Suppression of Plasma Renin and Plasma Aldosterone During Water Immersion in Normal Man MURRAY EPSTEIN,* DAVID S. PINS, JOSE SANCHO, AND EDGAR HABER Medical Service, Veterans Administration Hospital and Department of Medicine, University of Miami School of Medicine, Miami, Florida 33125; and the Cardiac Unit, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114 ABSTRACT. Previous studies from this laboratory have demonstrated that the redistribution of blood volume and concomitant relative central hypervolemia induced by water immersion to the neck (NI) results in a significant natriuresis which is quantitatively identical to that induced by the acute administration of 2 liters of saline. Since the central hypervolemia induced by NI occurs without concomitant alterations in serum sodium and potassium concentration, the NI model was utilized to assess the role of volume in the regulation of both plasma renin (PRA) and plasma aldosterone (PA) in man. Nine normal subjects were studied on two occasions while in balance on a 10 meq Na, 100 meq K diet: Control and NI. The conditions of seated posture and time of day were identical. Blood for PRA and PA was obtained at 30-min intervals for 6 h. NI produced a profound suppression of PRA as early as
P
REVIOUS studies from this laboratory have demonstrated that the redistribution of blood volume and concomitant central hypervolemia that accompany headout water immersion to the neck result in a significant and reproducible natriuresis and a suppression of the renin-angiotensin-aldosterone system (1-3). Since changes in aldosterone secretion were determined by measurements of aldosterone excretion (1) rather than plasma aldosterone, the timecourse of the aldosterone alterations in response to immersion and their relation to concomitant changes in PRA could not be elucidated (1). Furthermore, although a 70% decrease in plasma renin activity was documented in these studies (1), determination of PRA at two-hourly intervals precluded assessment of the rapidity of changes resulting from initiation or discontinuation of immersion. Received March 31, 1975. * Dr. Epstein is an Investigator of the Howard Hughes Medical Institute.
30 min with maximal suppression (62%) by 180 min (P < 0.001). Recovery from NI was associated with a prompt return to pre-study levels. The changes in PA paralleled those of PRA with regard to both the rapidity and magnitude of the suppression (r = 0.993: P < 0.001). These data emphasize the importance of central volume per se as a primary determinant of PRA and PA regulation in normal man. Furthermore, the current studies confirm the importance of the renin-angiotensin axis in the control of volumerelated changes in PA in normal man. The ability of NI to induce a prompt and parallel suppression of PRA and PA without concomitant alterations in plasma composition, suggests that NI may be a preferred investigative tool for assessing the effects of volume expansion on renin-aldosterone. (/ Clin Endocrinol Metab 41: 618, 1975)
The development of a sensitive and specific radioimmunoassay for plasma aldosterone (4) has made possible the assessment, by direct measurement, of the effect of water immersion on aldosterone. The current study was undertaken to characterize the temporal profile of the suppression of PRA and plasma aldosterone during immersion and to elucidate further the role of aldosterone suppression in mediating the natriuresis of water immersion. Materials and Methods Nine men between the ages of 19 and 25 y were studied. There was no prior history of hypertension, cardiovascular disease or diabetes. Significant renal disease was excluded by documenting a normal urinary sediment, creatinine clearance and sterile urine cultures. The subjects were housed during the study in an environmentally controlled metabolic ward at a constant temperature. Each consumed a diet, the composition of which remained unchanged throughout the study, containing 10 meq of sodium, 100 meq of potassium and 2,500 ml of water. Daily 24-h
618
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION urine collections were made for determination of sodium, potassium and creatinine. After dietary equilibration, each subject underwent a Control study on day 5, followed on days 7-8 by an Immersion study. The experimental protocols on the 2 study days were similar and were carried out as follows: After 14 h of overnight water deprivation, the subject was awakened at 0700, voided and sat quietly for 1 h. At 0800, an oral water load of 700 ml was administered and at 0815 the subject voided again. Immediately after voiding, a 20 gauge lVfe" Teflon catheter with a flash chamber and an accompanying Teflon stylet (Jelco Co., Raritan, N.J.) was inserted into a forearm vein, permitting sequential sampling of blood without the use of an anti-coagulant or an intravenous infusion. Immediately after insertion, venous blood was drawn using a pre-chilled 10 ml polystyrene disposable syringe and an aliquot was collected for renin and aldosterone measurements in a pre-chilled vacutainer tube containing EDTA. After obtaining the sample, a sterile stylet was inserted into the catheter. Blood samples were obtained before and after the study for sodium, potassium, creatinine and osmolality determinations. During the control study, the subject sat quietly outside the immersion tank for the 6-h period beginning with the 0815 voiding. During the control study, the subject sat quietly outside the immersion tank for the 6-h period beginning with the 0815 voiding. During the immersion study, the subject sat in the tank immersed in water to the neck for 4 h (915-1315), preceded and followed by 1 h of quiet sitting outside the tank (Pre-Study and Recovery hours, respectively). Each subject stood briefly to void spontaneously at hourly intervals during the study. To maintain adequate urine flow, 200 ml of water was administered orally every hour during the study. Sodium, potassium, creatinine and osmolality were measured in aliquots of the hourly urine collections. Blood was collected at 30-min intervals for renin and aldosterone determinations. All subjects were weighed every morning at 0700 after voiding and before and after each study. Immersion was carried out in a waterproof tank described in detail in previous communications (2,3). A constant water temperature of 34 ± 0.5 C was maintained by two heat exchangers, controlled by an adjustable temperature-calibrated
619
control meter with input derived from two thermistors immersed at different water levels. Plasma renin was measured by radioimmunoassay according to the method of Haber et al. (5). Plasma aldosterone was measured by a modification of the radioimmunoassay technique of Poulsen et al. (4). The modification utilizes a new antibody with a very high affinity constant (10" n M/liter), so that 50% displacement is obtained with only 8 pg of aldosterone (Sancho and Haber, to be published). With this modification, aldosterone could be assayed utilizing only 0.1 ml of plasma. The coefficient of variation for interassay determinations is 10.2%. Analytic methods for sodium, potassium and creatinine determinations have been reported previously (2,3). In the presentation of the data, mean values are followed by the standard error of the mean as an index of dispersion. Data were evaluated statistically using Student's t test for paired values or, where appropriate by the Wilcoxon signed rank test. Differences with P < 0.05 were considered significant. Permission for the study was obtained from each subject after a detailed description of the procedure and the potential complications. The protocol was approved by the Human Experimentation Committees of the University of Miami School of Medicine and the Miami Veterans Administration Hospital and was in compliance with the principles outlined in the Declaration of Helsinki. No complications occurred.
Results Urinary electrolyte excretion The effect of 4 h of water immersion on sodium and potassium excretion is shown in Table 1. During quiet sitting (Control), the rate of sodium excretion (UNa V) was constant, ranging from 3 to 6 /aeq/min. Immersion resulted in a significant increase in UNaV compared to Control beginning with the second hour of Immersion. By hour 4, UNaV was 10-fold greater than during the comparable Control period. Recovery was associated with a decline in UNaV, but UNaV continued to exceed both preimmersion values and the values during the comparable Control hour. The rate of potassium excretion (UKV)
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TABLE 1. Effect of immersion on urinaiy excretory patterns (results are mean ± SE of 9 subjects) Time (h) Group
1
Pre-study
Control Immersion
3.5 ± 0.8 3.1 ± 0.5
(jxeq/min)
Control Immersion
5.1 ± 1.5 2.5 ± 0.7
UKV (/neq/min)
Control Immersion
Ccr
Control Immersion
ml/vmin
u Na v
(ml/min)
51 49
± 12 ± 11
4
4.0 ± 0.4 7.6 ± 0.6*
3.7 ± 0.6 5.6 ± 0.6
3.8 ± 0.8 4.3 ± 0.5
2.5 ± 0.3 1.8 ± 0.5
6.0 ± 2.1 28.6 ± 10.3*
5.3 ± 2.5 39.5 ± 15.9*
5.6 ± 2.5 55.6 ± 19.9*
3.0 ± 0.7 38.2 ± 16.8*
2.6 ± 0.5 5.8 ± 0.9* 5.5 ± 2.4 13.4 ± 3.8 ± 11 ± 9*
58 78
120 ± 8 114 ± 5
3
2
121 ± 5 133 ± 8
60 ± 7 111 ± 8*
55 ± 9 106 ± 10*
118 ± 5 121 ± 4
109 122
± 6 ± 5
61 87
± 7 ± 8
111 ± 6 114 ± 8
Recovery
55 50
± 3 ± 7
108 ± 6 118 ± 8
* P < 0.05 as compared with Control.
was significantly increased during each of the initial 3 h of Immersion. Creatinine clearance (CCr) during Control was not significantly different from the Control pre-study hour, ranging from 108 to
121 ml/min. Immersion did not significantly alter CCr. Positive fluid balance (water intake minus urine volume) during Control of 253 ± 75 ml was observed. In contrast, fluid
IMMERSION
12 r-
10 L-'TN PRA ng/ml/hr
•i
8
N.S.
\ . . .
CONTROL
—
IMMERSION
N.S.
; \
(n = 9) MEAN + S.E. r
p < 0.001
>
p < 0.005
0
60
120
180
FlG. 1. Effect of water immersion to the neck on plasma renin activity (PRA) in subjects in balance on a 10 meq sodium diet. Immersion resulted in a progressive decrement in PRA beginning as early as 30 min. Cessation of immersion (Recovery) was associated with a prompt return of PRA to prestudy values.
240 30 60
TIME (minutes)
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION
621
IMMERSION (n = 9) MEAN 1 S.E.
500 FIG. 2. Effect of water immersion to the neck on plasma aldosterone in subjects in balance on a 10 meq sodium diet. Immersion resulted in a progressive decrement in plasma aldosterone beginning as early as 60 min. Cessation of immersion (Recovery) was associated with a prompt return of plasma aldosterone to pre-study values.
PLASMA
r
y?\
400
ALDOSTERONE LEVELS
300
N.S.
- —
pg/ml
N.s\ 200
100
CONTROL
— — IMMERSION \
IXl * p < 0.001 0 p < 0.005 I
I
0
60
120
180
240 30 60
TIME (minutes)
balance was negative by 299 ± 64 ml during Immersion, significantly different from Control (P < 0.001).
was associated with significant suppression of aldosterone beginning as early as 30 min. By 210 min of Immersion, aldosterone was maximally suppressed to 34% of the prePlasma renin activity (PRA) study value. Recovery was associated with a prompt return to pre-study values. As shown in Fig. 1, plasma renin activity was appropriately elevated in re- Relationship between PRA and plasma sponse to dietary sodium restriction through- aldosterone out the Control study and in the pre-study As shown in Fig. 3, the suppression of hour of Immersion. Immersion resulted in PRA and PA in response to immersion and a significant suppression of PRA beginning their subsequent recovery following cessawithin 30 min of study. By 180 min, PRA tion of immersion occurred in parallel was suppressed to 38% of the pre-study (r = 0.993; P < 0.001). value. Cessation of Immersion was associated with a prompt return toward pre- Serum electrolytes study values as early as 30 min of Recovery; Serum sodium (137 ± 1 meq/liter) and osby 60 min of Recovery, PRA was not difmolality (282 ± 1 mOsm/liter) were identiferent from the pre-study value. cal during the Control and Immersion studies. Similarly, serum potassium concenChanges in plasma aldosterone (PA) tration was similar during the two studies As shown in Fig. 2, plasma aldosterone and did not change significantly during the levels were also appropriately elevated control and immersion studies when comthroughout the Control study and during the pared with values obtained immediately pre-study hour of Immersion. Immersion prior to each study.
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EPSTEIN, PINS, SANCHO AND HABER
Discussion Water immersion to the neck has previously been shown to produce a profound natriuresis and a suppression of the reninaldosterone system (1-3,6). The nature of prior studies did not permit a kinetic examination of changes in plasma renin and aldosterone in relation to onset and offset of immersion nor their precise correlation with renal sodium and potassium excretion. The present study demonstrates that immersion induces a rapid (30 min) and profound suppression of both PRA and plasma aldosterone with an equally rapid recovery following discontinuation of immersion. The dissociation between plasma aldosterone and renal sodium handling during the recovery hour merits comment. During recovery, at a time when both PRA and aldosterone levels had returned to prestudy levels, the rate of sodium excretion continued to exceed both the pre-immersion value and the comparable control values.
JCE & M • 1975 VoUl • No 3
The prompt return of PRA and plasma aldosterone to pre-study levels suggests that cessation of immersion was associated with normalization ofblood volume distribution which was sensed by the appropriate volume receptors. Despite this apparent normalization of the distribution of circulating blood volume, the natriuresis persisted suggesting that a humoral factor rather than more rapidly acting hemodynamic and neural mechanisms mediated the natriuresis of immersion (7,8). The current demonstration of a progressive kaliuresis during immersion helps to elucidate further the mechanism of the natriuresis of immersion. In view of the concomitant decrease in plasma aldosterone concentration, one would have anticipated that potassium excretion would have diminished. The current demonstration of increased K excretion is consistent, however, with previous studies from this laboratory suggesting that the natriuresis of immer-
100 FIG. 3. Comparison of the effects of immersion on plasma renin activity (PRA) and plasma aldosterone in subjects in balance on a 10 meq Na diet. Data are expressed in terms of percent change from the pre-immersion hour. As can be seen, the suppression of plasma aldosterone paralleled the suppression of PRA throughout the immersion period. Similarly, following cessation of immersion, the recovery of both PRA and plasma aldosterone occurred in parallel.
80 0/
/o
DECREASE FROM
6 0
PRE-STUDY
= 0.993
40
20
0
0
•—•
PLASMA RENIN
o—o
PLASMA
60
ACTIVITY
ALDOSTERONE
120
180
240
60
TIME (minutes)
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RENIN-ALDOSTERONE SUPPRESSION BY IMMERSION sion is multi-factorial in nature and is mediated in part by an increased proximal rejection of sodium in addition to the aldosterone-mediated effects. This interpretation is supported by previous data from our laboratory which show: 1) an augmented free water clearance at a time when sodium excretion was enhanced suggesting an increase in sodium delivery to the diluting site (2,3): 2) that immersion induces an increase in bicarbonate excretion with a concomitant increase in U-PP C O 2 (urine to blood Pco2 gradient), suggesting an increased proximal tubular rejection of sodium bicarbonate (9). Several lines of evidence have suggested that plasma sodium concentration per se can influence aldosterone secretion in both experimental animals and in man (10,11). Thus, McCaa et al. (10) have reported that a reduction in plasma sodium concentration without a significant change in fluid volume, plasma potassium concentration or pituitary secretion of ACTH can stimulate aldosterone secretion in the absence of a functional renin-angiotensin system. This concept has recently been extended to suggest that the saline-induced suppression of the renin-angiotensin-aldosterone system may be related to either saline or the sodium ion per se, rather than the volume expansion induced by the infusion. Thus, Tuck et al. (11) demonstrated in sodium-depleted subjects that the rate of suppression of the renin-aldosterone system by acute volume expansion with saline was more rapid than that with a dextran and glucose infusion calculated to produce equivalent expansion. The authors suggested that their data support a specific role for the sodium ion per se in the regulation of renin and aldosterone system. In the present study, the suppression by immersion of PRA and PA was indistinguishable from that produced by acute saline expansion in the study of Tuck et al. as far as the time course and magnitude. Thus the results of the present study militate against the interpretation that the rapid
623
suppression of PRA and PA induced by saline represents an effect unique to saline. Rather, other factors common to both saline administration and the immersion model, including an increase in central blood volume (12), might explain these differences. Although there have been a few previous investigations of the acute responsiveness of the renin-angiotensin-aldosterone axis to volume expansion, significant differences in experimental design render comparisons difficult. Thus, Pickens and Enoch assessed the response of PRA to the administration of 6% dextran (500 ml in 2 - 3 h) to 2 normal seated subjects and reported that PRA fell 37% from control 4 h after the infusion (13). Kem et al. (14) administered 2 liters of isotonic saline over a 4-h period in a group of normal supine subjects and reported a 50% decline in plasma aldosterone levels following the infusion. Recently Tuck et al. (11) examined the simultaneous responses of PRA, angiotensin II and plasma aldosterone to acute volume expansion induced by either saline or dextran infusion in a group of normal recumbent subjects in balance on a 10 meq Na/100 meq K diet. Blood samples were obtained every 10 min for 30 min and hourly thereafter. These authors demonstrated a rapid decline in mean PRA levels after the start of saline infusion, with a significant fall of 20% by 10 min and a 50% decline by 60 min. In contrast to PRA, there was an initial delay in the decrease of plasma aldosterone with a subsequent rapid decline to 50% of the control value by 60 min. The results of the present study demonstrating a 50% decline in PRA and a 52% decline in PA at 60 min of immersion, are generally in accord with the above studies. Recent demonstrations in our laboratory have shown that water immersion and acute saline administration (2 liters/120 min) in the seated posture both induce similar increments in cardiac output and central blood volume (Begin, Epstein and Sackner,
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EPSTEIN, PINS, SANCHO AND HABER
unpublished observations). This suggests that the acute induction of central blood volume expansion, regardless of the experimental model utilized, results in a rapid and parallel suppression of PRA and aldosterone. Although many studies have demonstrated that alterations of aldosterone secretion in response to volume changes are mediated primarily by the renin-angiotensin system (15-17), some investigators (18-20) have suggested that the renin-angiotensin system is not necessarily the primary mechanism leading to increased secretion of aldosterone during sodium or volume changes. Thus, Bestet al. (20) have recently reported that dietary sodium restriction does not stimulate a rise in circulating angiotensin II in normal man, thereby raising a question concerning the role of angiotensin II as the stimulus to aldosterone secretion during dietary sodium restriction. Similarly, Lowenstein and Steele have recently reported that following propranolol-induced beta adrenergic blockage, plasma aldosterone concentration increased in response to dietary sodium restriction despite continued suppression of PRA and angiotensin II (21). The current demonstration of a parallel rise in the pre-study PRA and PA values in response to 5-8 days of dietary sodium restriction and the parallelism of the suppression of PRA and PA in response to immersion is consistent with the postulate that the stimulation of aldosterone secretion during sodium restriction is indeed mediated by the renin-angiotensin system. However, since measurement of PRA and aldosterone were not obtained at intervals frequent enough to discern a lag between the decline in PRA and that of aldosterone, a causal relationship cannot be established with certainty. While previous studies have attempted to assess the relationship between PRA and aldosterone by inducing volume and/or postural manipulation, such manipulations have been designed to favor a stimulation
JCE & M • 1975 Vol 41 • No 3
rather than a suppression of PRA and aldosterone (15,16,19). The current demonstration of a parallel suppression of PRA and aldosterone in response to immersion extends these earlier findings, and emphasizes the importance of the renin-angiotensin axis in mediating the control of aldosterone in many diverse clinical and experimental settings. In conclusion, the present observations further define the temporal profile of the suppression of PRA and aldosterone during immersion and demonstrate that PRA and PA respond in a parallel manner. Furthermore, the present findings suggest that in selected circumstances, immersion may constitute a preferred investigative model for comparing the relative autonomy of PRA and aldosterone in response to suppressive manuevers. Since previous studies have demonstrated that immersion induces a natriuresis and kaliuresis without a concomitant increase in total blood volume and with a decrease in body weight rather than the increase which attends saline infusion (22), it is suggested that the water immersion model constitutes an alternative means of assessing the effects of volume expansion on renin-aldosterone without the complexities and potential hazards which might accompany acute saline expansion in edematous subjects. Acknowledgments The authors are indebted to David C. Duncan, Roger D. Loutzenhiser, Arthur G. DeNunzio, Robert Arrington and Peggy D. Cumby for their outstanding assistance in various aspects of the study. Excellent technical assistance in the performance of the immunoassays was provided by Virginia Lucas and Elizabeth Dimock. This work was supported by grants from the National Aeronautics and Space Administration (NGR 10-007097 and NAS-9-11846), and by designated Veterans Administration Research Funds. Parts of this study were conducted on the Clinical Research Unit of the University of Miami School of Medicine, supported by a grant from the General Clinical Research Centers Program of the Division of Research Resources, National Institutes of Health (RR-261).
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION References 1. Epstein, M. and T. Saruta,/ Appl Physiol 31: 368, 1971. 2. , D. Duncan, and L. M. Fishman, Clin Sci 43: 275, 1972. 3. , J. L. Katsikas, and D. C. Duncan, Circ Res 32: 228, 1973. 4. Poulsen, K.,J. Sancho, andE. Haber, Clin Immunol Immunopathol 2: 373, 1974. 5. Haber, E., T. Koerner, L. B. Page, B. Kliman, and A. J. PurnodeJ. Clin Endocrinol Metab 29: 1349, 1969. 6. Crane, M. C , and J. J. Harris, Metabolism 23: 359, 1974. 7. Buckalew, V. M., and C. D. Lancaster, Clin Sci 42: 69, 1972. 8. Bourgoignie, J. J., K. H. Hwang, E. Ipakchi, and N. S. BrickerJ Clin Invest 53: 1559, 1974. 9. Epstein, M., N. S. Schneider, and C. A. Vaamonde, J Lab Clin Med 84: 777, 1974. 10. McCaa, R. E., J. D. Bower, and C. S. McCaa, Circ Res 32: 555, 1973. 11. Tuck, M. L., R. G. Dluhy, and G. H. Williams, 7 Clin Invest 53: 988, 1974. 12. Begin, R., M. Epstein, M. Sackner, R. Levinson, R.
13. 14. 15. 16. 17. 18. 19.
20. 21. 22.
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Dougherty, and D. Duncan, Fed Proc 34:416,1975 (Abstract). Pickens, P. T., and B. A. Enoch, Cardiovasc Res 2: 157, 1968. Kem, D. C., M. H. Weinberger, D. M. Mayes, and C. A. Nugent, Arch Intern Med 128: 380, 1971. Liddle, G. W., L. E. Duncan, Jr., and F. C. Bartter, Am J Med 21: 380, 1956. Laragh, J. H., and H. C. StoerkJ Clin Invest 36: 383, 1957. William, G. H., J. P. Cain, R. G. Dluhy, and R. H. Underwood, J Clin Invest 51: 1731, 1972. Miiller, J., Regulation of Aldosterone Biosynthesis, Springer-Verlag, New York, 1971, p. 108. Blair-West, J. R., M. Cain, K. Catt, J. P. Coghlan, D. A. Denton, J. W. Funder, B. A. Scoggins, and R. D. Wright, In Gaul, C. (ed.), Progress in Endocrinology, Excerpta Medica Foundation Publishers, Amsterdam, 1969, p. 276. Best, J. B., J. P. Coghlan, J. H. N. Bett, E. J. Cran, and B. A. Scoggins, Lancet 2: 1353, 1971. Lowenstein, J., and J. M. Steele, Jr., Proc Am Soc Nephrol, Washington, D.C., 1974, p. 54 (abstract). Epstein, M., D. S. Pins, R. Arrington, A. G. Denunzio, and R. Engstrom, J Appl Physiol 39: 66, 1975.
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P
REVIOUS studies from this laboratory have demonstrated that the redistribution of blood volume and concomitant central hypervolemia that accompany headout water immersion to the neck result in a significant and reproducible natriuresis and a suppression of the renin-angiotensin-aldosterone system (1-3). Since changes in aldosterone secretion were determined by measurements of aldosterone excretion (1) rather than plasma aldosterone, the timecourse of the aldosterone alterations in response to immersion and their relation to concomitant changes in PRA could not be elucidated (1). Furthermore, although a 70% decrease in plasma renin activity was documented in these studies (1), determination of PRA at two-hourly intervals precluded assessment of the rapidity of changes resulting from initiation or discontinuation of immersion. Received March 31, 1975. * Dr. Epstein is an Investigator of the Howard Hughes Medical Institute.
30 min with maximal suppression (62%) by 180 min (P < 0.001). Recovery from NI was associated with a prompt return to pre-study levels. The changes in PA paralleled those of PRA with regard to both the rapidity and magnitude of the suppression (r = 0.993: P < 0.001). These data emphasize the importance of central volume per se as a primary determinant of PRA and PA regulation in normal man. Furthermore, the current studies confirm the importance of the renin-angiotensin axis in the control of volumerelated changes in PA in normal man. The ability of NI to induce a prompt and parallel suppression of PRA and PA without concomitant alterations in plasma composition, suggests that NI may be a preferred investigative tool for assessing the effects of volume expansion on renin-aldosterone. (/ Clin Endocrinol Metab 41: 618, 1975)
The development of a sensitive and specific radioimmunoassay for plasma aldosterone (4) has made possible the assessment, by direct measurement, of the effect of water immersion on aldosterone. The current study was undertaken to characterize the temporal profile of the suppression of PRA and plasma aldosterone during immersion and to elucidate further the role of aldosterone suppression in mediating the natriuresis of water immersion. Materials and Methods Nine men between the ages of 19 and 25 y were studied. There was no prior history of hypertension, cardiovascular disease or diabetes. Significant renal disease was excluded by documenting a normal urinary sediment, creatinine clearance and sterile urine cultures. The subjects were housed during the study in an environmentally controlled metabolic ward at a constant temperature. Each consumed a diet, the composition of which remained unchanged throughout the study, containing 10 meq of sodium, 100 meq of potassium and 2,500 ml of water. Daily 24-h
618
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION urine collections were made for determination of sodium, potassium and creatinine. After dietary equilibration, each subject underwent a Control study on day 5, followed on days 7-8 by an Immersion study. The experimental protocols on the 2 study days were similar and were carried out as follows: After 14 h of overnight water deprivation, the subject was awakened at 0700, voided and sat quietly for 1 h. At 0800, an oral water load of 700 ml was administered and at 0815 the subject voided again. Immediately after voiding, a 20 gauge lVfe" Teflon catheter with a flash chamber and an accompanying Teflon stylet (Jelco Co., Raritan, N.J.) was inserted into a forearm vein, permitting sequential sampling of blood without the use of an anti-coagulant or an intravenous infusion. Immediately after insertion, venous blood was drawn using a pre-chilled 10 ml polystyrene disposable syringe and an aliquot was collected for renin and aldosterone measurements in a pre-chilled vacutainer tube containing EDTA. After obtaining the sample, a sterile stylet was inserted into the catheter. Blood samples were obtained before and after the study for sodium, potassium, creatinine and osmolality determinations. During the control study, the subject sat quietly outside the immersion tank for the 6-h period beginning with the 0815 voiding. During the control study, the subject sat quietly outside the immersion tank for the 6-h period beginning with the 0815 voiding. During the immersion study, the subject sat in the tank immersed in water to the neck for 4 h (915-1315), preceded and followed by 1 h of quiet sitting outside the tank (Pre-Study and Recovery hours, respectively). Each subject stood briefly to void spontaneously at hourly intervals during the study. To maintain adequate urine flow, 200 ml of water was administered orally every hour during the study. Sodium, potassium, creatinine and osmolality were measured in aliquots of the hourly urine collections. Blood was collected at 30-min intervals for renin and aldosterone determinations. All subjects were weighed every morning at 0700 after voiding and before and after each study. Immersion was carried out in a waterproof tank described in detail in previous communications (2,3). A constant water temperature of 34 ± 0.5 C was maintained by two heat exchangers, controlled by an adjustable temperature-calibrated
619
control meter with input derived from two thermistors immersed at different water levels. Plasma renin was measured by radioimmunoassay according to the method of Haber et al. (5). Plasma aldosterone was measured by a modification of the radioimmunoassay technique of Poulsen et al. (4). The modification utilizes a new antibody with a very high affinity constant (10" n M/liter), so that 50% displacement is obtained with only 8 pg of aldosterone (Sancho and Haber, to be published). With this modification, aldosterone could be assayed utilizing only 0.1 ml of plasma. The coefficient of variation for interassay determinations is 10.2%. Analytic methods for sodium, potassium and creatinine determinations have been reported previously (2,3). In the presentation of the data, mean values are followed by the standard error of the mean as an index of dispersion. Data were evaluated statistically using Student's t test for paired values or, where appropriate by the Wilcoxon signed rank test. Differences with P < 0.05 were considered significant. Permission for the study was obtained from each subject after a detailed description of the procedure and the potential complications. The protocol was approved by the Human Experimentation Committees of the University of Miami School of Medicine and the Miami Veterans Administration Hospital and was in compliance with the principles outlined in the Declaration of Helsinki. No complications occurred.
Results Urinary electrolyte excretion The effect of 4 h of water immersion on sodium and potassium excretion is shown in Table 1. During quiet sitting (Control), the rate of sodium excretion (UNa V) was constant, ranging from 3 to 6 /aeq/min. Immersion resulted in a significant increase in UNaV compared to Control beginning with the second hour of Immersion. By hour 4, UNaV was 10-fold greater than during the comparable Control period. Recovery was associated with a decline in UNaV, but UNaV continued to exceed both preimmersion values and the values during the comparable Control hour. The rate of potassium excretion (UKV)
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JCE & M • 1975 Vol41« iNo3
EPSTEIN, PINS, SANCHO AND HABER
TABLE 1. Effect of immersion on urinaiy excretory patterns (results are mean ± SE of 9 subjects) Time (h) Group
1
Pre-study
Control Immersion
3.5 ± 0.8 3.1 ± 0.5
(jxeq/min)
Control Immersion
5.1 ± 1.5 2.5 ± 0.7
UKV (/neq/min)
Control Immersion
Ccr
Control Immersion
ml/vmin
u Na v
(ml/min)
51 49
± 12 ± 11
4
4.0 ± 0.4 7.6 ± 0.6*
3.7 ± 0.6 5.6 ± 0.6
3.8 ± 0.8 4.3 ± 0.5
2.5 ± 0.3 1.8 ± 0.5
6.0 ± 2.1 28.6 ± 10.3*
5.3 ± 2.5 39.5 ± 15.9*
5.6 ± 2.5 55.6 ± 19.9*
3.0 ± 0.7 38.2 ± 16.8*
2.6 ± 0.5 5.8 ± 0.9* 5.5 ± 2.4 13.4 ± 3.8 ± 11 ± 9*
58 78
120 ± 8 114 ± 5
3
2
121 ± 5 133 ± 8
60 ± 7 111 ± 8*
55 ± 9 106 ± 10*
118 ± 5 121 ± 4
109 122
± 6 ± 5
61 87
± 7 ± 8
111 ± 6 114 ± 8
Recovery
55 50
± 3 ± 7
108 ± 6 118 ± 8
* P < 0.05 as compared with Control.
was significantly increased during each of the initial 3 h of Immersion. Creatinine clearance (CCr) during Control was not significantly different from the Control pre-study hour, ranging from 108 to
121 ml/min. Immersion did not significantly alter CCr. Positive fluid balance (water intake minus urine volume) during Control of 253 ± 75 ml was observed. In contrast, fluid
IMMERSION
12 r-
10 L-'TN PRA ng/ml/hr
•i
8
N.S.
\ . . .
CONTROL
—
IMMERSION
N.S.
; \
(n = 9) MEAN + S.E. r
p < 0.001
>
p < 0.005
0
60
120
180
FlG. 1. Effect of water immersion to the neck on plasma renin activity (PRA) in subjects in balance on a 10 meq sodium diet. Immersion resulted in a progressive decrement in PRA beginning as early as 30 min. Cessation of immersion (Recovery) was associated with a prompt return of PRA to prestudy values.
240 30 60
TIME (minutes)
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION
621
IMMERSION (n = 9) MEAN 1 S.E.
500 FIG. 2. Effect of water immersion to the neck on plasma aldosterone in subjects in balance on a 10 meq sodium diet. Immersion resulted in a progressive decrement in plasma aldosterone beginning as early as 60 min. Cessation of immersion (Recovery) was associated with a prompt return of plasma aldosterone to pre-study values.
PLASMA
r
y?\
400
ALDOSTERONE LEVELS
300
N.S.
- —
pg/ml
N.s\ 200
100
CONTROL
— — IMMERSION \
IXl * p < 0.001 0 p < 0.005 I
I
0
60
120
180
240 30 60
TIME (minutes)
balance was negative by 299 ± 64 ml during Immersion, significantly different from Control (P < 0.001).
was associated with significant suppression of aldosterone beginning as early as 30 min. By 210 min of Immersion, aldosterone was maximally suppressed to 34% of the prePlasma renin activity (PRA) study value. Recovery was associated with a prompt return to pre-study values. As shown in Fig. 1, plasma renin activity was appropriately elevated in re- Relationship between PRA and plasma sponse to dietary sodium restriction through- aldosterone out the Control study and in the pre-study As shown in Fig. 3, the suppression of hour of Immersion. Immersion resulted in PRA and PA in response to immersion and a significant suppression of PRA beginning their subsequent recovery following cessawithin 30 min of study. By 180 min, PRA tion of immersion occurred in parallel was suppressed to 38% of the pre-study (r = 0.993; P < 0.001). value. Cessation of Immersion was associated with a prompt return toward pre- Serum electrolytes study values as early as 30 min of Recovery; Serum sodium (137 ± 1 meq/liter) and osby 60 min of Recovery, PRA was not difmolality (282 ± 1 mOsm/liter) were identiferent from the pre-study value. cal during the Control and Immersion studies. Similarly, serum potassium concenChanges in plasma aldosterone (PA) tration was similar during the two studies As shown in Fig. 2, plasma aldosterone and did not change significantly during the levels were also appropriately elevated control and immersion studies when comthroughout the Control study and during the pared with values obtained immediately pre-study hour of Immersion. Immersion prior to each study.
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622
EPSTEIN, PINS, SANCHO AND HABER
Discussion Water immersion to the neck has previously been shown to produce a profound natriuresis and a suppression of the reninaldosterone system (1-3,6). The nature of prior studies did not permit a kinetic examination of changes in plasma renin and aldosterone in relation to onset and offset of immersion nor their precise correlation with renal sodium and potassium excretion. The present study demonstrates that immersion induces a rapid (30 min) and profound suppression of both PRA and plasma aldosterone with an equally rapid recovery following discontinuation of immersion. The dissociation between plasma aldosterone and renal sodium handling during the recovery hour merits comment. During recovery, at a time when both PRA and aldosterone levels had returned to prestudy levels, the rate of sodium excretion continued to exceed both the pre-immersion value and the comparable control values.
JCE & M • 1975 VoUl • No 3
The prompt return of PRA and plasma aldosterone to pre-study levels suggests that cessation of immersion was associated with normalization ofblood volume distribution which was sensed by the appropriate volume receptors. Despite this apparent normalization of the distribution of circulating blood volume, the natriuresis persisted suggesting that a humoral factor rather than more rapidly acting hemodynamic and neural mechanisms mediated the natriuresis of immersion (7,8). The current demonstration of a progressive kaliuresis during immersion helps to elucidate further the mechanism of the natriuresis of immersion. In view of the concomitant decrease in plasma aldosterone concentration, one would have anticipated that potassium excretion would have diminished. The current demonstration of increased K excretion is consistent, however, with previous studies from this laboratory suggesting that the natriuresis of immer-
100 FIG. 3. Comparison of the effects of immersion on plasma renin activity (PRA) and plasma aldosterone in subjects in balance on a 10 meq Na diet. Data are expressed in terms of percent change from the pre-immersion hour. As can be seen, the suppression of plasma aldosterone paralleled the suppression of PRA throughout the immersion period. Similarly, following cessation of immersion, the recovery of both PRA and plasma aldosterone occurred in parallel.
80 0/
/o
DECREASE FROM
6 0
PRE-STUDY
= 0.993
40
20
0
0
•—•
PLASMA RENIN
o—o
PLASMA
60
ACTIVITY
ALDOSTERONE
120
180
240
60
TIME (minutes)
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RENIN-ALDOSTERONE SUPPRESSION BY IMMERSION sion is multi-factorial in nature and is mediated in part by an increased proximal rejection of sodium in addition to the aldosterone-mediated effects. This interpretation is supported by previous data from our laboratory which show: 1) an augmented free water clearance at a time when sodium excretion was enhanced suggesting an increase in sodium delivery to the diluting site (2,3): 2) that immersion induces an increase in bicarbonate excretion with a concomitant increase in U-PP C O 2 (urine to blood Pco2 gradient), suggesting an increased proximal tubular rejection of sodium bicarbonate (9). Several lines of evidence have suggested that plasma sodium concentration per se can influence aldosterone secretion in both experimental animals and in man (10,11). Thus, McCaa et al. (10) have reported that a reduction in plasma sodium concentration without a significant change in fluid volume, plasma potassium concentration or pituitary secretion of ACTH can stimulate aldosterone secretion in the absence of a functional renin-angiotensin system. This concept has recently been extended to suggest that the saline-induced suppression of the renin-angiotensin-aldosterone system may be related to either saline or the sodium ion per se, rather than the volume expansion induced by the infusion. Thus, Tuck et al. (11) demonstrated in sodium-depleted subjects that the rate of suppression of the renin-aldosterone system by acute volume expansion with saline was more rapid than that with a dextran and glucose infusion calculated to produce equivalent expansion. The authors suggested that their data support a specific role for the sodium ion per se in the regulation of renin and aldosterone system. In the present study, the suppression by immersion of PRA and PA was indistinguishable from that produced by acute saline expansion in the study of Tuck et al. as far as the time course and magnitude. Thus the results of the present study militate against the interpretation that the rapid
623
suppression of PRA and PA induced by saline represents an effect unique to saline. Rather, other factors common to both saline administration and the immersion model, including an increase in central blood volume (12), might explain these differences. Although there have been a few previous investigations of the acute responsiveness of the renin-angiotensin-aldosterone axis to volume expansion, significant differences in experimental design render comparisons difficult. Thus, Pickens and Enoch assessed the response of PRA to the administration of 6% dextran (500 ml in 2 - 3 h) to 2 normal seated subjects and reported that PRA fell 37% from control 4 h after the infusion (13). Kem et al. (14) administered 2 liters of isotonic saline over a 4-h period in a group of normal supine subjects and reported a 50% decline in plasma aldosterone levels following the infusion. Recently Tuck et al. (11) examined the simultaneous responses of PRA, angiotensin II and plasma aldosterone to acute volume expansion induced by either saline or dextran infusion in a group of normal recumbent subjects in balance on a 10 meq Na/100 meq K diet. Blood samples were obtained every 10 min for 30 min and hourly thereafter. These authors demonstrated a rapid decline in mean PRA levels after the start of saline infusion, with a significant fall of 20% by 10 min and a 50% decline by 60 min. In contrast to PRA, there was an initial delay in the decrease of plasma aldosterone with a subsequent rapid decline to 50% of the control value by 60 min. The results of the present study demonstrating a 50% decline in PRA and a 52% decline in PA at 60 min of immersion, are generally in accord with the above studies. Recent demonstrations in our laboratory have shown that water immersion and acute saline administration (2 liters/120 min) in the seated posture both induce similar increments in cardiac output and central blood volume (Begin, Epstein and Sackner,
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624
EPSTEIN, PINS, SANCHO AND HABER
unpublished observations). This suggests that the acute induction of central blood volume expansion, regardless of the experimental model utilized, results in a rapid and parallel suppression of PRA and aldosterone. Although many studies have demonstrated that alterations of aldosterone secretion in response to volume changes are mediated primarily by the renin-angiotensin system (15-17), some investigators (18-20) have suggested that the renin-angiotensin system is not necessarily the primary mechanism leading to increased secretion of aldosterone during sodium or volume changes. Thus, Bestet al. (20) have recently reported that dietary sodium restriction does not stimulate a rise in circulating angiotensin II in normal man, thereby raising a question concerning the role of angiotensin II as the stimulus to aldosterone secretion during dietary sodium restriction. Similarly, Lowenstein and Steele have recently reported that following propranolol-induced beta adrenergic blockage, plasma aldosterone concentration increased in response to dietary sodium restriction despite continued suppression of PRA and angiotensin II (21). The current demonstration of a parallel rise in the pre-study PRA and PA values in response to 5-8 days of dietary sodium restriction and the parallelism of the suppression of PRA and PA in response to immersion is consistent with the postulate that the stimulation of aldosterone secretion during sodium restriction is indeed mediated by the renin-angiotensin system. However, since measurement of PRA and aldosterone were not obtained at intervals frequent enough to discern a lag between the decline in PRA and that of aldosterone, a causal relationship cannot be established with certainty. While previous studies have attempted to assess the relationship between PRA and aldosterone by inducing volume and/or postural manipulation, such manipulations have been designed to favor a stimulation
JCE & M • 1975 Vol 41 • No 3
rather than a suppression of PRA and aldosterone (15,16,19). The current demonstration of a parallel suppression of PRA and aldosterone in response to immersion extends these earlier findings, and emphasizes the importance of the renin-angiotensin axis in mediating the control of aldosterone in many diverse clinical and experimental settings. In conclusion, the present observations further define the temporal profile of the suppression of PRA and aldosterone during immersion and demonstrate that PRA and PA respond in a parallel manner. Furthermore, the present findings suggest that in selected circumstances, immersion may constitute a preferred investigative model for comparing the relative autonomy of PRA and aldosterone in response to suppressive manuevers. Since previous studies have demonstrated that immersion induces a natriuresis and kaliuresis without a concomitant increase in total blood volume and with a decrease in body weight rather than the increase which attends saline infusion (22), it is suggested that the water immersion model constitutes an alternative means of assessing the effects of volume expansion on renin-aldosterone without the complexities and potential hazards which might accompany acute saline expansion in edematous subjects. Acknowledgments The authors are indebted to David C. Duncan, Roger D. Loutzenhiser, Arthur G. DeNunzio, Robert Arrington and Peggy D. Cumby for their outstanding assistance in various aspects of the study. Excellent technical assistance in the performance of the immunoassays was provided by Virginia Lucas and Elizabeth Dimock. This work was supported by grants from the National Aeronautics and Space Administration (NGR 10-007097 and NAS-9-11846), and by designated Veterans Administration Research Funds. Parts of this study were conducted on the Clinical Research Unit of the University of Miami School of Medicine, supported by a grant from the General Clinical Research Centers Program of the Division of Research Resources, National Institutes of Health (RR-261).
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RENIN-ALDOSTERONE SUPPRESION BY IMMERSION References 1. Epstein, M. and T. Saruta,/ Appl Physiol 31: 368, 1971. 2. , D. Duncan, and L. M. Fishman, Clin Sci 43: 275, 1972. 3. , J. L. Katsikas, and D. C. Duncan, Circ Res 32: 228, 1973. 4. Poulsen, K.,J. Sancho, andE. Haber, Clin Immunol Immunopathol 2: 373, 1974. 5. Haber, E., T. Koerner, L. B. Page, B. Kliman, and A. J. PurnodeJ. Clin Endocrinol Metab 29: 1349, 1969. 6. Crane, M. C , and J. J. Harris, Metabolism 23: 359, 1974. 7. Buckalew, V. M., and C. D. Lancaster, Clin Sci 42: 69, 1972. 8. Bourgoignie, J. J., K. H. Hwang, E. Ipakchi, and N. S. BrickerJ Clin Invest 53: 1559, 1974. 9. Epstein, M., N. S. Schneider, and C. A. Vaamonde, J Lab Clin Med 84: 777, 1974. 10. McCaa, R. E., J. D. Bower, and C. S. McCaa, Circ Res 32: 555, 1973. 11. Tuck, M. L., R. G. Dluhy, and G. H. Williams, 7 Clin Invest 53: 988, 1974. 12. Begin, R., M. Epstein, M. Sackner, R. Levinson, R.
13. 14. 15. 16. 17. 18. 19.
20. 21. 22.
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Dougherty, and D. Duncan, Fed Proc 34:416,1975 (Abstract). Pickens, P. T., and B. A. Enoch, Cardiovasc Res 2: 157, 1968. Kem, D. C., M. H. Weinberger, D. M. Mayes, and C. A. Nugent, Arch Intern Med 128: 380, 1971. Liddle, G. W., L. E. Duncan, Jr., and F. C. Bartter, Am J Med 21: 380, 1956. Laragh, J. H., and H. C. StoerkJ Clin Invest 36: 383, 1957. William, G. H., J. P. Cain, R. G. Dluhy, and R. H. Underwood, J Clin Invest 51: 1731, 1972. Miiller, J., Regulation of Aldosterone Biosynthesis, Springer-Verlag, New York, 1971, p. 108. Blair-West, J. R., M. Cain, K. Catt, J. P. Coghlan, D. A. Denton, J. W. Funder, B. A. Scoggins, and R. D. Wright, In Gaul, C. (ed.), Progress in Endocrinology, Excerpta Medica Foundation Publishers, Amsterdam, 1969, p. 276. Best, J. B., J. P. Coghlan, J. H. N. Bett, E. J. Cran, and B. A. Scoggins, Lancet 2: 1353, 1971. Lowenstein, J., and J. M. Steele, Jr., Proc Am Soc Nephrol, Washington, D.C., 1974, p. 54 (abstract). Epstein, M., D. S. Pins, R. Arrington, A. G. Denunzio, and R. Engstrom, J Appl Physiol 39: 66, 1975.
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