- Email: [email protected]
Physical exercise at night blunts the nocturnal increase of plasma melatonin levels in healthy humans
Llfe Sciences, Vol. 47, pp. 1989-1995 Printed In the U.S.A.
Pergamon Press
PHYSICAL EXERCISE AT NIGHT BLUNTS THE NOCTURNAL INCREASE OF PLASMA MELATONIN LEVELS IN HEALTHY HUMANS P. Monteleone, M. Maj, M. Fusco, C. Orazzo and D. Kemali Instituteof Medical Psychology and Psychiatry, 1st Medical School, University of Naples, Largo Madonna delle Grazie, 80138 Naples, Italy. (Received in final form September 20, 1990)
Summar_y The effects of physical exercise on nighttime melatonin secretion have never been investigated in humans. For this purpose, plasma melatonin levels were measured at different times during the day and the night in seven healthy men (aged 26-33 yrs), both in resting condition and before and alter a physical exercise performed between 10.40 and 11.00 p.m.. The exercise consisted in bicycling on a bicycle ergometer at 50% of the personal maximal work capacity (MWC) for 10 min, followed by other 10 rain of bicycling at 80% of the MWC. The results clearly showed that physical stress at night significantly blunts the nocturnal increase in plasma msiatonin levels (group X time interaction: p <0.00001 ; two-way ANOVA with repeated measures). These findings, taken together with the data of the literature, suggest that the response of the pineal gland to provocative stimuli may depend on its level of activity when the stimulus is applied. It is well established that the synthesis of the pineal indeolamine melatonin is influenced primarily by light and that norepinenephrine (NE) released from sympathetic nerve terminals within the pineal gland acts on beta- and, to a lesser degree, on alphaadrenergic receptors to stimulate melatonin production in the dark (1, 2). As a consequence, melatonin shows a characteristic circadian rhythm with low plasma levels during the day and high plasma concentrations during the night. This circadian pattern of melatonin secretion has been documented in several animal species, including man (3, 4). Although it has been argued that the sympathetic nerve endings in the pineal gland may take up circulating catecholamines protecting the pinealocytes from daytime stimulation (5), it has clearly been demonstrated that, in the rat, peripheral catecholamines released by certain stressful stimuli may promote pineal melatonin production during the day (6-9). Particularly, exposure of rats to various stressful procedures, which increase circulating catecholamines, enhances both pineal and serum melatonin levels (6-8). On the contrary, adrenalectomy blocks the increase in rat pineal melatonin content brought about by daytime physical immobilization stress and insulin-induced hypoglycemia (6, 8). When the effects of stress on melatonin have been investigated at night, opposite results have been obtained. In fact, a hind leg injection of saline into rats at night led to a dramatic decrease of pineal melatonin content (10) and swimming for 10 min at 23.10 h. depressed nighttime melatonin content in the rat epiphysis (11). However, the presence of species differences in the pineal responsiveness to provocative stressful stimuli has been demonstrated. In fact, in the 0024-3205/90 $3.00 +.00 Copyright (c) 1990 Pergamon Press plc
1990
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
Syrian hamster, insulin-induced hypoglycemia was not able to induce any rise in pineal melatonin content during the light hours (7). As regards the human beings, although some studies show that physical exercise may increase daytime plasma levels of melatonin in healthy subjects (12-16), the weight of the evidence suggests that a variety of stressful situations, some of which provoke intense responses in the sympathetic and other hormonal systems, do not induce any increase in plasma meiatonin levels during the day (17, 18). These data emphasize that the human epiphysis is not responsive to daytime sympathetic stimulation. At present, no study has been performed to evaluate the effect of stress on nighttime plasma meiatonin levels in humans. Therefore, we decided to assess the nocturnal pattern of melatonin secretion in healthy male subjects following a physical exercise applied during the dark hours. _Methods Seven healthy male subjects volunteered for the study. They were aged 26-36 yrs (mean :1: SD = 29.2:1: 2.2), had normal endocrine and metabolic functions and no family history of diabetes mellitus, and were drug-free for at least one month. All were within 15% of their ideal body weight and smoked less than 10 cigarettes per day. None of them participated regularly in any kind of sport. Only male subjects were included into the study to avoid any distortion in the response to physical exercise caused by sex differences. Prior to experimental sessions, the maximal working capacity (MWC) was determined for each subject using a bicycle ergometer (Table I). The initial work load applied was 50 watt and the load was increased by 10 watt every one min until exhaustion (19). The experimental sessions were two for each subject. In one of them, the subject underwent a submaximal work test, starting to bicycle on a work load of 50% of his own MWC for 10 min, immediately followed by 80% of MWC for another 10 rain. In the other, the subject rested supine in a bed. The two sessions were performed in random order at one week interval, on an ambulatory basis, in the months of February and March. Each subject came to our research unit at 12.00 h, when a first blood sample was collected by venipuncture. Then he was free to go away and was requested to come back at 6.00 p.m.. At 7.00 p.m., a standardized meal was eaten; at 8.00 p.m. a butterfly needle was inserted into an antecubital vein and kept patent with a slow saline infusion. Immediately after, a second blood sample was drawn, then the subject rested supine. At 9.00 p.m., light was turned off and all the subsequent procedures were carried out by the help of a red light. Further blood samples were collected at 10.00, 11.00, 11.30, 12.00 p.m. and at 1.00,2.00, 4.00, 6.00 and 8.00 a.m.. Physical exercise was started at 10.40 p.m. and, according to the above mentioned procedure, lasted until 11.00 p.m.. Blood pressure and heart rate were recorded, in the session involving physical exercise, at the same time points of blood sampling between 10.40 p.m. and 2.00 a.m.. Subjects were allowed to sleep between 11.30 p.m. and 7.00 a.m., when light was turned on. They did not smoke or drink coffee during the experimental sessions. Plasma was separated by centrifugation at 3000 rpm and stored at -20 °C until assayed for meiatonin. Plasma melatonin concentrations were directly determined in plasma by using a sheep melatonin antiserum from Guildhay Antisera (University of
Vol. 47, No. 22, 1990
Nighttime Melatonln and Physical Stress
1991
Surrey, UK, batch No. 704/6483), 3[H]melatonin tracer (spec. acty., 85 kCi/mol) from Amersham International plc (Amersham, Buchs, UK) and unlabeled melatonin from Sigma Chemical Co. (St. Louis, MO, USA). We assayed 500 pL duplicate aliquots, comparing results directly against a standard curve prepared with melatonin-fres human plasma. The reaction mixtures were incubated for 18 h at 4 °C, and free and antibody=bound fractions of 3[H]melatonin were separeted using a dextran-coated charcoal solution (2% : 0.2%). The lower detection limit of the assay was 2.5 pg/ml; the upper limit was 250 pg/ml. Intra- and inter-assay coefficients of variation were 5.1% and 9.8%, respectively. Results were expressed as mean + SEM and statistically anlyzed by twoway analysis of variance (ANOVA) with repeated measures and Student's t-test for paired data, where appropriate. Results Table I shows clinical and demographic characteristics and the MWC of each volunteer. TABLE I Clinical Data and Maximal Working Capacity (MWC) of Healthy Subjects
AGE yrs
HEIGHT cm
31 30 28 29 28 33 26
172 173 172 175 180 178 t85
MEAN _+ SD 92 176A 22 4~,
WEIGHT Kg
MWC w~t
62 65 56 72 80 86 88
220 220 150 240 240 200 160
72.7
2042 36.4
123
BASAL SBP/ POST-EXERCISE DI3P (mmHg) SBP/DBP (mmHg) 130/80 140/85 110/80 135/90 120/85 120/75 130/70
160/65 160/70 120150 180/70 140/60 140/60 160/50
126.4/80.7 102 6.7
151.4/60.7 195 8.3
BAS,N_HR beats/rain
POST-EXERCISE HR (beats/min)
72 88 84 80 70 68 64
140 160 152 160 120 140 110
75.1 89
1402 19.3
SBP = Systolic Blood Pressure; DBP = Diastolic Blood Pressure; HR = Heart Rate. Plasma melatonin levels, in resting healthy volunteers, showed a characteristic nocturnal pattern with a peak at 1.00 a.m. and a subsequent return toward low daytime levels (Fig. 1). Physical exercise, between 10.40 and 11.00 p.m., significantly blunted the nocturnal increase in plasma melatonin levels (Fig. 1). In fact, two-way ANOVA with repeated measures showed that no significant difference was present in plasma msiatonin secretion between resting and exercising conditions (F = 1.691, NS), whereas there was a significant effect for time (F = 13.262, p <0.00001) and a significant group X time interaction (F = 4.913, p <0.00001) indicating that the temporal pattern of nighttime melatonin secretion was different in the two conditions. Following exercise, plasma melatonin levels observed at 23.30, 24.00, 1.00 and 2.00 h were lower than the corresponding time point values during resting condition, with a statistical significance of p <0.02 at 23.30, p <0.008 at 24.00, P <0.01 at 1.00 and p <0.03 at 2.00 a.m. (Student's t-test for paired data).
1992
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
150 o----o
R.,,
T I1\ /
H
Z
w
I
OJr_.~, 1--12
20
,
,,
,
,
,
,
,
,
22
23
24
1
2
4
6
8
CLOCK
RG_.
HO URS
!
Plasma melatonin rhythm in seven healthy men (Mean + SEM) during rest and exercise conditions. * p <0.02, ** p <0.01, *** p <0.008, + p <0.03 (Student's t-test for paired data).
The effects of physical exercise on systolic and diastolic blood pressure and on heart rate are shown in table I1.
Discu~i_o_n The results of the present study show for the first time that, in healthy men, physical exercise at night dramatically blocks the nocturnal increase of plasma melatonin levels. Although the results of previous studies concerning the effects of stressful procedures on plasma melatonin in humans are conflicting (12-18), the main difference between these studies and the present one is that the former examined the effects of sb'essful
Vol. 47, No. 22, 1990
Nighttime Helatonin and Physical Stress
1993
TABLE II Systolic and Diastolic Blood Pressure (MEAN + SD) and Heart Rate in Healthy Subjects Before and After Nighttime Physical Exercise
CLOCK HOURS 22.00
22.30
23.00
23.30
24.00
1.00
2.00
155.8+ 245
108.3± 196
106,6+ 15.0
107.5+ 103
109.1 + 10.6
SBP
1175+_ 133
1166+_. 15.7
DBP
733± 103
775+ 11.7
61.6_+ 14.7
61.6+ 93
62,5_+ 8.8
62,5± 9.8
61.6± 9.6
HR
67.6+ 1P2
63.6+ 8.8
1243_+ 32A
70.6_+ 10,3
70.0¢ 14.0
643_+ 99
623_+ 10.1
SBP = Systolic Blood Pressure; DBP = Diastolic Blood Pressure; HR = Heart Rate.
stimuli on daytime plasma melatonin levels, when the pineal gland is quiescent. In the present experiment, instead, the physical stress was applied when the pineal gland is physiologically activated by darkness and plasma melatonin levels are increasing. Consistent with our findings, animal studies have shown that acute stress at night significantly decreases nighttime pineal melatonin content (10, 11). Moreover, Wu et al. (20) have recently reported that swimming is able to depress the elevated daytime pineal and serum melatonin levels induced by acute administration of isoproterenol in rats. These data, taken together, seem to suggest that the manner in which the pineal gland responds to stress may depend on its level of activity at the time the stress is applied. The mechanism through which physical stress at night blunts the physiological increase of plasma melatonin in humans is difficult to explain at present. In fact, the clear-cut drop of nighttime plasma melatonin levels following the physical performance may have at least three possible interpretations. First of all, an increase in melatonin catabolism may be hypothesized. The degradation of melatonin occurs primarily in the liver and, to a lesser extent, in the brain (21, 22), whereas no degradation process within the pineal gland has never been identified (23). We cannot exclude that factors such as catecholamines, glucocorticoids, free fatty acid and lactate produced in response to physical stress (24) and/or the stressinduced increase in heart rate and blood flow may accelerate melatonin degradation leading to a decrease of its levels in the blood even if the release of the indoleamine from the pineal gland is presumably increased by darkness. The second interpretation may be that stress has an inhibitory effect on dark-induced melatonin release from the epiphysis. However, a decrease of melatonin secretion from the pineal without a concomitant reduction of its synthesis seems unlikely, because this would imply that melatonin should be stored in the epiphysis, while no storage sites for
1994
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
the hormone have been identified in this gland. On the other hand, a storage of melatonin during the exercise would have been followed by a rebound increase in the plasma levels of the indoleamine, which we did not observe. Finally, a reduction in melatonin synthesis may be postulated. According to this hypothesis, one would expect that the activity of the N-acetyl-transferase (NAT), the NEdependent rate-limiting enzyme involved in melatonin production, should be reduced following nighttime physical exercise. Animal data suggest that this is not the case (10), but no data are available, at present, in humans. Anyway, it seems likely to suggest that the inhibitory effects of physical stress on the nocturnal surge of plasma melatonin in humans may be mediated by adrenocortical hormones. In fact, it has been shown that glucocorticoids decrease the NE-stimulated melatonin secretion in vitro (25) and dexamethasone reduces plasma msiatonin levels in humans (26, 27), whereas the inhibition of cortisol synthesis with the use of metirapone results in increased melatonin urinary excretion (28). Furthermore, Soszynski et al. (29) have very recently demonstrated that the nocturnal surge of plasma melatonin is absent in patients with endogenous hypercortisolaemia. Therefore, it seems possible to hypothesize that the stress-induced increase in adrenocortical activity, by acting via one of the above postulated mechanisms, may lead to a decrease in the nocturnal levels of plasma melatonin. In conclusion, our data provide the first evidence that in humans physical stress may induce a striking decrease in plasma melatonin levels if it is applied at a time when melatonin production is physiologically increasing, such as during the dark phase of the light/dark cycle. This effect resembles the suppressant effect of light on nocturnal melatonin secretion (30), and might represent a possible model to study the msiatonin production without manipulating environmental light conditions. References 1. 2.
R.J. WURTMAN, H.M. SHEIN and F. LARIN, J. Neurochem. 18, 1683-1689 (1971). A.J. LEWY, The Pineal Gand, R.M. Relkin ed., pp. 77-128, Elsevier Science Publishing Co Inc, New York (1983). 3. R.J. REITER, Ufe Sci. 40, 2119-2131 (1987). 4. J. ARENDT, M. ALDHOUS, C. BOJKOWSKI, J. ENGLISH, C. FRANEY, A.L. POULTON and D. SKENE, Advances in Pineal R_.~___arq_h;_-2, R.J. Reiter and F. Fraschini eds., pp. 223-229, John Libbey & Sons Ltd, London (1987). 5. D.C. KLEIN and A. PARFITT, Cat ech~ami_nee and__Stres_s_-Rec_ent Ad_ya_nc__es,E. Usdin, R. Kvetnansky, and T.J. Kopin eds., pp. 119-128, Pergamon Press, New York (1976). 6. M.G. TANNENBAUM, R.J. REITER, M.K. VAUGHAN, M.E. TROIANI and A. GONZALES-BRITO, J. Pineal Res. ~, 395-402 (1987). 7. Toll. CHAMPNEY, R.W. STEGER, D.S. CHRISTIE and R.J. REITER, Brain Res. 328, 25-32 (1985). 8. H.J. LINCH, M. HO and R.J. WURTMAN, J. Neural Transm. 40, 87-97 (1977). 9. J. SEGGIE, L. CAMPBELL, G.M. BROWN and L.J. GROTA, J. Pineal Res. 2, 39-49 (1985). 10. B.N. JOSHI, M.E. TROIANI, J. MILIN, F. NURNBERGER and R.J. REITER, Life Sci. 38, 1573-1580 (1986). 11. M.E. TROIANI, R.J. REITER, M.K. VAUGHAN, S. OAKNIN and G.M. VAUGHAN, Neuroendocrinology 47, 55-60 (1988). 12. D. CARR, S. REPERT, B. BULLEN, G. SKRINAR, I. BEITINS, M. ARNOLD, M. ROSENBLA'FI', J.B. MARTIN and J.W. McARTHUR, J. Clin. Endocrinol. Metab. 53,
Vol. 47, No. 22, 1990
13. 14. 15. 16. 17. 18. 19. 2O. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30.
Nighttime Melatonin and Physical Stress
1995
224-225 (1981). J. THERON, J. OOSTHUIZEN and M. TAUTENBACH, S. Afr. Med. J. 666, 838-842 (1984). H. RONKAINEN, O. VAKKURI and A. KAUPPILA, Acta Ostet. Gynecol. Scand. 65, 827-829 (1986). M. L'HERMI'I-I'E-BALERIAUX, S. CASTEELS, K. DE MEIRLEIR, L. BAEYENS and M. L'HERMITTE, J. Neural Transm. Supp121,46 (1986). R.J. STRASSMAN, O. APPENZELLER, A.J. LEWY, C.R. QUALLS and G.T. PEAKE, J. Clin. Endocrinol. Metab; 6_99,540-545 (1989). G.M. VAUGHAN, J. Neural Transm. Supp121,199-215 (1986). G. THIENTZ, U. LANG, O. DERIAZ, P. CERETELLI and P. SIZONENKO, J. Steroid Biochem 2_0, 1470 (1984). H. ASTROM and B. JONSSON, Br. Heart J. 388,289-296 (1976). W. WU, R.J. REITER, M.E. TROIANI and G.M. VAUGHAN, Life Sci. 4_!, 1473-1479, (1987). S. KVEDER and W.M. MclSAAC, J. Biol. Chem. 236, 3214-3217 (1961). A.J. FELLENBERG, G. PHILIPPOU and R.F. SEAMARK, Pineal Function, C.D. Matthews and R.F. Seamark eds., pp. 143-150, Elsevier/North Holland, Amsterdam (1981). R.J. REITER, Am. J. Anat. 162, 287-313 (1981). H. GALBO, Hormonal and Metabolic Adaptation to Exercise, Thieme and Stratton Inc.. New York (1983). M. FEUVRE-MONTAGNE, J. TOURNIAIRE, B. ESTOUR and L. BAJARD, Clin. Endocrinol. _18,175-181 (1983). L. DEMISH, K. DEMISH and T. NICKELSEN, J. Pineal Res. 5_,317-321 (1988). J. BECK-FRIIS, T. HANSSEN, B.F. KJELLMAN, J.G. LJUNGGREN, F. UNDEN and L. WETTERBERG, Psychopharmacol. Bull. 1_99,646-648 (1983). K. BRISMAR, S. WERNER, M. THOREN and L. WETTERBERG, J. Endocrinol. Invest. 8, 91-95 (1985). P. SOSZYNSKI, J. STOWlNSKA-SRZEDNICKA, A. KASPERLIK-ZATUSKA and S. ZGLICZYNSKI, Horm. Metab. Res. 21, 673-674 (1989). A.J. LEWY, T.A. WEHR, F.K. GOODWlN, D.A. NEWSOME and S.P. MARKEY, Science 21___00,1267-1269 (1980).
Pergamon Press
PHYSICAL EXERCISE AT NIGHT BLUNTS THE NOCTURNAL INCREASE OF PLASMA MELATONIN LEVELS IN HEALTHY HUMANS P. Monteleone, M. Maj, M. Fusco, C. Orazzo and D. Kemali Instituteof Medical Psychology and Psychiatry, 1st Medical School, University of Naples, Largo Madonna delle Grazie, 80138 Naples, Italy. (Received in final form September 20, 1990)
Summar_y The effects of physical exercise on nighttime melatonin secretion have never been investigated in humans. For this purpose, plasma melatonin levels were measured at different times during the day and the night in seven healthy men (aged 26-33 yrs), both in resting condition and before and alter a physical exercise performed between 10.40 and 11.00 p.m.. The exercise consisted in bicycling on a bicycle ergometer at 50% of the personal maximal work capacity (MWC) for 10 min, followed by other 10 rain of bicycling at 80% of the MWC. The results clearly showed that physical stress at night significantly blunts the nocturnal increase in plasma msiatonin levels (group X time interaction: p <0.00001 ; two-way ANOVA with repeated measures). These findings, taken together with the data of the literature, suggest that the response of the pineal gland to provocative stimuli may depend on its level of activity when the stimulus is applied. It is well established that the synthesis of the pineal indeolamine melatonin is influenced primarily by light and that norepinenephrine (NE) released from sympathetic nerve terminals within the pineal gland acts on beta- and, to a lesser degree, on alphaadrenergic receptors to stimulate melatonin production in the dark (1, 2). As a consequence, melatonin shows a characteristic circadian rhythm with low plasma levels during the day and high plasma concentrations during the night. This circadian pattern of melatonin secretion has been documented in several animal species, including man (3, 4). Although it has been argued that the sympathetic nerve endings in the pineal gland may take up circulating catecholamines protecting the pinealocytes from daytime stimulation (5), it has clearly been demonstrated that, in the rat, peripheral catecholamines released by certain stressful stimuli may promote pineal melatonin production during the day (6-9). Particularly, exposure of rats to various stressful procedures, which increase circulating catecholamines, enhances both pineal and serum melatonin levels (6-8). On the contrary, adrenalectomy blocks the increase in rat pineal melatonin content brought about by daytime physical immobilization stress and insulin-induced hypoglycemia (6, 8). When the effects of stress on melatonin have been investigated at night, opposite results have been obtained. In fact, a hind leg injection of saline into rats at night led to a dramatic decrease of pineal melatonin content (10) and swimming for 10 min at 23.10 h. depressed nighttime melatonin content in the rat epiphysis (11). However, the presence of species differences in the pineal responsiveness to provocative stressful stimuli has been demonstrated. In fact, in the 0024-3205/90 $3.00 +.00 Copyright (c) 1990 Pergamon Press plc
1990
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
Syrian hamster, insulin-induced hypoglycemia was not able to induce any rise in pineal melatonin content during the light hours (7). As regards the human beings, although some studies show that physical exercise may increase daytime plasma levels of melatonin in healthy subjects (12-16), the weight of the evidence suggests that a variety of stressful situations, some of which provoke intense responses in the sympathetic and other hormonal systems, do not induce any increase in plasma meiatonin levels during the day (17, 18). These data emphasize that the human epiphysis is not responsive to daytime sympathetic stimulation. At present, no study has been performed to evaluate the effect of stress on nighttime plasma meiatonin levels in humans. Therefore, we decided to assess the nocturnal pattern of melatonin secretion in healthy male subjects following a physical exercise applied during the dark hours. _Methods Seven healthy male subjects volunteered for the study. They were aged 26-36 yrs (mean :1: SD = 29.2:1: 2.2), had normal endocrine and metabolic functions and no family history of diabetes mellitus, and were drug-free for at least one month. All were within 15% of their ideal body weight and smoked less than 10 cigarettes per day. None of them participated regularly in any kind of sport. Only male subjects were included into the study to avoid any distortion in the response to physical exercise caused by sex differences. Prior to experimental sessions, the maximal working capacity (MWC) was determined for each subject using a bicycle ergometer (Table I). The initial work load applied was 50 watt and the load was increased by 10 watt every one min until exhaustion (19). The experimental sessions were two for each subject. In one of them, the subject underwent a submaximal work test, starting to bicycle on a work load of 50% of his own MWC for 10 min, immediately followed by 80% of MWC for another 10 rain. In the other, the subject rested supine in a bed. The two sessions were performed in random order at one week interval, on an ambulatory basis, in the months of February and March. Each subject came to our research unit at 12.00 h, when a first blood sample was collected by venipuncture. Then he was free to go away and was requested to come back at 6.00 p.m.. At 7.00 p.m., a standardized meal was eaten; at 8.00 p.m. a butterfly needle was inserted into an antecubital vein and kept patent with a slow saline infusion. Immediately after, a second blood sample was drawn, then the subject rested supine. At 9.00 p.m., light was turned off and all the subsequent procedures were carried out by the help of a red light. Further blood samples were collected at 10.00, 11.00, 11.30, 12.00 p.m. and at 1.00,2.00, 4.00, 6.00 and 8.00 a.m.. Physical exercise was started at 10.40 p.m. and, according to the above mentioned procedure, lasted until 11.00 p.m.. Blood pressure and heart rate were recorded, in the session involving physical exercise, at the same time points of blood sampling between 10.40 p.m. and 2.00 a.m.. Subjects were allowed to sleep between 11.30 p.m. and 7.00 a.m., when light was turned on. They did not smoke or drink coffee during the experimental sessions. Plasma was separated by centrifugation at 3000 rpm and stored at -20 °C until assayed for meiatonin. Plasma melatonin concentrations were directly determined in plasma by using a sheep melatonin antiserum from Guildhay Antisera (University of
Vol. 47, No. 22, 1990
Nighttime Melatonln and Physical Stress
1991
Surrey, UK, batch No. 704/6483), 3[H]melatonin tracer (spec. acty., 85 kCi/mol) from Amersham International plc (Amersham, Buchs, UK) and unlabeled melatonin from Sigma Chemical Co. (St. Louis, MO, USA). We assayed 500 pL duplicate aliquots, comparing results directly against a standard curve prepared with melatonin-fres human plasma. The reaction mixtures were incubated for 18 h at 4 °C, and free and antibody=bound fractions of 3[H]melatonin were separeted using a dextran-coated charcoal solution (2% : 0.2%). The lower detection limit of the assay was 2.5 pg/ml; the upper limit was 250 pg/ml. Intra- and inter-assay coefficients of variation were 5.1% and 9.8%, respectively. Results were expressed as mean + SEM and statistically anlyzed by twoway analysis of variance (ANOVA) with repeated measures and Student's t-test for paired data, where appropriate. Results Table I shows clinical and demographic characteristics and the MWC of each volunteer. TABLE I Clinical Data and Maximal Working Capacity (MWC) of Healthy Subjects
AGE yrs
HEIGHT cm
31 30 28 29 28 33 26
172 173 172 175 180 178 t85
MEAN _+ SD 92 176A 22 4~,
WEIGHT Kg
MWC w~t
62 65 56 72 80 86 88
220 220 150 240 240 200 160
72.7
2042 36.4
123
BASAL SBP/ POST-EXERCISE DI3P (mmHg) SBP/DBP (mmHg) 130/80 140/85 110/80 135/90 120/85 120/75 130/70
160/65 160/70 120150 180/70 140/60 140/60 160/50
126.4/80.7 102 6.7
151.4/60.7 195 8.3
BAS,N_HR beats/rain
POST-EXERCISE HR (beats/min)
72 88 84 80 70 68 64
140 160 152 160 120 140 110
75.1 89
1402 19.3
SBP = Systolic Blood Pressure; DBP = Diastolic Blood Pressure; HR = Heart Rate. Plasma melatonin levels, in resting healthy volunteers, showed a characteristic nocturnal pattern with a peak at 1.00 a.m. and a subsequent return toward low daytime levels (Fig. 1). Physical exercise, between 10.40 and 11.00 p.m., significantly blunted the nocturnal increase in plasma melatonin levels (Fig. 1). In fact, two-way ANOVA with repeated measures showed that no significant difference was present in plasma msiatonin secretion between resting and exercising conditions (F = 1.691, NS), whereas there was a significant effect for time (F = 13.262, p <0.00001) and a significant group X time interaction (F = 4.913, p <0.00001) indicating that the temporal pattern of nighttime melatonin secretion was different in the two conditions. Following exercise, plasma melatonin levels observed at 23.30, 24.00, 1.00 and 2.00 h were lower than the corresponding time point values during resting condition, with a statistical significance of p <0.02 at 23.30, p <0.008 at 24.00, P <0.01 at 1.00 and p <0.03 at 2.00 a.m. (Student's t-test for paired data).
1992
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
150 o----o
R.,,
T I1\ /
H
Z
w
I
OJr_.~, 1--12
20
,
,,
,
,
,
,
,
,
22
23
24
1
2
4
6
8
CLOCK
RG_.
HO URS
!
Plasma melatonin rhythm in seven healthy men (Mean + SEM) during rest and exercise conditions. * p <0.02, ** p <0.01, *** p <0.008, + p <0.03 (Student's t-test for paired data).
The effects of physical exercise on systolic and diastolic blood pressure and on heart rate are shown in table I1.
Discu~i_o_n The results of the present study show for the first time that, in healthy men, physical exercise at night dramatically blocks the nocturnal increase of plasma melatonin levels. Although the results of previous studies concerning the effects of stressful procedures on plasma melatonin in humans are conflicting (12-18), the main difference between these studies and the present one is that the former examined the effects of sb'essful
Vol. 47, No. 22, 1990
Nighttime Helatonin and Physical Stress
1993
TABLE II Systolic and Diastolic Blood Pressure (MEAN + SD) and Heart Rate in Healthy Subjects Before and After Nighttime Physical Exercise
CLOCK HOURS 22.00
22.30
23.00
23.30
24.00
1.00
2.00
155.8+ 245
108.3± 196
106,6+ 15.0
107.5+ 103
109.1 + 10.6
SBP
1175+_ 133
1166+_. 15.7
DBP
733± 103
775+ 11.7
61.6_+ 14.7
61.6+ 93
62,5_+ 8.8
62,5± 9.8
61.6± 9.6
HR
67.6+ 1P2
63.6+ 8.8
1243_+ 32A
70.6_+ 10,3
70.0¢ 14.0
643_+ 99
623_+ 10.1
SBP = Systolic Blood Pressure; DBP = Diastolic Blood Pressure; HR = Heart Rate.
stimuli on daytime plasma melatonin levels, when the pineal gland is quiescent. In the present experiment, instead, the physical stress was applied when the pineal gland is physiologically activated by darkness and plasma melatonin levels are increasing. Consistent with our findings, animal studies have shown that acute stress at night significantly decreases nighttime pineal melatonin content (10, 11). Moreover, Wu et al. (20) have recently reported that swimming is able to depress the elevated daytime pineal and serum melatonin levels induced by acute administration of isoproterenol in rats. These data, taken together, seem to suggest that the manner in which the pineal gland responds to stress may depend on its level of activity at the time the stress is applied. The mechanism through which physical stress at night blunts the physiological increase of plasma melatonin in humans is difficult to explain at present. In fact, the clear-cut drop of nighttime plasma melatonin levels following the physical performance may have at least three possible interpretations. First of all, an increase in melatonin catabolism may be hypothesized. The degradation of melatonin occurs primarily in the liver and, to a lesser extent, in the brain (21, 22), whereas no degradation process within the pineal gland has never been identified (23). We cannot exclude that factors such as catecholamines, glucocorticoids, free fatty acid and lactate produced in response to physical stress (24) and/or the stressinduced increase in heart rate and blood flow may accelerate melatonin degradation leading to a decrease of its levels in the blood even if the release of the indoleamine from the pineal gland is presumably increased by darkness. The second interpretation may be that stress has an inhibitory effect on dark-induced melatonin release from the epiphysis. However, a decrease of melatonin secretion from the pineal without a concomitant reduction of its synthesis seems unlikely, because this would imply that melatonin should be stored in the epiphysis, while no storage sites for
1994
Nighttime Melatonin and Physical Stress
Vol. 47, No. 22, 1990
the hormone have been identified in this gland. On the other hand, a storage of melatonin during the exercise would have been followed by a rebound increase in the plasma levels of the indoleamine, which we did not observe. Finally, a reduction in melatonin synthesis may be postulated. According to this hypothesis, one would expect that the activity of the N-acetyl-transferase (NAT), the NEdependent rate-limiting enzyme involved in melatonin production, should be reduced following nighttime physical exercise. Animal data suggest that this is not the case (10), but no data are available, at present, in humans. Anyway, it seems likely to suggest that the inhibitory effects of physical stress on the nocturnal surge of plasma melatonin in humans may be mediated by adrenocortical hormones. In fact, it has been shown that glucocorticoids decrease the NE-stimulated melatonin secretion in vitro (25) and dexamethasone reduces plasma msiatonin levels in humans (26, 27), whereas the inhibition of cortisol synthesis with the use of metirapone results in increased melatonin urinary excretion (28). Furthermore, Soszynski et al. (29) have very recently demonstrated that the nocturnal surge of plasma melatonin is absent in patients with endogenous hypercortisolaemia. Therefore, it seems possible to hypothesize that the stress-induced increase in adrenocortical activity, by acting via one of the above postulated mechanisms, may lead to a decrease in the nocturnal levels of plasma melatonin. In conclusion, our data provide the first evidence that in humans physical stress may induce a striking decrease in plasma melatonin levels if it is applied at a time when melatonin production is physiologically increasing, such as during the dark phase of the light/dark cycle. This effect resembles the suppressant effect of light on nocturnal melatonin secretion (30), and might represent a possible model to study the msiatonin production without manipulating environmental light conditions. References 1. 2.
R.J. WURTMAN, H.M. SHEIN and F. LARIN, J. Neurochem. 18, 1683-1689 (1971). A.J. LEWY, The Pineal Gand, R.M. Relkin ed., pp. 77-128, Elsevier Science Publishing Co Inc, New York (1983). 3. R.J. REITER, Ufe Sci. 40, 2119-2131 (1987). 4. J. ARENDT, M. ALDHOUS, C. BOJKOWSKI, J. ENGLISH, C. FRANEY, A.L. POULTON and D. SKENE, Advances in Pineal R_.~___arq_h;_-2, R.J. Reiter and F. Fraschini eds., pp. 223-229, John Libbey & Sons Ltd, London (1987). 5. D.C. KLEIN and A. PARFITT, Cat ech~ami_nee and__Stres_s_-Rec_ent Ad_ya_nc__es,E. Usdin, R. Kvetnansky, and T.J. Kopin eds., pp. 119-128, Pergamon Press, New York (1976). 6. M.G. TANNENBAUM, R.J. REITER, M.K. VAUGHAN, M.E. TROIANI and A. GONZALES-BRITO, J. Pineal Res. ~, 395-402 (1987). 7. Toll. CHAMPNEY, R.W. STEGER, D.S. CHRISTIE and R.J. REITER, Brain Res. 328, 25-32 (1985). 8. H.J. LINCH, M. HO and R.J. WURTMAN, J. Neural Transm. 40, 87-97 (1977). 9. J. SEGGIE, L. CAMPBELL, G.M. BROWN and L.J. GROTA, J. Pineal Res. 2, 39-49 (1985). 10. B.N. JOSHI, M.E. TROIANI, J. MILIN, F. NURNBERGER and R.J. REITER, Life Sci. 38, 1573-1580 (1986). 11. M.E. TROIANI, R.J. REITER, M.K. VAUGHAN, S. OAKNIN and G.M. VAUGHAN, Neuroendocrinology 47, 55-60 (1988). 12. D. CARR, S. REPERT, B. BULLEN, G. SKRINAR, I. BEITINS, M. ARNOLD, M. ROSENBLA'FI', J.B. MARTIN and J.W. McARTHUR, J. Clin. Endocrinol. Metab. 53,
Vol. 47, No. 22, 1990
13. 14. 15. 16. 17. 18. 19. 2O. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30.
Nighttime Melatonin and Physical Stress
1995
224-225 (1981). J. THERON, J. OOSTHUIZEN and M. TAUTENBACH, S. Afr. Med. J. 666, 838-842 (1984). H. RONKAINEN, O. VAKKURI and A. KAUPPILA, Acta Ostet. Gynecol. Scand. 65, 827-829 (1986). M. L'HERMI'I-I'E-BALERIAUX, S. CASTEELS, K. DE MEIRLEIR, L. BAEYENS and M. L'HERMITTE, J. Neural Transm. Supp121,46 (1986). R.J. STRASSMAN, O. APPENZELLER, A.J. LEWY, C.R. QUALLS and G.T. PEAKE, J. Clin. Endocrinol. Metab; 6_99,540-545 (1989). G.M. VAUGHAN, J. Neural Transm. Supp121,199-215 (1986). G. THIENTZ, U. LANG, O. DERIAZ, P. CERETELLI and P. SIZONENKO, J. Steroid Biochem 2_0, 1470 (1984). H. ASTROM and B. JONSSON, Br. Heart J. 388,289-296 (1976). W. WU, R.J. REITER, M.E. TROIANI and G.M. VAUGHAN, Life Sci. 4_!, 1473-1479, (1987). S. KVEDER and W.M. MclSAAC, J. Biol. Chem. 236, 3214-3217 (1961). A.J. FELLENBERG, G. PHILIPPOU and R.F. SEAMARK, Pineal Function, C.D. Matthews and R.F. Seamark eds., pp. 143-150, Elsevier/North Holland, Amsterdam (1981). R.J. REITER, Am. J. Anat. 162, 287-313 (1981). H. GALBO, Hormonal and Metabolic Adaptation to Exercise, Thieme and Stratton Inc.. New York (1983). M. FEUVRE-MONTAGNE, J. TOURNIAIRE, B. ESTOUR and L. BAJARD, Clin. Endocrinol. _18,175-181 (1983). L. DEMISH, K. DEMISH and T. NICKELSEN, J. Pineal Res. 5_,317-321 (1988). J. BECK-FRIIS, T. HANSSEN, B.F. KJELLMAN, J.G. LJUNGGREN, F. UNDEN and L. WETTERBERG, Psychopharmacol. Bull. 1_99,646-648 (1983). K. BRISMAR, S. WERNER, M. THOREN and L. WETTERBERG, J. Endocrinol. Invest. 8, 91-95 (1985). P. SOSZYNSKI, J. STOWlNSKA-SRZEDNICKA, A. KASPERLIK-ZATUSKA and S. ZGLICZYNSKI, Horm. Metab. Res. 21, 673-674 (1989). A.J. LEWY, T.A. WEHR, F.K. GOODWlN, D.A. NEWSOME and S.P. MARKEY, Science 21___00,1267-1269 (1980).