J. Endocrinol. Invest. 13: 727 -735, 1990
Nocturnal oscillations of plasma aldosterone in relation to sleep stages M.O. Krauth, J. Saini, M. Follenius, and G. Brandenberger Laboratoire de Physiologie et de Psychologie Environnementales CNRS/INRS UMR 32, 21 rue Becquerel, 67087 Strasbourg Cedex, France ABSTRACT. This study examined the relationship between aldosterone secretion and sleep stages in conjunction with two aldosterone regulating hormone systems, the renin-angiotensin system (RAS) and adrenocorticotropin (ACTH), and also K+. Nocturnal plasma patterns of aldosterone, plasma renin activity (PRA), ACTH and K+ were established in blood collected at 10-min intervals in two groups of 6 subjects. Both groups underwent two 9 hour overnight-studies, consisting of one control night and one experimental night. The first group was maintained on a low Na diet and the other was given a beta-blocker, atenolol. Polygraphic recordings of sleep were scored according to established criteria. For the control night, REM sleep usually began at peak level or in the descending phase of aldosterone oscillations. As previously described, PRA reflected REM-NREM sleep alteration, levels increased in NREM and decreased during REM sleep. ACTH fluctuations did not oscillate with sleep stages, but levels were very seldom in the ascending phase at REM sleep onset.
Plasma K+ remained almost constant throughout the night. The relative importance of the ACTH and the RAS on nocturnal aldosterone secretion and the relationship between aldosterone oscillations and sleep stages remained unclear. Modulating renin levels by either consuming a low Na diet or administration of a beta-blocker enabled this relationship to be clarified. The RAS dominated aldosterone secretion when stimulated by a low sodium diet. Aldosterone oscillations then reflected PRA oscillations with a delay of about 20 min and the relationship of aldosterone to sleep stages was dependent on the relationship of PRA with sleep stages. Atenolol depressed PRA and in this case aldosterone levels reflected those of ACTH with a delay of about 10 min. To conclude, this study shows that the nocturnal secretion of aldosterone is under control of both the RAS and ACTH. Stimulating and depressing the RAS revealed that the relationship of aldosterone to sleep stages is dependent on the relationship of these hormonal systems to sleep stages.
INTRODUCTION The secretion of aldosterone has been referred to as episodic with a general increase in mean levels during the last hours of sleep (1-3). The two major hormonal systems involved in the regulation of aldosterone are the renin-angiotensin system (RAS) and the adrenocorticotropic system (4). The relative importance of these two hormonal systems in controlling the ultradian aldosterone secretory pattern remains unclear. Studies of nocturnal aldosterone secretion differ in their conclusions. A relationship
between plasma peaks of aldosterone and cortisol but not between PRA and aldosterone was reported in sleeping subjects using a 20 to 30 min blood sampling method (5). Alternatively, dexamethasone administration has been shown to abolish the cortisol response but not to affect nocturnal plasma aldosterone levels (5, 6) or alter the quantity excreted (7), suggesting ACTH has little effect on aldosterone secretion. Apart from Rubin et al. (8) who established a 90min nocturnal oscillatory pattern which could not be related to a specific sleep stage, the relationship of aldosterone with sleep stages has not been examined. Plasma renin activity (PRA), measured as an index of renin secretion, and adrenocorticotropin (ACTH) have different nocturnal secretory patterns. PRA is strongly associated with sleep stages (9),
Key-words: Aldosterone, ACTH, renin, sleep, ultradian rhythm. Correspondence: Dr. G. Brandenberger L.P.P.E., 21 rue Becquerel, 67087 Strasbourg Cedex, France. Received July 17, 1989; accepted June 28, 1990.
Mo. Krauth, J. Saini, M Fol/enius, et al.
rapid eye movement (REM) sleep phases always occur when PRA levels are decreasing and non (N)REM sleep phases invariably coincide with increasing PRA levels (10, 11 ). Contrarily, the secretion of ACTH and cortisol is considered to be circadian rather than sleep-related (12), although recent studies suggest that there may be a temporal relationship between ACTH and REM sleep onset as REM sleep seems to coincide with a period of reduced hypothalamo-pituitary activity (13). The aim of this study was to investigate the relationship between aldosterone secretion and sleep stages in conjunction with the two major stimulating factors, RAS and ACTH. A 10-min blood sampling technique was employed to examine firstly the nocturnal plasma profiles of aldosterone, PRA and ACTH together with continuous sleep recordings in normal healthy subjects. Secondly, the effect of either consuming a low Na diet, which stimulates the RAS, or administration of a beta-blocker which depresses the RAS, was investigated.
trooculograms, one electromyogram and one electrocardiogram. Sleep stages were scored from the polygraphic recordings according to established criteria (14). At 21 :00 h a catheter was inserted into an antecubital vein. Blood was removed continuously using a peristaltic pump and collected in an adjoining room at 10-min intervals from 23:00 to 08:00 h. Light was switched off at 23:00 h and the subjects were awaked at 08:00 h. A maximum of 200 ml of blood was removed.
Plasma sample measurements Blood samples were immediately centrifuged at 4 C and the plasma stored at -20 C. Plasma aldosterone was measured by a radioimmunoassay method based on the method of Gianotti et at. (15). The aldosterone antiserum was prepared against a bovine serum albumine conjugato of aldosterone-3O-carboxymethyl-oxime-18-21 -diacetate. PRA was measured by radioimmunoassay of angiotensin I generated in vitro (16). ACTH was measured by radioimmunoassay using kits from CEA-Sorin (Saluggia, Italy). Plasma K+ was measured by flame photometry. All samples from one night were processed in the same assay to eliminate the effect of interassay variability. In duplicate samples, intraassay precision was for aldosterone 7.5% for samples > 10 ng 100 ml- 1 and 10% for samples < 10 ng 100 ml- 1 , the limit of sensitivity was 0.5 ng 100 ml- 1 . For PRA the intraassay coefficient of variation was 4% for levels between 10 and 20 ng ml-1.h- 1 , 6% for levels between 2 and 10 ng ml- 1 h- 1 , 10% for levels between 1 and 2 ng ml-1.h- 1 and 30% for levels < 1 ng ml- 1 . h- 1 , the limit of sensitivity was 0.18 ng ml- 1 . h- 1 . ACTH had an intraassay coefficient of variation of 3.5% for values of> 10 pg ml- 1 and 5% for values < 10 pg ml- 1 , the limit of assay sensitivity was 5 pg ml- 1.
MATERIALS AND METHODS
Subjects and procedures Two groups of 6 healthy male subjects aged 21 to 26 yr were studied. They all had normal routines of work, meals and sleep, they had no evidence of any disease and were not taking any medication. Before their final enlistment, they took part in an experimental session to familiarize themselves with the experimental environment and with catheter insertion. Informed written consent was obtained from all subjects. Both groups underwent two 9 hour overnight studies separated by at least 2 weeks. During the control study all subjects remained on their usual Na diet (individual 24-h urinary Na secretion was between 104 and 205 mEq / day). For the second experiment, one group was maintained on a low Na diet during the three days preceding the night study (individual 24-h urinary Na secretion was between 24 and 75 mEq/ day) and the other group received, at 22:00 h on the experimental night, a single dose of 100 mg of atenolol (Tenormine, ICI-Pharma, Cergy, France). The studies were performed in sound-proof air conditioned sleep chambers. Before the subjects entered the sleep chamber, electrodes were attached for the following uninterrupted electrophysiological recordings: two electroencephalograms, two elec-
Data analysis Analysis of hormone profiles and sleep stages Hormonal peaks were identified using the pulse detection program ULTRA (17) at a threshold of 3 times the coefficient of variation. The program excludes all peaks of plasma hormone concentration for which the increase and the decrease are less than the predefined threshold. For each significant pulse, its total duration, duration of the ascending and descending phase and magnitude are quantified. Time-intervals between oscillations were taken as the duration of time between maximum levels of
Plasma aldosterone and sleep stages
two hormone oscillations. The significant oscillations were then analyzed peak by peak in relation to sleep stages and for aldosterone in relation to PRA and ACTH. The nocturnal trends for the different hormones were determined using an algorithm calculating polynomial regression line of the 3rd degree. To compare the three hormonal trends, the calculation was based on the percentage change from mean levels.
LOW SODIUM DIET
Mean nocturnal profiles For the control night, mean nocturnal plasma aldosterone, PRA and ACTH were 12.4 ± 2.4 ng 100 ml- 1 , 2.0 ± 0.2 ng ml- 1'h- 1 and 21.9 ± 1.5 pg ml- 1 , respectively. Plasma K+ remained constant throughout the night, giving a mean value of 4.1 ± 0.1 mEq 1- 1 . The nocturnal trends for each hormone are given in Figure 1 in the form of a polynomial regression curve calculated using the percentage changes from the mean value. ACTH and aldosterone tended to increase towards the latter part of the night whereas the curve for PRA was flatter indicating less overall change in nocturnal levels during the night. Stimulating the RAS by a low sodium diet and depressing its activity by administration of the betablocker atenolol, increased and decreased respectively mean nocturnal plasma PRA and aldosterone levels. Mean values were 6.5 ± 1.1 ng ml-1.h- 1 for PRA and 16.6 ± 2.6 ng 100 ml- 1 aldosterone for the group that had been on a low sodium diet and 1.0 ± 0.2 ng ml-1. h-1 for PRA and 5.8 ± 0.9 ng 100 ml-1 for aldosterone for the group that received atenolol. Plasma ACTH and K+ were not significantly affected by either treatments. Despite changes in mean levels of both PRA and aldosterone, polynomial regression analysis revealed that under both conditions the nocturnal trend of aldosterone strongly resembled that of ACTH (Fig. 1).
Fig. 1 - Mean nocturnal trends of aldosterone, ACTH and PRA
(least squared polynome calculated from the percentage changes of mean levels).
Individual nocturnal profiles The ULTRA peak detection program was used to detect significant hormone oscillations and to define the peak characteristics. These data were then used to examine the relationship between plasma aldosterone, ACTH, PRA and sleep stages.
Table 1 - Percentage of ACTH, PRA and Aldosterone oscillations in ascending (A), peak (P), descending (D), nadir (N) phase at
REM sleep onset. ACTH Night
Control (no. = 12) Low Na+ diet (no. = 6) B-bloker (no. = 6)
Number of REM sleep phases
48 26 20
6 15 15
23 27 35
44 38 45
27 19 5
2 0 0
4 4 5
75 85 50
17 12 45
23 12 20
19 15 20
33 58 25
35 15 35
M.G. Krauth, J. Saini, M. Fol/enius, et aJ.
Subjects on control nights Significant oscillations were identified for all 3 hormones. For the control night PRA oscillations, as previously described (11), reflected sleep stage cycles, levels were increasing when NREM sleep occurred and decreasing levels coincided with REM sleep (Table 1). The mean amplitude and timeinterval (time duration between peak levels) of the PRA oscillation were 1.0 ± 0.2 ng ml- 1 . h-1 and 107 ± 9 min, respectively. The periodicities ranged from 70 to 100 min in 9 out of the 12 subjects with regular sleep cycles, as shown by the nocturnal profiles of an individual subject in Figure 2. For the remaining 3 subjects a large variation of time-intervals was observed reflecting disturbed sleep patterns (11). The amplitude and frequency of ACTH fluctuations varied widely both between subjects and between the beginning and end of the night. ACTH did not reflect the sleep stage cycles, but a temporal relationship was observed in that REM sleep onset rarely occurred when ACTH was ascending (Table 1). Aldosterone oscillations had a mean amplitude of 5.5 ± 0.6 ng 100 ml-1 and showed a time-interval of between 90 and 130 min. The relationship of aldosterone to sleep stages, ACTH and PRA was unclear. The number of aldosterone oscillations preceded by either ACTH or PRA were identified (Table 2) however 35% of the peaks were preceded by both PRA and ACTH oscillations.
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2300 2400 100
Fig. 2 - Individual nocturnal profiles of plasma renin activity, aldosterone, ACTH and K in one typical subject on a normal Na diet.
Subjects on a low Na diet Following a low sodium diet the amplitude of PRA and aldosterone oscillations were enhanced, reaching 2.9 ± 0.6 ng ml-1 . h-1 for PRA and 7.4 ± 1.4 ng 100 ml-1 for aldosterone. The periodicities of the two hormone systems were similar and ranged from 80 to 110 min for PRA and 70 to 100 min for aldosterone. Under these conditions the nocturnal oscillatory pattern of aldosterone appeared to reflect that of PRA, as illustrated by the individual profiles
of 2 subjects (Fig. 3). The number of significant peaks that were preceded by the well defined PRA oscillations, increased to 48% (Table 2) giving evidence of the predominant influence of renin secretion in these conditions. The time lag between the PRA and aldosterone peak was approximately 20
Table 2 - Oscillation characteristics of plasma aldosterone under control conditions, following a low sodium diet, and following 8blocker administration. Condition Control (no. = 12) Low Na' diet (no. = 6) B-bloker (no. = 6)
Number of oscillations
% associated with PRA
% associated with ACTH
% associaled with PRA and ACTH
68 33 44
38 21 70
35 30 5
Plasma aldosterone and sleep stages