AMERICAN
JOURNAL
OF
Vol. 230, No. 4, April
PHYSIOLOGY
1976.
Printed
in U.S.A.
Effect of prolonged infusion on thirst
angiotensin
NICK C. TRIPPODO, ROBERT E. McCAA, Department of Physiology and Biophysics, Mississippi 39216
AND ARTHUR C. GUYTON University of Mississippi School
TRIPPODO, NICK C., ROBERT E. MCCAA, AND ARTHUR C. GUYTON. Effect of prolonged angiotension II infusion on thirst. Am. J. Physiol. 230(4): 1063-1066. 1976. -To determine the effect of prolonged angiotensin II (A-II) infusion on thirst, daily water intake by drinking was measured in dogs during a 4day control period, a 4day period of vehicle infusion without A-II, a lo-day period of A-II infusion, and a 4-day recovery period of vehicle infusion without A-II. During the control period and the periods of vehicle infusion in the absence of A-II, daily water intake by drinking in four dogs averaged 118 2 20 ml/day (mean t SE). During the lo-day period of A-II infusion at the rate of 13.0 ngfkg per min drinking increased to 269 2 49 ml/day (paired t; P < 0.05). Angiotensin II infusion at the rate of 26.0 rig/kg per min produced a sustained increase in water intake in two dogs during an 8-day period of infusion. These results demonstrate that in dogs, prolonged infusion of angiotensin II stimulates the thirst mechanism and that the effect lasts for more than a few days.
aldosterone ance
secretion
in dogs; water
balance;
electrolyte
II
bal-
(9, 10) found that bilateral nephrectomy reduced the increased water drinking induced in rats by either ligation of the inferior vena cava or constriction of the renal artery. He also found that drinking was elicited by the administration of renal extract to rats (10). A number of investigators have since shown that administration of angiotensin II (A-II), either peripherally or centrally, into the brain stimulates thirst in a variety of species (2, 12). According to Fitzsimons’ (11) model during low hemodynamic states, increased amounts of A-II (resulting from increased renal renin release) enhances drinking by acting on thirst areas in the diencephalon. In support of the concept that circulating A-II affects central thirst receptors, Cooling and Day (8) found in cats that centrally administered antagonist of A-II reduced the dipsogenic response to intravenously administered A-II, and Simpson and Routtenberg (20) demonstrated that lesions in the subfornical organ in the brain of rats also reduced drinking induced by intravenous injections of A-II. Increased water drinking has been reported in dogs with Goldblatt hypertension (15) and in patients with juxtaglomerular cell tumors (5, 7, 13, 19). Brown et al. (4) reported in man that polydipsia associated with high levels of plasma renin ceased after nephrectomy. It is FITZSIMONS
of Medicine,
Jackson,
possible that the high concentration of blood A-II that was associated with these states played an important role in the increased water intake. However, Abraham et al. (1) questioned whether physiological changes in blood A-II concentration could act as a sole factor in the genesis of thirst. These investigators failed to stimulate water drinking in sheep by intravenous infusion of A-II at levels within the physiological range that has been observed so far in that species. Yet when pharmacological levels were produced in the brain by intracarotid or centrally administered A-II, drinking occurred immediately. They pointed out that similar quantitative factors should be considered in relation to data previously reported in other species. In most cases the dipsogenic effect of A-II has been studied during short-term experiments with infusions not lasting longer than 24 h. Abraham et al. (1) infused A-II for 3 days in sheep, but the animals did not show an increase in drinking. The present study was conducted to determine the effect of high blood levels of A-II on daily water intake in dogs and to see whether or not the effect persists for more than a few days. In contrast to the findings in sheep, we found that dogs showed a significant increase in water drinking in response to prolonged intravenous infusion of A-II. METHODS
Mongrel dogs of either sex with body weights between 14.4 and 23.6 kg were anesthetized with sodium pentobarbital (30 mg/kg) and surgically prepared with femoral arterial and venous catheters, which ran subcutaneously from the femoral triangle region to the region dorsal to the pectoral girdle and out through a small skin incision. The dogs were fitted with jackets made of heavy cotton material to protect the catheters. Two weeks were allowed for recovery from the surgery, during which time the dogs were placed on a fixed diet consisting of two cans per day of Prescription Diet k/d (Riviana Foods, Inc.) and water ad libitum. Sodium chloride was added to the food to give a final daily intake of 50 meq/day of sodium and 28 meq/day of potassium. This diet was maintained throughout the experimental period, was eaten completely on each day by all dogs in the study, and consisted of approximately 700 ml/day of water in the food. Each dog was placed in a metabolic cage and kept in
1063
Downloaded from www.physiology.org/journal/ajplegacy by ${individualUser.givenNames} ${individualUser.surname} (133.006.082.173) on January 14, 2019.
TRIPPODO,
1064 an air-conditioned room. Ambient temperature and humidity remained relatively constant. A plaster of Paris harness with a pulley system to counterbalance the harness was attached to the dog and held in place on its The venous catheter was back by leather straps. brought out through an opening in the dorsal aspect of the harness and connected to a Harvard syringe pump at the top of the cage. The arterial catheter was connected to a Statham pressure transducer that was built into the side of the harness. After each dog was placed in the harness, its daily water intake was measured for 1 wk to test its suitability for study. Five of eleven dogs were eliminated for the following reasons: elevated body temperature and suspected infection (two), diarrhea (one), persistent destruction of harness (one), and copious water drinking with volumes approaching 2 liters/ day (one). Data were collected on the selected dogs during four successive periods: 4 days of preinfusion, 4 days of vehicle infusion without angiotensin II, 10 days of vehicle infusion containing angiotensin II (A-II), and again 4 days of vehicle infusion without angiotensin. The vehicle infusion consisted of either 0.9% NaCl or 5% mannito1 infused continuously intravenously by a syringe pump at a flow rate of either 26 or 52 ml/day. In four dogs, A-II (Hypertensin, Ciba) was added to the vehicle to give a dose rate of 13 rig/kg per min during the lo-day A-II period. In two dogs, the dose rate of A-II was 13 ng/ kg per min for the first 2 days and 26 rig/kg per min for the next 8 days. In all dogs except one, mean arterial blood pressure was monitored continuously for approximately 6 h/day and recorded on a Grass polygraph. Except in cases where there were technical problems (clot in catheter, etc.) 7-10 readings (approximately 40 min apart) were taken from the 6-h record of mean arterial pressure and averaged to give the daily value for a particular dog. Arterial blood samples (25 ml) were collected every morning from the dogs that received A-II at 13 rig/kg per min, and every other morning from the dogs that received A-II at 26 rig/kg per min. The hematocrits were measured and the blood was centrifuged at a temperature of 4°C. The plasma was frozen for later analysis and the blood cells were resuspended in dextran 70 and reinfused into the animals. Daily water intake by drinking and urinary volumes were measured each morning. Plasma and urinary sodium and potassium concentrations were determined by flame photometry. Plasma renin activity was determined by radioimmunoassay using the Squibb Angiotensin I Immunotope Kit. Plasma aldosterone concentration was determined by radioimmunoassay as previously described (17). Plasma cortisol concentration was determined using the method reported by Murphy (18). Student’s t test for paired data was used to analyze the data from the four dogs that received A-II infusion at 13 rig/kg per min. Statistical significance was accepted at P 5 0.05. RESULTS
Water intake by drinking and urine output. Figure lA shows the mean daily water intake bv drinking of the
6
McCAA,
AND
GUYTON
i
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1. Effect of per min for 10 days fBj, and potassium tion (D,. Each bar sents average value FIG.
intravenous infusion of angiotensin II at 13 rig/kg on water intake by drinking (A), urinary sodium CC, excretion, and plasma aldosterone concentrain graphs A-C and each point in graph D repreof four dogs. Standard errors are indicated.
four dogs that received A-II at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, the average water intake by drinking was 118 2 20 ml/day (12-day mean t SE). With the 700 ml/day of water in the food, this gave an average total water intake of about 820 ml/day. During the 10 days of A-II infusion, water intake by drinking averaged 269 2 49 ml/day (IO-day mean t SE). The mean difference and its SE between the average daily water drunk while the dogs received A-II and the average daily water drunk during the three periods when A-II was not infused was 151 + 45 ml/day (paired t; P ~0.05). Figure 2 illustrates the daily water intake by drinking of the two dogs that received A-II at 26 rig/kg per min for 8 days following 2 days at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, water intake by drinking in the two dogs averaged 302 and 903 ml/day; during A-II infusion it averaged 657 and 1,705 ml/day, respectively. The average daily urine output of the four dogs that received A-II at 13 rig/kg per min during the “non-A-II” and “A-II” periods were 425 t 29 and 527 5 43 ml/day, respectively. The mean difference and its SE between these two values was 102 2 24 ml/day (paired t; P < 0.05). Similar increases in urine output were observed during A-II infusion in the two dogs that received A-II at 26 rig/kg per min.
Downloaded from www.physiology.org/journal/ajplegacy by ${individualUser.givenNames} ${individualUser.surname} (133.006.082.173) on January 14, 2019.
ANGIOTENSIN
II AND ,,,,$,&,,
1065
THIRST DISCUSSION
VEHICLE
24
Electrolyte balance and hematocrit. There were no large changes in plasma sodium and potassium concentrations or in hematocrit in the dogs that received A-II at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, the average values for plasma sodium concentration, plasma potassium concentration, and hematocrit were: 143 meq/liter, 4.4 meq/liter, and 38%. During A-II infusion the respective values were 142 meq/liter, 4.3 meq/liter and 36%. Urinary sodium and potassium excretion are shown in Fig. 1B and C. The average daily sodium excretion during the non-A-II and A-II periods were 40 t 2 and 38 t 3 meq/day, respectively (not significantly different,). For potassium excretion the respective values were 25 ? 2 and 27 _ + 4 meq/day (not significantly different). Similar values were observed for these variables in the two dogs that received A-II at 26 rig/kg per min.
The daily water intake by drinking in dogs was significantly elevated during prolonged intravenous infusion of angiotensin (A-II) at a rate of 13 rig/kg per min. By radioimmunoassay, B. H. Douglas (personal communication) found that intravenous infusion of A-II in dogs at dose rates comparable to 13 rig/kg per min for 4 h produced blood A-II levels in the range of 200-300 pg/ml. He also found that at the end of a 3-day sodium-depletion period in dogs, blood levels of A-II often attain concentrations of 200-300 pg/ml. Values as high as 1,000 pg/ml have been reported in sodium-depleted sheep (3). In man with renal hypertension, blood A-II levels reached 175 ,t 91 pg/ml (6). Thus, it is reasonable to state that the infusion rate of 13 rig/kg per min used in the present study produces a blood A-II concentration that is within the physiologically feasible range for dogs and that it is capable of stimulating the thirst mechanism. These results are in disagreement with the findings in sheep, which showed no increase in drinking during intravenous infusion of A-II at rates of 5-40 pg/h (2-17 rig/kg per min) for 3 days (1). Such infusion rates in sheep produce levels of A-II within the physiological limits of that species. Since sheep are ruminant animals, the discrepancy between the two studies might be related to differences in experimental animals. The two dogs that received 26 rig/kg per min of A-II showed sustained increased drinking throughout the 8 days of infusion at this rate. Although the blood levels of A-II produced by this rate of infusion may have been out of the physiological limits, the results suggest that the dipsogenic response to pharmacological doses of A-II does not show tachyphylaxis. There is a paucity of data on the dipsogenic effect of A-II in dogs; however, in one study Kozlowski et al. (14) reported that A-II infused intravenously into dogs at a rate of 50 ng/min (2-3 rig/kg per min) lowered the “thirst threshold.” That is, A-II decreased the degree of cellular dehydration (produced by 5% sodium chloride infusion)
Plasma aldosterone concentration, plasma cortisol concentration, mean arterial pressure, and plasma renin actiuity. The mean daily plasma aldosterone con-
on mean
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DAYS
FIG. 2. Effect of intravenous per min for 8 days (following intake by drinking. ’
infusion 13 rig/kg
of angiotensin II at 26 rig/kg per min for 2 days) on water
centration of the dogs that received A-II at 13 nglkg per min are shown in Fig. m. Plasma aldosterone averaged 11.4 t 0.2 ng/lOO ml during the non-A-II period and 13.7 t 1.3 ng/lOO ml during the A-II period (not significantly different). Plasma cortisol concentration did not change significantly. Mean arterial pressure remained elevated 30-35 mm Hg above the control value during A-II infusion at either dose rate in all dogs in which it was measured, except dog 3 (Table 1). Likewise, in all dogs except dog 3, plasma renin activity remained suppressed during the greater part of A-II infusion. Control values for plasma renin activity in dog 11 were probably high during the control period because this animal, which weighed 21 kg, lost approximately 300 ml of blood through the arterial catheter several days before the start of the experiment. The changes in mean arterial pressure and plasma renin activity could not be shown to have statistical significance.
TABLE
DOG No. Days
1. Effect
of prolonged infusion pressure and plasma
arterial Preinfusion 2
Vehicle 4
2 Mean
2 3 5 12 12
86 87 100 101 83
94 77 100 99 95
I
99 76 98 95 92
Plasma
Angiotensin 4
2
arterial 101 88 95 93 78
133 81 130 117 113
renin
actrvitv,
II + Vehicle
4
pressure,
of angiotensin renin activity
6
II
Vehicle
8
10
2
4
134 82 144 144 134
80 130 134 128
68 99 84 74
84 99 88 91
1.12 1.29 0.67 0.16 0.91 0.51
0.48 1.15 0.62 0.24 0.86 1.20
mmHg
131 75 123 130 121 ng A-Ilml
82 133 128 116
per h
During the vehicle infusion periods, either 0.9% saline or 5% mannitol continuously by way of indwelling venous catheters and syringe pumps at either day. The dose rate of angiotensin II was 13 rig/kg per min in dogs 2,3. 5, and and 12, angiotensin II was infused at 13 rig/kg per min for 2 days, followed by min for 8 days. Mean arterial pressure was monitored for 6 h each day by way arterial catheters.
was infused 26 or 52 ml/ 7. In dogs 21 26 rig/kg per of indwelling
Downloaded from www.physiology.org/journal/ajplegacy by ${individualUser.givenNames} ${individualUser.surname} (133.006.082.173) on January 14, 2019.
TRIPPODO,
1066 necessary to induce a drinking response. A-II infused alone at this rate (without simultaneous sodium chloride infusion) did not induce the dogs to drink spontaneously. However, the infusion was carried out only for 3 h. Also, unlike the findings in sheep (l), there was no increase in urinary sodium excretion during the first 2 days of A-II infusion in the present study. In fact, there was a tendency for sodium excretion to’decrease during that time. This agrees with the results of Urquhart et al. (21), who reported sodium retention in dogs during the first 2 days of A-II infusion at 20 pg/kg per day (14 rig/kg per min). In the dogs that received A-II at 13 rig/kg per min
McCAA,
AND
GUYTON
plasma aldosterone concentration was only slightly above the control value after the third day of A-II infusion. McCaa et al. (16) demonstrated that continuous infusion of A-II at 5 rig/kg per min for 14 days produced a significant increase in plasma aldosterone concentration in dogs only during the first day of infusion. The authors are grateful to Terry S. Finch, Florence N. son, Michael O’Ryan McGee, and Mary Frances Plummer excellent technical assistance. This work was supported by NIH grants HL 11678,09921, 00316. N. C. Trippodo was an American and Mississippi Heart tion Fellow during the time of this study. Received
for publication
30 June
Hutchifor their and GM Associa-
1975.
REFERENCES 1. ABRAHAM, S. F., R. M. BAKER, E. H. BLAINE, D. A. DENTON, AND M. J. MCKINLEY. Water drinking induced in sheep by angiotensin - a physiological or pharmacological effect? J. Comp. Physiol. Psychol. 8: 503-518, 1975. 2. ANDERSON, B., AND 0. WESTBYE. Synergistic action of sodium and angiotensin on brain mechanisms controlling water and salt balance. Nature 228: 75, 1970. 3. BLAIR-WEST, J. R., M. D. CAIN, K. J. GATT, J. P. COGHLAN, D. A. DENTON, J. W. FUNDER, B. A. SCOGGINS, AND R. D. WRIGHT. The dissociation of aldosterone secretion and systemic renin and angiotensin II levels during the correction of sodium deficiency. Acta. Endocrinol. 66: 229-247, 1971. 4. BROWN, J. J., J. R. CURTIS, A. F. LEVER, J. I. S. ROBERTSON, H. E. DEWARDENER, AND A. J. WING. Plasma renin concentration and the control of blood pressure in patients on maintenance haemodialysis. Nephron 6: 329-349, 1969. 5. BROWN, J. J., A. F. LEVER, J. I. S. ROBERTSON, R. FRASER, J. J. MORTON, M. TREE, P. R. F. BELL, J. K. DAVIDSON, AND I. S. RUTHVEN. Hypertension and secondary hyperaldosteronism associated with a renin-secreting renal juxtaglomerular-cell tumor. Lancet 2: 1228-1232, 1973. 6. CATT, K. J., M. D. CAIN, J. P. COGHLAN, P. Z. ZIMMET, E. CRAN, AND J. B. BEST. Metabolism and blood levels of angiotensin II in normal subjects, renal disease, and essential hypertension. Circulation Res. 27, Suppl. 2: 11177-11193, 1970. 7. CONN, J. W., E. L. COHEN, C. P. LUCAS, W. J. MCDONALD, G. H. MAYOR, W. M. BLOUGH, JR., W. C. EVELAND, J. J. BOOKSTEIN, AND J. LAPIDES. Primary reninism. Arch. fntern. Med. 130: 682696, 1972. 8. COOLING, M. J., AND M. D. DAY. Inhibition of renin-angiotensin induced drinking in the cat by enzyme inhibitors and by analogue antagonists of angiotensin II. Chin. Exptl. PharmacoZ. PhysioZ. 1: 389-396, 1974. 9. FITZSIMONS, J. T. Drinking caused by constriction of the inferior vena cava in the rat. Nature 204: 479-480, 1964. 10. FITZSIMONS, J. T. The role of a renal thirst factor in drinking induced by extracellular stimuli. J. Physiol., London 201: 349368, 1969. 11. FITZSIMONS, J. T. The phvsiologv of thirst: a review of the extra-
12. 13.
14.
15.
16.
17.
18.
19.
20.
21.
neural aspects of the mechanisms of drinking. In: Progress in PhysiologicaL Psychology, edited by E. Stellar and J. Sprague. New York: Academic, 1971, vol. 4, p. 119-201. FITZSIMONS, J. T. Thirst. Physiol. Rev. 52: 468-561, 1972. KIHARA, I., S. KITAMURA, T. HOSHINO, S. HITOSHI, AND T. WATANABE. A hitherto unreported vascular tumor of the kidney: a proposal of “juxtaglomerular cell tumor.” Acta Physiol. Japan. 18: 197-206, 1968. KOZLOWSKI, S., K. DRZEWIECKI, AND W. ZURAWSKI. Relationship between osmotic reactivity of the thirst mechanism and the angiotensin and aldosterone level in the blood of dogs. Acta. Physiol. Polon. 23: 369-376, 1972. LIARD, J., A. W. COWLEY, JR., R. E. MCCAA, C. S. MCCAA, AND A. C. GUYTON. Renin, aldosterone, body fluid volumes, and the baroreceptor reflex in the development and reversal of Goldblatt hypertension in conscious dogs. Circulation Res. 34: 549-560, 1974. MCCAA, R. E., C. S. MCCAA, AND A. C. GUYTON. Role of angiotensin II and potassium in the long-term regulation of aldosterone secretion in intact conscious dogs. CircuZation Res. 36, Suppl. 1: 157-167, 1975. MCCAA, R. E., C. S. MCCAA, D. G. READ, J. D. Bower, and A. C. GUYTON. Increased plasma aldosterone concentration in response to hemodialysis in nephrectomized man. Circulation Res. 31: 473-480, 1972. MURPHY, B. E. P. Some studies of the protein-binding of steroids and their application to the routine micro and ultra-micro measurement of various steroids in body fluids by competitive protein-binding radioassay. J. Clin. Endocrinol. Metab. 27: 973-990, 1967. SCHAMBELAN, M., E. L. HOWES, J. R. STOCKIGT, C. A. NOAKES, AND E. G. BIGLIERI. Role of renin and aldosterone in hypertension due to a renin-secreting tumor. Am. J. Med. 55: 86-92, 1973. SIMPSON, J. B., AND A. ROUTTENBERG. Subfornical organ lesions reduce intravenous angiotensin-induced drinking. Brain Res. 88: 154-161, 1975. URQUHART, J., J. 0. DAVIS, AND J. T. HIGGINS, JR. Effects of prolonged infusion of angiotensin II in normal dogs. Am. J. Phwiol. 205: 1241-1246. 1963.
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JOURNAL
OF
Vol. 230, No. 4, April
PHYSIOLOGY
1976.
Printed
in U.S.A.
Effect of prolonged infusion on thirst
angiotensin
NICK C. TRIPPODO, ROBERT E. McCAA, Department of Physiology and Biophysics, Mississippi 39216
AND ARTHUR C. GUYTON University of Mississippi School
TRIPPODO, NICK C., ROBERT E. MCCAA, AND ARTHUR C. GUYTON. Effect of prolonged angiotension II infusion on thirst. Am. J. Physiol. 230(4): 1063-1066. 1976. -To determine the effect of prolonged angiotensin II (A-II) infusion on thirst, daily water intake by drinking was measured in dogs during a 4day control period, a 4day period of vehicle infusion without A-II, a lo-day period of A-II infusion, and a 4-day recovery period of vehicle infusion without A-II. During the control period and the periods of vehicle infusion in the absence of A-II, daily water intake by drinking in four dogs averaged 118 2 20 ml/day (mean t SE). During the lo-day period of A-II infusion at the rate of 13.0 ngfkg per min drinking increased to 269 2 49 ml/day (paired t; P < 0.05). Angiotensin II infusion at the rate of 26.0 rig/kg per min produced a sustained increase in water intake in two dogs during an 8-day period of infusion. These results demonstrate that in dogs, prolonged infusion of angiotensin II stimulates the thirst mechanism and that the effect lasts for more than a few days.
aldosterone ance
secretion
in dogs; water
balance;
electrolyte
II
bal-
(9, 10) found that bilateral nephrectomy reduced the increased water drinking induced in rats by either ligation of the inferior vena cava or constriction of the renal artery. He also found that drinking was elicited by the administration of renal extract to rats (10). A number of investigators have since shown that administration of angiotensin II (A-II), either peripherally or centrally, into the brain stimulates thirst in a variety of species (2, 12). According to Fitzsimons’ (11) model during low hemodynamic states, increased amounts of A-II (resulting from increased renal renin release) enhances drinking by acting on thirst areas in the diencephalon. In support of the concept that circulating A-II affects central thirst receptors, Cooling and Day (8) found in cats that centrally administered antagonist of A-II reduced the dipsogenic response to intravenously administered A-II, and Simpson and Routtenberg (20) demonstrated that lesions in the subfornical organ in the brain of rats also reduced drinking induced by intravenous injections of A-II. Increased water drinking has been reported in dogs with Goldblatt hypertension (15) and in patients with juxtaglomerular cell tumors (5, 7, 13, 19). Brown et al. (4) reported in man that polydipsia associated with high levels of plasma renin ceased after nephrectomy. It is FITZSIMONS
of Medicine,
Jackson,
possible that the high concentration of blood A-II that was associated with these states played an important role in the increased water intake. However, Abraham et al. (1) questioned whether physiological changes in blood A-II concentration could act as a sole factor in the genesis of thirst. These investigators failed to stimulate water drinking in sheep by intravenous infusion of A-II at levels within the physiological range that has been observed so far in that species. Yet when pharmacological levels were produced in the brain by intracarotid or centrally administered A-II, drinking occurred immediately. They pointed out that similar quantitative factors should be considered in relation to data previously reported in other species. In most cases the dipsogenic effect of A-II has been studied during short-term experiments with infusions not lasting longer than 24 h. Abraham et al. (1) infused A-II for 3 days in sheep, but the animals did not show an increase in drinking. The present study was conducted to determine the effect of high blood levels of A-II on daily water intake in dogs and to see whether or not the effect persists for more than a few days. In contrast to the findings in sheep, we found that dogs showed a significant increase in water drinking in response to prolonged intravenous infusion of A-II. METHODS
Mongrel dogs of either sex with body weights between 14.4 and 23.6 kg were anesthetized with sodium pentobarbital (30 mg/kg) and surgically prepared with femoral arterial and venous catheters, which ran subcutaneously from the femoral triangle region to the region dorsal to the pectoral girdle and out through a small skin incision. The dogs were fitted with jackets made of heavy cotton material to protect the catheters. Two weeks were allowed for recovery from the surgery, during which time the dogs were placed on a fixed diet consisting of two cans per day of Prescription Diet k/d (Riviana Foods, Inc.) and water ad libitum. Sodium chloride was added to the food to give a final daily intake of 50 meq/day of sodium and 28 meq/day of potassium. This diet was maintained throughout the experimental period, was eaten completely on each day by all dogs in the study, and consisted of approximately 700 ml/day of water in the food. Each dog was placed in a metabolic cage and kept in
1063
Downloaded from www.physiology.org/journal/ajplegacy by ${individualUser.givenNames} ${individualUser.surname} (133.006.082.173) on January 14, 2019.
TRIPPODO,
1064 an air-conditioned room. Ambient temperature and humidity remained relatively constant. A plaster of Paris harness with a pulley system to counterbalance the harness was attached to the dog and held in place on its The venous catheter was back by leather straps. brought out through an opening in the dorsal aspect of the harness and connected to a Harvard syringe pump at the top of the cage. The arterial catheter was connected to a Statham pressure transducer that was built into the side of the harness. After each dog was placed in the harness, its daily water intake was measured for 1 wk to test its suitability for study. Five of eleven dogs were eliminated for the following reasons: elevated body temperature and suspected infection (two), diarrhea (one), persistent destruction of harness (one), and copious water drinking with volumes approaching 2 liters/ day (one). Data were collected on the selected dogs during four successive periods: 4 days of preinfusion, 4 days of vehicle infusion without angiotensin II, 10 days of vehicle infusion containing angiotensin II (A-II), and again 4 days of vehicle infusion without angiotensin. The vehicle infusion consisted of either 0.9% NaCl or 5% mannito1 infused continuously intravenously by a syringe pump at a flow rate of either 26 or 52 ml/day. In four dogs, A-II (Hypertensin, Ciba) was added to the vehicle to give a dose rate of 13 rig/kg per min during the lo-day A-II period. In two dogs, the dose rate of A-II was 13 ng/ kg per min for the first 2 days and 26 rig/kg per min for the next 8 days. In all dogs except one, mean arterial blood pressure was monitored continuously for approximately 6 h/day and recorded on a Grass polygraph. Except in cases where there were technical problems (clot in catheter, etc.) 7-10 readings (approximately 40 min apart) were taken from the 6-h record of mean arterial pressure and averaged to give the daily value for a particular dog. Arterial blood samples (25 ml) were collected every morning from the dogs that received A-II at 13 rig/kg per min, and every other morning from the dogs that received A-II at 26 rig/kg per min. The hematocrits were measured and the blood was centrifuged at a temperature of 4°C. The plasma was frozen for later analysis and the blood cells were resuspended in dextran 70 and reinfused into the animals. Daily water intake by drinking and urinary volumes were measured each morning. Plasma and urinary sodium and potassium concentrations were determined by flame photometry. Plasma renin activity was determined by radioimmunoassay using the Squibb Angiotensin I Immunotope Kit. Plasma aldosterone concentration was determined by radioimmunoassay as previously described (17). Plasma cortisol concentration was determined using the method reported by Murphy (18). Student’s t test for paired data was used to analyze the data from the four dogs that received A-II infusion at 13 rig/kg per min. Statistical significance was accepted at P 5 0.05. RESULTS
Water intake by drinking and urine output. Figure lA shows the mean daily water intake bv drinking of the
6
McCAA,
AND
GUYTON
i
4
I I1 I I 12341234123456789101234
I
I !
11
11 I
i I1
I
I1
1
I,
I,,,, I
DAYS
1. Effect of per min for 10 days fBj, and potassium tion (D,. Each bar sents average value FIG.
intravenous infusion of angiotensin II at 13 rig/kg on water intake by drinking (A), urinary sodium CC, excretion, and plasma aldosterone concentrain graphs A-C and each point in graph D repreof four dogs. Standard errors are indicated.
four dogs that received A-II at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, the average water intake by drinking was 118 2 20 ml/day (12-day mean t SE). With the 700 ml/day of water in the food, this gave an average total water intake of about 820 ml/day. During the 10 days of A-II infusion, water intake by drinking averaged 269 2 49 ml/day (IO-day mean t SE). The mean difference and its SE between the average daily water drunk while the dogs received A-II and the average daily water drunk during the three periods when A-II was not infused was 151 + 45 ml/day (paired t; P ~0.05). Figure 2 illustrates the daily water intake by drinking of the two dogs that received A-II at 26 rig/kg per min for 8 days following 2 days at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, water intake by drinking in the two dogs averaged 302 and 903 ml/day; during A-II infusion it averaged 657 and 1,705 ml/day, respectively. The average daily urine output of the four dogs that received A-II at 13 rig/kg per min during the “non-A-II” and “A-II” periods were 425 t 29 and 527 5 43 ml/day, respectively. The mean difference and its SE between these two values was 102 2 24 ml/day (paired t; P < 0.05). Similar increases in urine output were observed during A-II infusion in the two dogs that received A-II at 26 rig/kg per min.
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ANGIOTENSIN
II AND ,,,,$,&,,
1065
THIRST DISCUSSION
VEHICLE
24
Electrolyte balance and hematocrit. There were no large changes in plasma sodium and potassium concentrations or in hematocrit in the dogs that received A-II at 13 rig/kg per min. During the preinfusion period and both periods of vehicle infusion without A-II, the average values for plasma sodium concentration, plasma potassium concentration, and hematocrit were: 143 meq/liter, 4.4 meq/liter, and 38%. During A-II infusion the respective values were 142 meq/liter, 4.3 meq/liter and 36%. Urinary sodium and potassium excretion are shown in Fig. 1B and C. The average daily sodium excretion during the non-A-II and A-II periods were 40 t 2 and 38 t 3 meq/day, respectively (not significantly different,). For potassium excretion the respective values were 25 ? 2 and 27 _ + 4 meq/day (not significantly different). Similar values were observed for these variables in the two dogs that received A-II at 26 rig/kg per min.
The daily water intake by drinking in dogs was significantly elevated during prolonged intravenous infusion of angiotensin (A-II) at a rate of 13 rig/kg per min. By radioimmunoassay, B. H. Douglas (personal communication) found that intravenous infusion of A-II in dogs at dose rates comparable to 13 rig/kg per min for 4 h produced blood A-II levels in the range of 200-300 pg/ml. He also found that at the end of a 3-day sodium-depletion period in dogs, blood levels of A-II often attain concentrations of 200-300 pg/ml. Values as high as 1,000 pg/ml have been reported in sodium-depleted sheep (3). In man with renal hypertension, blood A-II levels reached 175 ,t 91 pg/ml (6). Thus, it is reasonable to state that the infusion rate of 13 rig/kg per min used in the present study produces a blood A-II concentration that is within the physiologically feasible range for dogs and that it is capable of stimulating the thirst mechanism. These results are in disagreement with the findings in sheep, which showed no increase in drinking during intravenous infusion of A-II at rates of 5-40 pg/h (2-17 rig/kg per min) for 3 days (1). Such infusion rates in sheep produce levels of A-II within the physiological limits of that species. Since sheep are ruminant animals, the discrepancy between the two studies might be related to differences in experimental animals. The two dogs that received 26 rig/kg per min of A-II showed sustained increased drinking throughout the 8 days of infusion at this rate. Although the blood levels of A-II produced by this rate of infusion may have been out of the physiological limits, the results suggest that the dipsogenic response to pharmacological doses of A-II does not show tachyphylaxis. There is a paucity of data on the dipsogenic effect of A-II in dogs; however, in one study Kozlowski et al. (14) reported that A-II infused intravenously into dogs at a rate of 50 ng/min (2-3 rig/kg per min) lowered the “thirst threshold.” That is, A-II decreased the degree of cellular dehydration (produced by 5% sodium chloride infusion)
Plasma aldosterone concentration, plasma cortisol concentration, mean arterial pressure, and plasma renin actiuity. The mean daily plasma aldosterone con-
on mean
I
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i
18
i
-
16
00” \ m
14 12
i i i
.-h
IO
s
06
E z i5
0.4
: CK
08 0 5
I
0.2
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DAYS
FIG. 2. Effect of intravenous per min for 8 days (following intake by drinking. ’
infusion 13 rig/kg
of angiotensin II at 26 rig/kg per min for 2 days) on water
centration of the dogs that received A-II at 13 nglkg per min are shown in Fig. m. Plasma aldosterone averaged 11.4 t 0.2 ng/lOO ml during the non-A-II period and 13.7 t 1.3 ng/lOO ml during the A-II period (not significantly different). Plasma cortisol concentration did not change significantly. Mean arterial pressure remained elevated 30-35 mm Hg above the control value during A-II infusion at either dose rate in all dogs in which it was measured, except dog 3 (Table 1). Likewise, in all dogs except dog 3, plasma renin activity remained suppressed during the greater part of A-II infusion. Control values for plasma renin activity in dog 11 were probably high during the control period because this animal, which weighed 21 kg, lost approximately 300 ml of blood through the arterial catheter several days before the start of the experiment. The changes in mean arterial pressure and plasma renin activity could not be shown to have statistical significance.
TABLE
DOG No. Days
1. Effect
of prolonged infusion pressure and plasma
arterial Preinfusion 2
Vehicle 4
2 Mean
2 3 5 12 12
86 87 100 101 83
94 77 100 99 95
I
99 76 98 95 92
Plasma
Angiotensin 4
2
arterial 101 88 95 93 78
133 81 130 117 113
renin
actrvitv,
II + Vehicle
4
pressure,
of angiotensin renin activity
6
II
Vehicle
8
10
2
4
134 82 144 144 134
80 130 134 128
68 99 84 74
84 99 88 91
1.12 1.29 0.67 0.16 0.91 0.51
0.48 1.15 0.62 0.24 0.86 1.20
mmHg
131 75 123 130 121 ng A-Ilml
82 133 128 116
per h
During the vehicle infusion periods, either 0.9% saline or 5% mannitol continuously by way of indwelling venous catheters and syringe pumps at either day. The dose rate of angiotensin II was 13 rig/kg per min in dogs 2,3. 5, and and 12, angiotensin II was infused at 13 rig/kg per min for 2 days, followed by min for 8 days. Mean arterial pressure was monitored for 6 h each day by way arterial catheters.
was infused 26 or 52 ml/ 7. In dogs 21 26 rig/kg per of indwelling
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TRIPPODO,
1066 necessary to induce a drinking response. A-II infused alone at this rate (without simultaneous sodium chloride infusion) did not induce the dogs to drink spontaneously. However, the infusion was carried out only for 3 h. Also, unlike the findings in sheep (l), there was no increase in urinary sodium excretion during the first 2 days of A-II infusion in the present study. In fact, there was a tendency for sodium excretion to’decrease during that time. This agrees with the results of Urquhart et al. (21), who reported sodium retention in dogs during the first 2 days of A-II infusion at 20 pg/kg per day (14 rig/kg per min). In the dogs that received A-II at 13 rig/kg per min
McCAA,
AND
GUYTON
plasma aldosterone concentration was only slightly above the control value after the third day of A-II infusion. McCaa et al. (16) demonstrated that continuous infusion of A-II at 5 rig/kg per min for 14 days produced a significant increase in plasma aldosterone concentration in dogs only during the first day of infusion. The authors are grateful to Terry S. Finch, Florence N. son, Michael O’Ryan McGee, and Mary Frances Plummer excellent technical assistance. This work was supported by NIH grants HL 11678,09921, 00316. N. C. Trippodo was an American and Mississippi Heart tion Fellow during the time of this study. Received
for publication
30 June
Hutchifor their and GM Associa-
1975.
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