Thermal dehydration-induced role of angiotensin II CHRISTOPHER Department
C. BARNEY,
thirst in rats:
JEFFREY
S. WILLIAMS,
AND
DAVID
H. KUIPER
of Biology, Hope College, Holland, Michigan 49423
BARNEY, CHRISTOPHER C., JEFFREY S. WILLIAMS, AND DAVID H. KUIPER. Thermal dehydration-induced thirst in rats: role of angiotensin II. Am. J. Physiol. 261 (Regulatory Integra-
tive Comp. Physiol. 30): Rll71-R1175, 1991.-Dehydration can be brought about by either water deprivation or by heat exposure(thermal dehydration). Angiotensin II hasbeenshown to have a role in water deprivation-induced thirst. The current study was designedto determine whether angiotensin II is involved in thirst causedby thermal dehydration. Male Sprague-Dawley strain rats were dehydrated by exposure to a 40°C environment for 2-4 h or by water deprivation for 44 h. Water deprivation but not heat exposure significantly increased plasma renin activity. Neither ureteric ligation nor nephrectomy significantly altered water intake after thermal dehydration. Captopril, an inhibitor of angiotensin converting enzyme, given at a doseof 100 mg/kg ip, significantly decreasedwater intake in water-deprived rats but not in thermally dehydrated rats. Angiotensin II therefore doesnot appear to play a role in the control of water intake of thermally dehydrated rats. The physiological responsesto dehydration in rats are dependent on the way in which the dehydration is brought about. captopril; heat stress; nephrectomy; plasma renin activity; water balance;water deprivation
of Du Pont 753, a specific angiotensin II type I receptor antagonist (35). Although angiotensin II may have a physiological role in stimulating water intake in rats deprived of water for extended periods, the role of angiotensin II in thermal dehydration-induced thirst has not been previously elucidated. Thermal dehydration of rats is associated with loss of water and electrolytes in the saliva used for evaporative cooling (16, 17, 30); decreases in total body water, extracellular fluid volume and plasma volume (7, 18, 27, 30); and thirst (4, 17, 28-30). Thermal dehydration differs from dehydration caused by water deprivation in that there are significant increases in body temperature (16) and salivary electrolyte loss (30) and a smaller but still significant decrease in plasma volume during thermal dehydration (7, 18). Thirst after thermal dehydration in rats has a smaller volemic component than water deprivation-induced thirst (4, 27, 28, 32). Thus, it is less likely that the renin-angiotensin system is involved in thermal dehydration-induced thirst. The studies reported here confirm this view. METHODS
IS ONE OF MANY stimuli that can cause thirst and an increase in water intake. Dehydration can be brought about by water deprivation or by exposure to hot environments. During exposure to heat, water is lost for evaporative cooling, leading to thermal dehydration. Water deprivation-induced water intake has been studied extensively (see Refs. 12 and 34 for reviews). In rats, the animal most used for thirst studies, water deprivation stimulates thirst by causing both a cellular water deficit and a reduction in plasma volume (5, 32). A reduction in plasma volume appears to induce thirst via either activation of the renin-angiotensin system or stimulation of venous and atria1 baroreceptors (5, 32). Although the role of angiotensin II in physiological thirst is controversial (24,33), several studies with rats provide support for angiotensin II’s involvement in water deprivation-induced thirst. Water deprivation increases plasma renin activity and plasma angiotensin II concentrations (3, 6, 22,23). In addition, captopril, an inhibitor of angiotensin converting enzyme (ACE) that has been shown to specifically attenuate thirst observed after activation of the renin-angiotensin system (2, 9, 10, 13), decreases water intake due to extended water deprivation (2, 3). Water deprivation-induced thirst in rats is also significantly attenuated after intracerebroventricular administration DEHYDRATION
0363-6119/91
$1.50
One hundred twenty-four male Sprague-Dawley strain rats (Harlan Sprague Dawley) weighing from 290 to 560 g were used in these experiments. In any experiment, there were no significant differences in pretreatment body weights between groups. Rats were housed individually in an animal room kept at 24 t 2°C in hanging stainless steel cages. The animal room was illuminated from 7:00 A.M. to 7:00 P.M. Rats were allowed Purina Rat Chow and water ad libitum, except during the experiments. The experimental protocol was approved by the Hope College Animal Care and Use Committee. Two types of thirst stimuli were used in these studies, water deprivation and thermal dehydration. In previous studies, we have examined the body water status and thirst responses of rats to varying levels of water deprivation (3) and thermal dehydration (4). The durations of water deprivation and thermal dehydration used in the current experiments were chosen to be those that had caused similar and significant increases in plasma sodium concentration, plasma osmolality, and water intake in the previous studies. Because angiotensin II appears to be significantly involved in the thirst response only at high levels of water deprivation-induced dehydration, the current studies of thermal dehydration were limited to similar high levels (6-8% of body wt).
Copyright 0 1991 the American Physiological Society
R1171
Downloaded from www.physiology.org/journal/ajpregu by ${individualUser.givenNames} ${individualUser.surname} (132.174.254.155) on December 2, 2018. Copyright © 1991 the American Physiological Society. All rights reserved.
R1172
ANG
II AND
THERMAL
DEHYDRATION-INDUCED
THIRST
Six groups of rats were used in the first experiment. ber as in experiment 1. After being placed in the heat Each rat was weighed and placed in a modified Nalgene chamber (3 h and 15 min later), nine rats were injected metabolism cage (4) at the beginning of the experiment. with captopril (100 mg/kg ip), and eight rats were inThe experiment began between 7:30 and 8:30 A.M. for jected with 0.9% NaCl solution (2 ml/kg ip). Previous the control and heat-exposed rats and at 6:30 P.M. for studies (2,9, 10) have shown that high doses of captopril the water-deprived rats. At the end of the experiment, (50-100 mg/kg) are needed if inhibition of both brain each rat was reweighed, and a 3-ml blood sample was and peripheral ACE activity is desired. Because angioobtained by decapitating the rat. Blood was collected in tensin II generated in either the periphery or the brain chilled centrifuge tubes containing 3 mg of sodiummight play a role in thermal dehydration-induced thirst EDTA. The blood was centrifuged at 12,000 g for 30 min the 100 mg/kg dose was chosen for these studies. The at 4OC. The plasma was removed and stored at -80°C. concentration of captopril was 50 mg/ml, and the saline Plasma renin activity (PRA) was determined in duplicate was adjusted to the same pH (2.3) as the captopril with using a lz51-labeled angiotensin I radioimmunoassay kit HCl. Forty-five minutes after injections the rats were reweighed and moved to clean metabolism cages in an (Rianen, New England Nuclear). Intra-assay variability of duplicate samples was 9.1%. environmental chamber kept at 25°C. Water was given Groups 1 through 4 did not have access to food or to each rat 15 min later, and water intake was measured water throughout the experimental period. Group 1 rats as in experiment 2. Cumulative water intake of the two (control, n = 7) were kept at 25 t 05°C for 4 h and 15 groups at each time interval was analyzed by Student’s t min before blood sampling. Group 2 (n = 8) was kept at test (11). 25°C for 2 h and was then moved to a walk-in environIn the fourth experiment the effect of captopril on mental chamber kept at 40 t 0.5”C for an additional 2 water intake of rats deprived of water for 44 h was h. Blood was sampled 15 min after removal from the heat studied. Rats were weighed and returned to their home chamber (4). Group 3 (n = 8) was kept in the heat cages in the animal room. Water but not food was removed at 6:30 P.M. Forty-three hours later the rats were chamber for 4 h, and blood was sampled 15 min after removal from the heat chamber. Group 4 (n = 7) was reweighed, injected with either captopril (n = 9) or saline kept in the heat chamber for 4 h and 15 min, and the (n = 9) as in experiment 3, and placed in metabolism blood samples were taken while the rats remained in the cages at 25°C. One hour later the rats were given water, heat chamber. Group 5 (n = 7) was kept without food or and water intake was measured and analyzed as in exwater in the heat chamber for 4 h and was then removed periment 3. to clean metabolism cages in an environmental chamber kept at 25 t 0.5OC. Fifteen minutes after removal from RESULTS the heat the rats in group 5 were allowed access to 25°C The rats in experiment 1 lost from 1.9% (control rats) water for l-h. At the end of the l-h water access period, to 10.0% (44 h water-deprived rats) of their body weight blood samples were taken. Water intake was measured during the experimental period. The rats exposed to the by the weight difference in the water bottle. Group 6 (n = 7) was kept in the animal room with access to food heat for 4 h lost 7.6% of their body weight, which was but not water for 44 h before blood sampling. The data significantly more than the 5.5% of body weight lost by the rats exposed to the heat for 2 h. The rats that were were analyzed by one-way analysis of variance (ANOVA) with significance set at the 95% confidence level (11). allowed accessto water for 1 h drank 18.3 t 2.4 (SE) ml/ Individual comparisons were made by the post hoc New- kg. Figure 1 shows the PRA data from experiment 1. man-Keuls test with significance also set at the 95% One-way ANOVA indicated a significant (P < 0.001) treatment effect on PRA. However, only the group deconfidence level (11). prived of water for 44 h had a PRA value significantly In the second experiment, the effects of nephrectomy or ureteric ligation on water intake of thermally dehy- different from any other group. Heat exposure for 2 or 4 h did not alter PRA. PRA was higher in the rats that drated rats were studied. Rats were anesthetized with had blood samples taken in the heat and was lower after methoxyflurane and were bilaterally nephrectomized, had both ureters ligated, or underwent sham surgery TX7 CN I 2 HR 40°C using the procedures of Elfont and Fitzsimons (8). Fifb 4 HR 40°C teen rats were used in each group. Two hours after the /?GT2!l4 HR 4c>“C, SAMPLED AT 40°C EII 4 HR 40°C, 1 HR ACC ESS TO HZ0 end of surgery, at between 10 A.M. and noon, the rats AT 25°C \’ \ I were placed in the metabolism cages at 4O"C, as in the ,” 15 I/l 44 HR WD CI first experiment. After 3 h of heat exposure the rats were removed to clean metabolism cages in an environmental E 3 IO chamber kept at 25OC and were given water to drink 15 min later. Water intake was measured after 0.5, 1, and 2 3 75 h. At the end of the experiment, each rat was killed by 7 overanesthetization with pentobarbital sodium. Cumu& lative water intake at each time interval was analyzed by 0 one-way ANOVA. FIG. 1. Mean plasma renin activity of control (CN) rats, rats exThe effect of inhibition of ACE on thermal dehydraposed to heat for 2 or 4 h, and rats deprived of water (WD) for 44 h. tion-induced thirst was studied in the third experiment. Bars indicate mean k 1 SE for 7 or 8 rats/group. Mean values not Rats were placed in metabolism cages in the heat cham- sharing same superscript are significantly different at P < 0.05.
Downloaded from www.physiology.org/journal/ajpregu by ${individualUser.givenNames} ${individualUser.surname} (132.174.254.155) on December 2, 2018. Copyright © 1991 the American Physiological Society. All rights reserved.
ANG
II AND
THERMAL
DEHYDRATION-INDUCED
1 h of access to water, but these differences were not statistically significant. Water intake of the sham, ureteric-ligated, and nephrectomized rats after 3 h of heat exposure is shown in Fig. 2. Although each group of rats drank a considerable amount of water indicative of thermal dehydration, there was no significant treatment effect on cumulative water intake at any time. The effects of captopril on water intake of thermally dehydrated and water-deprived rats are shown in Figs. 3 and 4, respectively. Rats exposed to the heat for 4 h showed an 8.2% decrease in body weight with no significant difference between the two groups. Captopril had no significant effect on cumulative water intake during the 1st h of access to water and significantly (P < 0.05) increased the 2-h cumulative water intake. Rats deprived of water for 44 h lost an average of 10.2% of their body weight with no significant difference between the two groups. Captopril significantly (P < 0.05) reduced cumulative water intake induced by water deprivation at each measurement interval. DISCUSSION
Angiotensin II has been shown to be a potent dipsogenie agent and to be involved in the thirst response to 35
1
‘;;;
30
s A w Y 5 -z
25-
0 LKJ
SHAM URETERIC
EB8
NEPHRECTOMIZED
LIGATED T
zo15-
E5 IO50 1
0.5
HOURS FIG. ligated, indicate
2
OF ACCESS
TO WATER
2. Mean cumulative water inta .ke of sham-operated, ure terican .d nephrectomized rats after 3 h of exposure to 40°C. Bars mean t 1 SE for 15 rats/group.
551 G
50
C. BARNEY,
thirst in rats:
JEFFREY
S. WILLIAMS,
AND
DAVID
H. KUIPER
of Biology, Hope College, Holland, Michigan 49423
BARNEY, CHRISTOPHER C., JEFFREY S. WILLIAMS, AND DAVID H. KUIPER. Thermal dehydration-induced thirst in rats: role of angiotensin II. Am. J. Physiol. 261 (Regulatory Integra-
tive Comp. Physiol. 30): Rll71-R1175, 1991.-Dehydration can be brought about by either water deprivation or by heat exposure(thermal dehydration). Angiotensin II hasbeenshown to have a role in water deprivation-induced thirst. The current study was designedto determine whether angiotensin II is involved in thirst causedby thermal dehydration. Male Sprague-Dawley strain rats were dehydrated by exposure to a 40°C environment for 2-4 h or by water deprivation for 44 h. Water deprivation but not heat exposure significantly increased plasma renin activity. Neither ureteric ligation nor nephrectomy significantly altered water intake after thermal dehydration. Captopril, an inhibitor of angiotensin converting enzyme, given at a doseof 100 mg/kg ip, significantly decreasedwater intake in water-deprived rats but not in thermally dehydrated rats. Angiotensin II therefore doesnot appear to play a role in the control of water intake of thermally dehydrated rats. The physiological responsesto dehydration in rats are dependent on the way in which the dehydration is brought about. captopril; heat stress; nephrectomy; plasma renin activity; water balance;water deprivation
of Du Pont 753, a specific angiotensin II type I receptor antagonist (35). Although angiotensin II may have a physiological role in stimulating water intake in rats deprived of water for extended periods, the role of angiotensin II in thermal dehydration-induced thirst has not been previously elucidated. Thermal dehydration of rats is associated with loss of water and electrolytes in the saliva used for evaporative cooling (16, 17, 30); decreases in total body water, extracellular fluid volume and plasma volume (7, 18, 27, 30); and thirst (4, 17, 28-30). Thermal dehydration differs from dehydration caused by water deprivation in that there are significant increases in body temperature (16) and salivary electrolyte loss (30) and a smaller but still significant decrease in plasma volume during thermal dehydration (7, 18). Thirst after thermal dehydration in rats has a smaller volemic component than water deprivation-induced thirst (4, 27, 28, 32). Thus, it is less likely that the renin-angiotensin system is involved in thermal dehydration-induced thirst. The studies reported here confirm this view. METHODS
IS ONE OF MANY stimuli that can cause thirst and an increase in water intake. Dehydration can be brought about by water deprivation or by exposure to hot environments. During exposure to heat, water is lost for evaporative cooling, leading to thermal dehydration. Water deprivation-induced water intake has been studied extensively (see Refs. 12 and 34 for reviews). In rats, the animal most used for thirst studies, water deprivation stimulates thirst by causing both a cellular water deficit and a reduction in plasma volume (5, 32). A reduction in plasma volume appears to induce thirst via either activation of the renin-angiotensin system or stimulation of venous and atria1 baroreceptors (5, 32). Although the role of angiotensin II in physiological thirst is controversial (24,33), several studies with rats provide support for angiotensin II’s involvement in water deprivation-induced thirst. Water deprivation increases plasma renin activity and plasma angiotensin II concentrations (3, 6, 22,23). In addition, captopril, an inhibitor of angiotensin converting enzyme (ACE) that has been shown to specifically attenuate thirst observed after activation of the renin-angiotensin system (2, 9, 10, 13), decreases water intake due to extended water deprivation (2, 3). Water deprivation-induced thirst in rats is also significantly attenuated after intracerebroventricular administration DEHYDRATION
0363-6119/91
$1.50
One hundred twenty-four male Sprague-Dawley strain rats (Harlan Sprague Dawley) weighing from 290 to 560 g were used in these experiments. In any experiment, there were no significant differences in pretreatment body weights between groups. Rats were housed individually in an animal room kept at 24 t 2°C in hanging stainless steel cages. The animal room was illuminated from 7:00 A.M. to 7:00 P.M. Rats were allowed Purina Rat Chow and water ad libitum, except during the experiments. The experimental protocol was approved by the Hope College Animal Care and Use Committee. Two types of thirst stimuli were used in these studies, water deprivation and thermal dehydration. In previous studies, we have examined the body water status and thirst responses of rats to varying levels of water deprivation (3) and thermal dehydration (4). The durations of water deprivation and thermal dehydration used in the current experiments were chosen to be those that had caused similar and significant increases in plasma sodium concentration, plasma osmolality, and water intake in the previous studies. Because angiotensin II appears to be significantly involved in the thirst response only at high levels of water deprivation-induced dehydration, the current studies of thermal dehydration were limited to similar high levels (6-8% of body wt).
Copyright 0 1991 the American Physiological Society
R1171
Downloaded from www.physiology.org/journal/ajpregu by ${individualUser.givenNames} ${individualUser.surname} (132.174.254.155) on December 2, 2018. Copyright © 1991 the American Physiological Society. All rights reserved.
R1172
ANG
II AND
THERMAL
DEHYDRATION-INDUCED
THIRST
Six groups of rats were used in the first experiment. ber as in experiment 1. After being placed in the heat Each rat was weighed and placed in a modified Nalgene chamber (3 h and 15 min later), nine rats were injected metabolism cage (4) at the beginning of the experiment. with captopril (100 mg/kg ip), and eight rats were inThe experiment began between 7:30 and 8:30 A.M. for jected with 0.9% NaCl solution (2 ml/kg ip). Previous the control and heat-exposed rats and at 6:30 P.M. for studies (2,9, 10) have shown that high doses of captopril the water-deprived rats. At the end of the experiment, (50-100 mg/kg) are needed if inhibition of both brain each rat was reweighed, and a 3-ml blood sample was and peripheral ACE activity is desired. Because angioobtained by decapitating the rat. Blood was collected in tensin II generated in either the periphery or the brain chilled centrifuge tubes containing 3 mg of sodiummight play a role in thermal dehydration-induced thirst EDTA. The blood was centrifuged at 12,000 g for 30 min the 100 mg/kg dose was chosen for these studies. The at 4OC. The plasma was removed and stored at -80°C. concentration of captopril was 50 mg/ml, and the saline Plasma renin activity (PRA) was determined in duplicate was adjusted to the same pH (2.3) as the captopril with using a lz51-labeled angiotensin I radioimmunoassay kit HCl. Forty-five minutes after injections the rats were reweighed and moved to clean metabolism cages in an (Rianen, New England Nuclear). Intra-assay variability of duplicate samples was 9.1%. environmental chamber kept at 25°C. Water was given Groups 1 through 4 did not have access to food or to each rat 15 min later, and water intake was measured water throughout the experimental period. Group 1 rats as in experiment 2. Cumulative water intake of the two (control, n = 7) were kept at 25 t 05°C for 4 h and 15 groups at each time interval was analyzed by Student’s t min before blood sampling. Group 2 (n = 8) was kept at test (11). 25°C for 2 h and was then moved to a walk-in environIn the fourth experiment the effect of captopril on mental chamber kept at 40 t 0.5”C for an additional 2 water intake of rats deprived of water for 44 h was h. Blood was sampled 15 min after removal from the heat studied. Rats were weighed and returned to their home chamber (4). Group 3 (n = 8) was kept in the heat cages in the animal room. Water but not food was removed at 6:30 P.M. Forty-three hours later the rats were chamber for 4 h, and blood was sampled 15 min after removal from the heat chamber. Group 4 (n = 7) was reweighed, injected with either captopril (n = 9) or saline kept in the heat chamber for 4 h and 15 min, and the (n = 9) as in experiment 3, and placed in metabolism blood samples were taken while the rats remained in the cages at 25°C. One hour later the rats were given water, heat chamber. Group 5 (n = 7) was kept without food or and water intake was measured and analyzed as in exwater in the heat chamber for 4 h and was then removed periment 3. to clean metabolism cages in an environmental chamber kept at 25 t 0.5OC. Fifteen minutes after removal from RESULTS the heat the rats in group 5 were allowed access to 25°C The rats in experiment 1 lost from 1.9% (control rats) water for l-h. At the end of the l-h water access period, to 10.0% (44 h water-deprived rats) of their body weight blood samples were taken. Water intake was measured during the experimental period. The rats exposed to the by the weight difference in the water bottle. Group 6 (n = 7) was kept in the animal room with access to food heat for 4 h lost 7.6% of their body weight, which was but not water for 44 h before blood sampling. The data significantly more than the 5.5% of body weight lost by the rats exposed to the heat for 2 h. The rats that were were analyzed by one-way analysis of variance (ANOVA) with significance set at the 95% confidence level (11). allowed accessto water for 1 h drank 18.3 t 2.4 (SE) ml/ Individual comparisons were made by the post hoc New- kg. Figure 1 shows the PRA data from experiment 1. man-Keuls test with significance also set at the 95% One-way ANOVA indicated a significant (P < 0.001) treatment effect on PRA. However, only the group deconfidence level (11). prived of water for 44 h had a PRA value significantly In the second experiment, the effects of nephrectomy or ureteric ligation on water intake of thermally dehy- different from any other group. Heat exposure for 2 or 4 h did not alter PRA. PRA was higher in the rats that drated rats were studied. Rats were anesthetized with had blood samples taken in the heat and was lower after methoxyflurane and were bilaterally nephrectomized, had both ureters ligated, or underwent sham surgery TX7 CN I 2 HR 40°C using the procedures of Elfont and Fitzsimons (8). Fifb 4 HR 40°C teen rats were used in each group. Two hours after the /?GT2!l4 HR 4c>“C, SAMPLED AT 40°C EII 4 HR 40°C, 1 HR ACC ESS TO HZ0 end of surgery, at between 10 A.M. and noon, the rats AT 25°C \’ \ I were placed in the metabolism cages at 4O"C, as in the ,” 15 I/l 44 HR WD CI first experiment. After 3 h of heat exposure the rats were removed to clean metabolism cages in an environmental E 3 IO chamber kept at 25OC and were given water to drink 15 min later. Water intake was measured after 0.5, 1, and 2 3 75 h. At the end of the experiment, each rat was killed by 7 overanesthetization with pentobarbital sodium. Cumu& lative water intake at each time interval was analyzed by 0 one-way ANOVA. FIG. 1. Mean plasma renin activity of control (CN) rats, rats exThe effect of inhibition of ACE on thermal dehydraposed to heat for 2 or 4 h, and rats deprived of water (WD) for 44 h. tion-induced thirst was studied in the third experiment. Bars indicate mean k 1 SE for 7 or 8 rats/group. Mean values not Rats were placed in metabolism cages in the heat cham- sharing same superscript are significantly different at P < 0.05.
Downloaded from www.physiology.org/journal/ajpregu by ${individualUser.givenNames} ${individualUser.surname} (132.174.254.155) on December 2, 2018. Copyright © 1991 the American Physiological Society. All rights reserved.
ANG
II AND
THERMAL
DEHYDRATION-INDUCED
1 h of access to water, but these differences were not statistically significant. Water intake of the sham, ureteric-ligated, and nephrectomized rats after 3 h of heat exposure is shown in Fig. 2. Although each group of rats drank a considerable amount of water indicative of thermal dehydration, there was no significant treatment effect on cumulative water intake at any time. The effects of captopril on water intake of thermally dehydrated and water-deprived rats are shown in Figs. 3 and 4, respectively. Rats exposed to the heat for 4 h showed an 8.2% decrease in body weight with no significant difference between the two groups. Captopril had no significant effect on cumulative water intake during the 1st h of access to water and significantly (P < 0.05) increased the 2-h cumulative water intake. Rats deprived of water for 44 h lost an average of 10.2% of their body weight with no significant difference between the two groups. Captopril significantly (P < 0.05) reduced cumulative water intake induced by water deprivation at each measurement interval. DISCUSSION
Angiotensin II has been shown to be a potent dipsogenie agent and to be involved in the thirst response to 35
1
‘;;;
30
s A w Y 5 -z
25-
0 LKJ
SHAM URETERIC
EB8
NEPHRECTOMIZED
LIGATED T
zo15-
E5 IO50 1
0.5
HOURS FIG. ligated, indicate
2
OF ACCESS
TO WATER
2. Mean cumulative water inta .ke of sham-operated, ure terican .d nephrectomized rats after 3 h of exposure to 40°C. Bars mean t 1 SE for 15 rats/group.
551 G
50
