate may result i n a significant increase in sodium excretion as well as a significant increase i n osmolal clearance. To avoid such d i f f i c u l t y in the present investigation the following criterion for a natriuresis has been established; a significant increase i n sodium excretion which is of the same magnitude as a significant increase in total solute excretion i s not a natriuretic effect, while a significant increase in sodium excretion of a significantly greater magnitude than the increase 7 4 . in total solute excretion i s a natriuretic effect. This criterion i s represented i n the results as the percentage of sodium comprising the total osmolal excretion. On this basis there was a natriuretic response to l e f t a t r i a l distension only during vasopressin infusion at a rate of 0.1 m-u./kg./min. If sodium or potassium excretion was increased by any specific effect more consistent results would be expected. Thus the protean nature of any natriuretic or kaluretic effects make i t unlikely that the increase in osmolal clearance was either accomplished or significantly augmented by a mechanism specific for sodium or potassium. The vasopressin infusions i n series two of the a t r i a l distension experiments significantly increased the percentage of sodium comprising the total osmolal excretion i n the control and experimental periods. This increase, however, i s of doubtful significance since the same effect was absent from series one. Atherton, Hai and Thomas (1969) reported a similar effect of increasing vasopressin infusion on sodium excretion i n the hydrated rat. Their results were also variable and they also questioned the significance of a continuous vasopressin infusion on sodium excretion. Due to the known effect of vasopressin on water absorption changes in free water clearance would be expected to provide the most sensitive index of any of the variables measured on the effects of changes i n the circulating concentration of vasopressin. Free water clearance increased in response to l e f t a t r i a l distension only during infusion of vasopressin at rates of 0.1 m-u./kg./min. or less. Thus infusion of vasopressin at rates of 0.U m-u./kg. /min. prevented any increase in free water clearance in response to l e f t a t r i a l distension. However, infusion of vasopressin at rates of 0.02$ m-u./kg./min. and 0.1 m-u./kg./min. prevented significant increases in free water clear-ance from occurring. Thus infusion of vasopressin at a rate of 0.02$ m-u./kg. /min. was effective i n reducing the increase i n free water clearance i n response to l e f t a t r i a l distension. 75. This dose of vasopressin appeared more effective i n preventing the increase in free water clearance i n some dogs than i n others. One dog i n which an infusion of vasopressin at a rate of 0.025 m-u./kg./min. was capable of producing a dramatic antidiuresis i s shown i n Fig. 5. Indirect evidence that increases i n free water clearance i n response to l e f t a t r i a l distension were prevented by the vasopressin infusions and not the control concentration of the urine i s given byTable VI. In Table VI highly significant increases in free water clearance occurred over the control osmolality range of 0 - 750 m.osm./kg. The infusions of vasopressin significantly reduced free water clearance in the control and experimental periods u n t i l at maximum antidiuretic a c t i v i t y free water clearance varied inversely to osmolal clearance as has already been mentioned. The time course of the increases in free water clearance that occurred i n response to l e f t a t r i a l distension were compatible with removal and replacement of antidiuretic hormone i n the blood. D. URINARY EFFECTS OF VASOPRESSIN  INFUSION EXPERIMENTS. 1. 30 min. Infusion Experiments. Reduction i n the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. for a 30 min. period was associated with a small diuresis. There was an increase i n free water clearance, a decrease i n solute excretion and the urine became more dilute. The decreased concentration of solutes without any large change in the rate of excretion of solutes and the time course of the changes were similar to the changes which occur during water diuresis. The small decrease in solute excretion observed i n the present experiments could be accounted for by a decrease in glomerular f i l t r a t i o n rate secondary to the decrease i n a r t e r i a l pressure. However, part of the decrease in sodium excretion which occurred may be directly attributable to'tffe 76. decrease in a r t e r i a l pressure i f this decrease i n pressure were transferred to the peritubular capillaries (Earley and Daugharty,1969). The fact that no changes in glomerular f i l t r a t i o n rate were measured may not be s i g n i f i -cant since the accuracy of the method of measurement of inulin clearance i s not adequate to detect small changes in glomerular f i l t r a t i o n rate, capable of altering solute excretion. It was mentioned i n the results that there were two dogs (dogs numbered 15 and 16) i n which six tests produced an antidiuresis i n the experimental period. Upon closer examination of the results from those two dogs i t was found that while osmolal clearance was decreased an average of 0.19 ml./min. and free water clearance was increased an average of 0.39 ml./min. i n the experimental period for a l l 21 tests i n 8 dogs these two dogs showed a decrease i n osmolal clearance of 0.67 ml./min. and an increase i n free water clearance of 0.41 ml./min. in the experimental period. Thus while these dogs responded to a decrease i n the rate of infusion of vasopressin with decreased • reabsorption of water a diuresis was not manifest due to the relatively large simultaneous decrease i n solute excretion. That changes i n the rate of vasopressin infusion w i l l increase urine flow i s also apparent from the results of the a t r i a l distension experiments presented i n Section D of the results. 2. 2 hr. Infusion Experiments. In the 2 hr. infusion experiments an osmotic diuresis occurred before the f i r s t control period. The appearance of this osmotic diuresis was not confined to this set of experiments since there were indications that i t occurred i n the other sets of experiments in this investigation (see Section D of the results). This osmotic diuresis appears to be related to com-pletion of the surgical procedures. In the present experiments a reduction in the rate of vasopressin infusion from 0.4 m-u./kg./min. to 0.04 m-u./kg./min. for 2 hours led to changes in urinary excretion which were qualitatively similar to those reported for the half hour infusion experiments. However, in the 2 hour infusion experiments there was a quantitatively greater dilution of the urine and after 70 minutes the urine flow and free water clearance began to decrease and osmolality to increase. The characteristics of the diuresis associated with the 2 hour infusions were compatible with the known actions of vasopressin. However, the time course of the diuresis was not as expected following removal and replacement of vasopressin i n the blood. If the h a l f - l i f e of vasopressin i n the blood i s 5 min. (Lauson and Bocanegra, 196l) then a reduced stable concentration i n the blood would be expected about 30 min. after the change to the lower rate of infusion. Infusions of vasopressin at rates as low as 0.01 m-u./kg./min. into conscious dogs during water diuresis have been shown to have maximum effect i n about 30 min. (Verney, 1947) and i n these experiments when vasopressin infusion was stopped urine flow returned to the diuretic level in about the same time. In the present experiments the urine flow and concentration continued to change over the whole 2 hour period as shown in Fig. 9. It must be con-cluded that i n the moderately hydrated anesthetized dog when the con-centration of vasopressin i n the blood i s reduced from a high level to a lower, but i n the conscious dog antidiuretic l e v e l , a transient, dilute, diuresis may be expected. E. EVIDENCE AGAINST ANTIDIURETIC HORMONE AS AN AGENT  PRODUCING THE DIURESIS TO LEFT ATRIAL DISTENSION. Evidence against antidiuretic hormone acting as an agent to produce the diuretic response to l e f t a t r i a l distension comes from the experiments by Ledsome et a l . (1961) and Lydtin and Hamilton (1964). Their demonstration of a dilute diuresis i n response to l e f t a t r i a l distension during an infusion of vasopressin at a rate of 0.025 m-u./kg./min., which had previously been demonstrated to completely inhibit water diuresis in a conscious dog, led to the argument that the diuretic response to l e f t a t r i a l distension was unlikely to be due to a decreased release of antidiuretic hormone from the neurohypophysis. This argument i s dependant on a relatively unchanged effectiveness of low doses of vasopressin to max-imally reduce urine flow with the experimental animal i n two different states; one state being conscious i n water diuresis; the other being anesthetized and overhydrated. The question arises as to whether the difference in these states could alter the effectiveness of vasopressin to concentrate urine. The effectiveness of an infusion of vasopressin i n reducing urine flow was investigated i n conscious dogs undergoing water diuresis by Shannon (1942) and Verney (1947). Their results agreed with Ledsome et a l . (1961) and indicated that vasopressin infused at a rate of 0.01 m-u./kg./min. had a maximum effect i n reducing urine flow. Unfortunately, assessment of the effectiveness of vasopressin i s d i f f i c u l t i n anesthetized dogs because water given intravenously or by stomach tube does not always produce a diuresis (Verney, 1929). However, Perlmutt (1961) obtained a dilute diuresis i n severely hydrated anesthetized dogs and found that a dose of vasopressin of approximately 0.07 m-u./kg./min. had a maximum effect i n reducing urine flow although even higher doses did not achieve maximum urine concentration. The differences i n the effectiveness of vasopressin demonstrated i n these studies have been attributed to the level of hydration (Perlmutt, 1962). Other investigators have also supported a similar relationship between hydration and alterations in the effective dose of vasopressin (Epstein, Kleeman and Hendrikx, 1957; Lavinsky, Davidson and Berliner, 1959). Orloff 79. et a l . (1957) have demonstrated that at less than the maximum vasopressin dose of 0.8 m-u./kg./min. used in their study the effectiveness of vaso-pressin i n concentrating urine was reduced by increased osmolal clearance. There i s also some evidence that the sensitivity to exogenous vasopressin i s increased when release of endogenous hormone i s reduced, for example after alcohol administration (Pickford, 1966). Shannon (1942) examined this possibility i n conscious dogs undergoing water diuresis but was unable to find evidence to support such a change i n sensitivity. However, variation in response to changing blood levels of antidiuretic activity was indicated by the experiments of Czaczkes, Kleeman, and Koenig (1964) who i n a study on man showed that an injection of vasopressin which increased plasma anti -diuretic a c t i v i t y to 5.5 uU/ml. was associated with a urine concentration of 641 m.osmole/l whereas when the same level of antidiuretic a c t i v i t y occurred during a decrease i n antidiuretic a c t i v i t y from a higher level the urine concentration was 224 m/osmole/l. Since the studies of Ledsome et a l . (I96l) and Lydtin and Hamilton (1964) used a moderate level of hydration and i n some instances anesthesia the possibility remained that an infusion of vasopressin at a rate of 0.025 m-u./kg./min. may not constitute a maximum effective dose. Results from the a t r i a l distension experiments indicated that i n the moderately hydrated anesthetized animal only an infusion of vasopressin at a rate of 0 .4 m-u./kg./min. or above constituted a maximum effective dose. The only significant effect on the diuresis to l e f t a t r i a l distension of an infusion of vasopressin at a rate of 0.025 m-u./kg./min. was to prevent a significant increase i n free water clearance from occurring. The 30 min. infusion experiments also indicated that when large changes i n vasopressin con-centration occur a diuresis may result even during vasopressin infusion at a rate of 0.04 m-u./kg./min. 80. Thus i t i s apparent from the results of the present investigation that in dogs anesthetized and hydrated in a similar fashion to those used by Ledsome et a l . (1961) and Lydtin and Hamilton (1964), a dose of vasopressin of 0.025 m-u./kg./min. v/ould not constitute a maximally effective dose. Furthermore i f relatively large changes in vasopressin concentration in the blood occurred during infusion of vasopressin at a rate of 0.025 m-u./kg./ min. a dilute diuresis may occur. The conclusion that the diuretic response to l e f t a t r i a l distension could not be due to a decrease in the rate of release of vasopressin from the neurohypophysis was therefore not j u s t i f i e d . Further evidence against antidiuretic hormone acting as an efferent agent has been the demonstration that the diuretic response i s transient despite continued distension of the l e f t atrium (Henry et a l . , 1956; Ledsome et a l . , 1961; Lydtin and Hamilton, 1964; Shu'ayb et a l . , 1965) and despite a continued reduction i n blood antidiuretic a c t i v i t y during the distension (Shu'ayb et a l . 1965? Figs. 2 and 3). Results from the 2 hr. infusion experiments indicated that when large changes i n vasopressin concentration occur a transient dilute diuresis may result despite a continued reduction i n the rate of infusion of vasopressin. Furthermore the transient dilute diuresis which occurred i n response to changing the rate of infusion of vasopressin reached a peak approximately 70 minutes after the change to the reduced rate of vasopressin infusion. The transient, dilute, diuretic response to l e f t a t r i a l distension has been reported to reach a peak 50 - 90 min. after beginning l e f t a t r i a l distension. Thus antidiuretic hormone could be acting as an efferent agent to produce the transient, dilute diuresis to l e f t a t r i a l distension i f the rate of release of antidiuretic hormone from the neurohypophysis were reduced from a rather high rate to a much lower rate. 8 1 . F. EVIDENCE FOR ANTIDIURETIC HORMONE AS AN  AGENT PRODUCING THE DIURESIS TO LEFT  ATRIAL DISTENSION. There i s good indirect evidence suggesting that one efferent mechanism responsible for the diuretic response to l e f t a t r i a l distension i s a change i n the circulating concentration of antidiuretic hormone. For example, a major component of the diuretic response to l e f t a t r i a l dis-tension in a moderately hydrated animal i s a water diuresis delayed by 5 to 10 minutes (Henry et a l . 1956; Ledsome et a l . , 1961; Arndt et a l . , 1963; Lydtin and Hamilton, 1964). This fact receives further support from the results of the a t r i a l distension experiments in the present investigations. The time course of the dilution of the urine following 30 min. of l e f t a t r i a l distension i s similar to the time course of the dilution of the urine following either cessation of vasopressin infusion i n hydrated dogs (Verney, 1947) or a 30 min. reduction i n the rate of vasopressin infusion i n hydrated anesthetized dogs (present experiments). In hydrated anesthetized dogs the time course of the dilution of the urine following prolonged l e f t a t r i a l distension i s similar to the time course of the dilution of the urine following a prolonged reduction i n the rate of infusion of vaso-pressin (present experiments). Furthermore, the study by Carswell et a l . (1970) demonstrated a diuretic response to l e f t a t r i a l distension i n an isolated kidney perfused at constant pressure indicating that a blood borne agent was involved. More direct supporting evidence for the suggestion that antidiuretic hormone i s an agent responsible for the diuresis to l e f t a t r i a l distension comes from studies i n which antidiuretic a c t i v i t y has been assayed during the diuretic response to l e f t a t r i a l distension (Baisset and Montastruc, 1959; Shu'ayb et a l . , 1965; Johnson et a l . , 1969). There are however several features i n the reports by Baisset and Montastruc (1959) and 82. Shu'ayb et a l . , (1965) which lessen the v a l i d i t y of their conclusions. Baisset and Montastruc (1959) claimed to have shown that distension of the l e f t atrium produces diuresis by inhibiting neurohypophysial release of antidiuretic hormone. They assayed an extract of jugular vein blood (Baisset, Douste-Blazy, Montastruc and Valdeguie, 1957) by injecting the extract into ethanol anesthetized rats and compared the effect on urine flow with that produced by injecting vasopressin. The antidiuretic a c t i v i t y of the extract from 0.25 ml. plasma was represented as corresponding to 0.1 m-u. vasopressin before balloon i n f l a t i o n and 0.06 m-u. immediately after balloon i n f l a t i o n . The method was reported to allow assay of antidiuretic a c t i v i t y to within 0.02 m-u. vasopressin, the difference reported may therefore be only just within the limits of the method. The authors claimed that diuresis did not follow balloon i n f l a t i o n after the supra-optic© hypophysial tracts had been sectioned, Fig. 5 of their paper shows such an experiment; a balloon was inflated i n the l e f t atrium for three minutes and was followed by a marked f a l l i n mean a r t e r i a l pressure of approximately 40 mm. Hg. which lasted about twenty-five minutes; under these conditions i t would indeed have been surprising i f urine flow had increased. Shu'ayb et a l . (1965) who employed the same methods of hydration, anesthesia and a t r i a l distension as those reported by Ledsome et a l . (1961) and the present investigation "concluded that the diuresis which follows distension of the l e f t atrium i s caused by a decrease i n blood antidiuretic hormone". However, of the seven figures i n their paper in which l e f t a t r i a l distension was associated with a diuresis four of these figures indicate that the level of antidiuretic a c t i v i t y was already decreasing before distension of the l e f t atrium, and began increasing before pressure i n the l e f t atrium was returned to normal, two other 83. figures indicate that the diuretic response to l e f t a t r i a l distension occurred although there was no decrease i n the level of antidiuretic a c t i v i t y from the period before a t r i a l distension, the remaining figure while demonstrating a decrease in blood antidiuretic a c t i v i t y i n response to l e f t a t r i a l distension from a previously steady level also demonstrates that the diuretic response occurred after release of the distension and at a time when the level of antidiuretic a c t i v i t y was increasing dramatically. The authors claimed that a significant or two fold increase i n urine flow did not occur unless levels of blood antidiuretic a c t i v i t y decreased i n response to l e f t a t r i a l distension to 2.2 uU/ml. or less. Upon closer examination of their results a decrease i n blood antidiuretic a c t i v i t y from 3.9 }iU/ml. to 1.9 jaU/ml. i s only associated with an increase i n urine flow from 0.20 ml./min. to 0.30 ml./min., a 50$ increase, while i n another instance a decrease i n blood antidiuretic a c t i v i t y from 29.7 ^iU/ml. to 6.7 uU/ml i s associated with an increase i n urine flow from 0.04 ml./min. to 0.42 ml./min., a ten fold increase. Thus i t cannot be concluded from the studies by Baisset and Montastruc (1959) and Shu'ayb et a l . (1965) that l e f t a t r i a l distension i s causing a diuresis by reducing the circulating concentration of antidiuretic hormone. However, the study by Johnson et a l . (1969) does demonstrate a direct relationship between an increase i n l e f t a t r i a l transmural pressure, a decrease i n the plasma concentration of antidiuretic a c t i v i t y and a dilute diuresis. These authors employed small balloon inflations in anesthetized dogs which increased l e f t a t r i a l transmural pressure an average of 4.4 cm. H2O. Their results indicated a highly significant linear correlation between increases i n l e f t a t r i a l transmural pressure from 1 - 7 cm. HgO and decreases i n plasma antidiuretic a c t i v i t y from 0.5 - 5.0 jaU/ml. They reported an average decrease i n plasma antidiuretic a c t i v i t y from 6.3 84. to 4.4 pU/ml. This decrease was associated with a highly significant decrease in urine concentration and water reabsorption and a highly significant increase i n urine flow. The magnitude of their reported changes in renal function were small however and their results could be interpreted as indicating that the decrease in the plasma antidiuretic a c t i v i t y pro-duced by an increase in l e f t a t r i a l transmural pressure i s not sufficient to contribute significantly to the diuretic response to l e f t a t r i a l dis-tension. It therefore remains with the results of the present investigation to elucidate the role of antidiuretic hormone in the diuretic response to l e f t a t r i a l distension. Had the a t r i a l distension experiments been carried out using only a single high dose of vasopressin infusion an interpretation of the results may have been complicated by changes in cardiovascular function which have been shown to occur at relatively low doses of vasopressin infusion (present experiments). Rather the adoption of an experimental design involving increasing doses of vasopressin in a dose-response relationship permits an assessment of the effects of vasopressin on the diuretic response to l e f t a t r i a l distension. This view is further j u s t i f i e d since the sequence of the vasopressin doses was randomized and between doses no significant changes in the measured cardiovascular or plasma variables occurred. The diuretic response to l e f t a t r i a l distension i s the result of an increase i n solute excretion and a decrease in the reabsorption of water. The fact that vasopressin could not abolish the diuretic response nor affect the increase i n solute excretion indicates that a significant part of the diuretic response to l e f t a t r i a l distension could be an increase i n solute excretion. The effect of increasing doses of vasopressin on the diuresis indicates that one role for vasopressin i s to alter the characteristics of the diuresis from a diuresis involving an increase in 85. solute excretion and water excretion to one involving solely an increase i n solute excretion. The fact that increases i n l e f t a t r i a l transmural pressure w i l l reduce the circulating concentration of plasma antidiuretic a c t i v i t y and cause a decrease in the reabsorption of water (Johnson et a l . , 1969), that this effect of decreased water reabsorption can be progressive-l y reduced and entirely prevented by increasing doses of vasopressin (present experiments) and that a decreased water reabsorption can be produced by changing the rate of vasopressin infusion at relatively higher rates of infusion (present experiments) i s conclusive evidence that the increase i n free water clearance in response to l e f t a t r i a l distension i s the result of a decrease i n the circulating concentration of antidiuretic hormone. Furthermore this increase i n free water clearance can account for a significant part of the diuretic response to l e f t a t r i a l distension as demonstrated in the present investigation during the saline infusion or i t can account for a l l of the diuretic response to l e f t a t r i a l distension as demonstrated i n the present investigation when urine osmolality i n the control period was very low. G. THE ROLE OF VASOPRESSIN AS CONTROLLED BY  LEFT ATRIAL DISTENSION IN THE CONTROL OF  BODY FLUID VOLUME. 1. Normal Function. In reference to the normal control of body f l u i d volume i t has often been observed that manouvers which distend the intrathoracic circulation result in the case of a well hydrated individual i n a rather dilute diuresis and in the case of a normohydrated or dehydrated individual i n a diuresis, a significant part of which i s an increase i n solute excretion (Lydtin and Hamilton, 1964, Behn, Gauer, Kirsch and Eckert, 1969). It i s possible to correlate a dehydrated individual at rest with a higher circulating level of antidiuretic hormone than a well hydrated individual at rest (Czaczkes 86. et a l . 1964). This p o s s i b i l i t y when placed i n conjunction with the results of the present investigation which indicate the effect increasing doses of vasopressin may have on altering the characteristics of the diuretic response to l e f t a t r i a l distension may provide an explanation for the different response patterns to distension of the intrathoracic circulation relative to the degree of hydration. The results of the present investigation are also significant in terms of the weightless condition achieved in space travel, a condition which i s known to cause a "relative degree of engorgement of the intrathoracic circulation", (Gauer et a l . 1970). The present results indicate that i t cannot be presumed that in the weightless condition the acquisition of relatively high circulating levels of antidiuretic hormone w i l l prevent a loss of solute from the body f l u i d s . The p o s s i b i l i t y must also be considered that i n the "normal" conscious dog i f small changes in l e f t a t r i a l pressure such as those reported by Johnson et a l . (1969) were to occur with a circulating level of antidiuretic much larger changes in renal function may result. Furthermore, there i s no reason to suppose that the effect of antidiuretic hormone on renal function ismanifest over such a limited range as the studies by Shannon (1941) and Verney (1947) would indicate. Their studies were done with a dog i n the "abnormal" state of water diuresis and there i s no reason to suppose that this state should give any more "normal" results than the results from the vasopressin infusion experiments in the present investigations. 2. Abnormal Function. Distension of the l e f t atrium and the control exerted by this dis-tension on the circulating l e v e l of antidiuretic hormone has been implicated as being involved in several pathological states of c l i n i c a l a c t i v i t y in the control period lower than them 87. interest. In chronic heart failure the greatly increased and prolonged a t r i a l stretch accompanying the elevated venous pressure may lead to a loss of sensitivity in the a t r i a l receptors controlling secretion of anti-diuretic hormone and thereby contribute to the water retention seen in this condition. The postcommissuratomy hyponatremic syndrome which may persist for days after commissurotomy and is accompanied by water retention has been reasonably explained by Shu'ayb et a l (1965) as being due to a sudden decrease in tension i n the l e f t atrium which results i n a remarkable increase i n the circulating level of antidiuretic hormone. This same mechanism has also been implicated as being important i n the defect i n water excretion observed in post operative cardiac patients in which even very modest oral water loads produced a severe hyponatremia. In some attacks of paroxysmal tachycardia i n which there may be an abrupt increase i n a t r i a l mean pressure and pulse pressure to double the resting state, together with a pulse rate up to 200 or more per min. patients have discharged abundant dilute urine during or immediately after the attacks. Kilburn (1964) reported that e l e c t r i c a l pacing of the normal dogs l e f t atrium to 200/min. or more produces a diuresis which could be inhibited by infusion of antidiuretic hormone and he attributed the diuresis resulting from an increased heart rate to stimulation of receptors i n the l e f t atrium. H- CONCLUSIONS CONCERNING THE ROLE OF  ANTIDIURETIC HORMONE IN THE DIURETIC JHESFONSE TO LEFT ATRIAL DISTENSION. The evidence clearly indicates that the diuretic response to l e f t a t r i a l distension can be composed of an increase i n osmolal clearance and/or free water clearance. The increase i n free water clearance in the present investigation i s dependent on the circulating level of vasopressin. As the circulating level of vasopressin increased the increase in free water clearance i s reduced or abolished. The increase i n free water clearance in response to l e f t a t r i a l distension can be reproduced by altering the rate of infusion of vasopressin in dogs anesthetized and hydrated i n similar manner to those used i n the a t r i a l distension experiments. These facts i n conjunction with the report by Johnson et a l . ( 1 9 6 9 ) indicate that the variable of free water clearance i n the diuretic response to l e f t a t r i a l distension i s controlled mainly by antidiuretic hormone. The increase i n osmolal clearance i n the present investigation appears to be independent of the circulating level of vasopressin and the mechanisms responsible for concentration and dilution of the urine. Large changes in vasopressin concentration which had hitherto been conceived of as representing maximum antidiuretic a c t i v i t y are capable of causing a dilute transient diuresis. P A R T V. B I B L I O G R A P H Y . Arndt, J.O., Reineck, H., and Gauer, O.H. (1963). Ausscheidungsfunktion und Hamodynamik der Wieren bei Dehnung des linken Vorhofes am narkotisierten Hund. Arch. ges. Physiol. 277, 1-15. Arndt, J.O. (1965). Diuresis induced by water infusion into the carotid loop and i t s inhibition by small hemorrhage. Pflugers Archiv. 282, 313 - 322. Atherton, J.C, Hai, M.A. and Thomas, S. (1969). Acute effects of lysine-vasopressin injection (single and continuous) on urinary com-position in the conscious water-diuretic rat. Pflugers Archiv. 310, 281 - 296. 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