Endocrine and chronobiological effects of fasting in women

FERTILITY AND STERILITY威 VOL. 75, NO. 5, MAY 2001 Copyright ©2001 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A.

Endocrine and chronobiological effects of fasting in women Sarah L. Berga, M.D.,a,b Tammy L. Loucks, M.P.H.,a and Judy L. Cameron, Ph.D.b,c,d Magee-Womens Hospital and the University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania

Received August 15, 2000; revised and accepted November 13, 2000. Supported by grants from the University of Pittsburgh School of Medicine to J.L.C. (HD 26888, NS09561), to S.L.B. (RO1MH-50748), and to the General Clinical Research Center (RR00056). Presented in part at the 75th Annual Meeting of the Endocrine Society, Las Vegas, Nevada, June 9 –12, 1993. Reprint requests: Sarah L. Berga, M.D., University of Pittsburgh School of Medicine, Magee-Womens Hospital, 300 Halket Street, Pittsburgh, Pennsylvania 15213 (FAX: 412-641-1133; E-mail: sberga@mail. magee.edu). a Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh School of Medicine. b Department of Psychiatry, University of Pittsburgh School of Medicine. c Department of Neuroscience, University of Pittsburgh School of Medicine. d Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine. 0015-0282/01/$20.00 PII S0015-0282(01)01686-7

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Objective: To determine whether fasting in women would suppress GnRH/LH drive in a high- versus low-gonadal steroid milieu. Design: Case-control study. Setting: Academic clinical research center. Patient(s): Eleven eumenorrheic women and eleven women taking combined oral contraceptives. Intervention(s): Seven of the eleven women in each group underwent an acute 72-hour fast. Blood samples were obtained at 15-minute intervals for 24 hours before the fast and during the last 24 hours of fasting. Main Outcome Measure(s): Twenty-four– hour profiles of LH, cortisol, and melatonin were assessed. Ovarian activity was tracked with estradiol and progesterone levels, and metabolic responses were gauged by measuring thyroid hormone and ␤-hydroxy-butyric acid levels. Result(s): Fasting increaseed ␤-hydroxy-butyric acid and reduced free thyronine. Fasting in the midfollicular phase had no effect on LH pulsatility or on FSH, estradiol, or subsequent luteal-phase progesterone levels. However, fasting elevated cortisol and resulted in a phase advance in melatonin secretion of 81 minutes in both the midfollicular and luteal phases. Conclusion(s): Fasting in women elicited expected metabolic responses and apparently advanced the central circadian clock without compromising reproductive function. (Fertil Steril威 2001;75:926 –32. ©2001 by American Society for Reproductive Medicine.) Key Words: Fasting, ovulation, melatonin

Nutritional deprivation has been implicated as a common cause of reproductive compromise. To understand the neuroendocrine mechanisms mediating this relationship, the effects of acute calorie deprivation were investigated. In male monkeys (1) and men (2– 4), brief periods of fasting of 1 to 3 days have been shown to decrease LH pulsatility as well as decrease circulating levels of FSH and testosterone. However, when women have fasted during the follicular phase, no effects on LH pulsatility and ovarian function have been seen (5). Because fasting-induced suppression of GnRH/LH drive is attenuated in castrate male monkeys and rats (6, 7), we wondered whether the preservation of gonadotropin drive in women fasted in the follicular phase was attributable to the initiation of the fast during a period of the menstrual cycle when there is a relatively low steroid hormone milieu. If this were true, then we would expect to observe a suppression of GnRH/LH drive when women fasted in the luteal phase.

Several lines of evidence support the notion that metabolic challenge is associated with reproductive compromise in women. For instance, sustained calorie deprivation sufficient to cause weight loss leads to anovulation in previously ovulatory women (8). Moreover, undernutrition has been linked to the development of functional hypothalamic amenorrhea (FHA), a condition in which GnRH/LH drive is significantly reduced (9). Given these considerations, we sought to determine whether the effects of fasting in women differed if the fast was initiated during the follicular as compared with the luteal phase of the menstrual cycle. Fasting-induced decrements in LH pulse frequency imply that fasting disrupts GnRH neuronal drive to the reproductive axis. Women with FHA concomitantly display reduced GnRH/LH drive and amplified nocturnal melatonin secretion (10 –13) and elevated cortisol secretion (14 –16). Amplification of hypothalamic-pituitary-adrenal secretion and pineal

FIGURE 1 Schematic diagram of the experimental protocol.

Berga. Endocrine effects of fasting in women. Fertil Steril 2001.

melatonin secretion have been implicated as modulators of GnRH pulsatility. We reasoned that if increases in CRH or melatonin were causally linked to diminished central reproductive drive, then fasting would elicit an amplification of nocturnal melatonin secretion and diurnal cortisol secretion while concomitantly producing a decrease in LH pulsatility. Accordingly, we also determined the effects of fasting upon melatonin and cortisol secretion.

MATERIALS AND METHODS

taking oral contraceptives. In those women not taking oral contraceptives, ovulation was verified by a midluteal phase serum progesterone concentration of 15 nm/L or greater (mean 35.8 ⫾ 15.8 nm/L) obtained in the cycle preceding the study cycle. Ovulation was similarly assessed in the study cycle. Women already taking oral contraceptives were asked to switch for the duration of the study to a preparation containing 35 ␮g ethinyl estradiol and 1 mg norethindrone. Subjects were required to be between 95–110% of IBW as determined by the 1983 Metropolitan height and weight table for women (17).

Subjects Twenty-two women 19 – 40 years of age and at 96 –108% of ideal body weight (IBW) participated in this study. The study was approved by the institutional review boards of the University of Pittsburgh School of Medicine and MageeWomens Hospital. Informed consent was obtained from all participants prior to the initiation of the study. Eleven women were studied in the midfollicular phase, days 4 –9 from last menses, and 11 were studied during a simulated luteal phase achieved by administering 35 ␮g of ethinyl estradiol and 1 mg norethindrone orally daily for ⬎7 and ⬍39 days. We anticipated that variability in luteal length and sex steroid secretion characteristic of endogenous luteal phases would complicate the assessment of fasting. For instance, variability in luteal-phase sex steroid secretion might acutely alter the pulsatile release of LH and obscure detection of LH responses to fasting. Therefore, we sought to assess the effects of fasting during a simulated luteal phase by giving a combination oral contraceptive. In each group of 11 women, 7 were randomized after enrollment to a 72-hour fast, and 4 were not fasted but otherwise underwent the identical study paradigm. To be eligible for the study, subjects gave a history of regular menstrual intervals of 27–31 days or were currently FERTILITY & STERILITY威

Protocol Participants were admitted to the General Clinical Research Center or to the Magee-Womens Clinical Research Center by 7 A.M. on day 1 (see Fig. 1). Four subjects in each group were randomized to a nonfasting arm. Blood samples were obtained at 15-minute intervals for 24 hours on the day before the fast began (day 1) and during the last 24 hours of the fast (days 4 –5). Blood sampling began at 10 A.M. on day 1, at least 30 minutes after placement of an indwelling intravenous catheter into a forearm vein. This timepoint was chosen because melatonin levels are at a nadir at this time of day. Throughout the study, sleeping was encouraged from 11 P.M. to 7 A.M. and was not permitted at any other time. During these hours, room lights were turned off, window shades were drawn, and the intravenous tubing was extended so that blood samples could be drawn from outside the subject’s room. Before the initiation of the fast, subjects were fed standard mixed meals at standard times. After the fast began, subjects were permitted water only for 72 hours (days 2–5). During the first 36 hours of the fast, subjects were allowed to leave the clinical research center but were advised not to exercise. During the last 36 hours of the fast, subjects were readmitted to the clinical research center for ongoing obser927

vation and to obtain blood samples. After the study was completed, participants were fed and observed before discharge.

Assays LH levels were determined in duplicate by a highly sensitive immunofluorometric assay (Delfia, hLHSpec; Wallac Inc., Turko, Finland) that requires only 25 ␮L of serum per well and has a sensitivity of 0.05 IU/L. There was a coefficient of variation (CV) of 4.8% within an assay and 8.5% between assays. Plasma melatonin levels were determined in duplicate at 30-minute intervals, according to methods described elsewhere (10), using a validated radioimmunoassay with a sensitivity of 8.3 pg/mL and a within-assay CV of 4.9% and a between-assay CV of 7.1%. All samples from a given subject were run in the same assay. Serum cortisol levels were measured in duplicate by a solid phase immunoradiometric assay (Coat-a-Count; Diagnostics Products Corp., Los Angeles, CA). The within- and between-assay CVs were 3.7% and 8.9%, respectively. The cortisol assay sensitivity was 2.0 ng/mL. Serum estradiol (E2) was assessed in baseline samples in duplicate using a direct immunoradiometric assay (Coat-aCount, DPC) with a sensitivity of 8.0 pg/mL, a within-assay CV of 3.0%, and a between-assay CV of 6.9%. Free thyronine (fT3) and free thyroxine (fT4) determinations were made at hourly intervals using immunoradiometric assays (Coat-a-Count, DPC). The within-assay and between-assay CVs for fT3 and fT4 were 3.8%, 4.0%, 5.9%, and 6.3%, respectively. The sensitivity of the fT3 assay was 0.2 pg/mL, and that of the fT4 was 0.01 ng/dL. FSH was determined in duplicate at hourly intervals by immunofluorometric assay (Delfia, hFSH, Wallac Inc.). The assay sensitivity was 0.05 U/L, and the within- and between-assay CVs were 3.6% and 9.4%, respectively.

␤-hydroxy-butyric acid levels were assessed by the clinical chemistry laboratory at the University of Pittsburgh Medical Center. The method employed the Ketorex kit (Sanwa Kagaku Kenkyusho Co., Ltd., Nagoya, Japan), which entails the enzymatic conversion of ␤-hydroxy-butyric acid to acetoacetate and subsequent spectrometric quantitation. The sensitivity of the assay was 5 ␮m/L, and the within-assay CV was 4%. Progesterone levels were measured by the clinical chemistry laboratory of Magee-Womens Hospital using an immunoradiometric assay (Coat-aCount, DPC). The sensitivity of this method was 0.06 nmol/L, and the within- and between-assay CVs were less than 10%.

Data Analysis LH pulse number and amplitude were determined by a computer-assisted algorithm, Cluster (18). A peak width of two, a nadir of one, and a t statistic of 3 for upstroke and downstroke were used. Variance for each point was estimated from a quadratic equation, externally derived from serial replicates at standard doses. The following parameters 928

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Endocrine effects of fasting in women

of the nocturnal melatonin secretory profile were determined (10): onset, offset, midpoint, duration, peak value, and area under the curve from onset to offset. To analyze onset, offset, and midpoint, clock hours were transformed into circular coordinates. For ease of interpretation, however, these variables are reported in clock hours. Other outcome variables that were determined were as follows: mean LH, mean FSH, mean 24-hour cortisol level, and mean 8-hour cortisol from 8 A.M. to 3:45 P.M., 4 to 10:45 P.M., and 11 P.M. to 7:45 A.M., mean fT3, mean fT4, baseline E2 on each day of study, midluteal progesterone levels in a preceding cycle and during the cycle in which the fast occurred, and ␤-hydroxy-butyric acid levels before and during fasting. Cortisol acrophase was determined by a computerized cosinor analysis program. Outcome variables were analyzed by one-factor and by mixed, two-factor analysis of variance, as appropriate. When the analysis of variance indicated significance (P⬍.05), post hoc, within-group responses were analyzed by paired t-tests and between-group responses, by group t-tests. As there were no differences for the melatonin parameters between the follicular phase and the simulated luteal-phase groups, these groups were combined to compare the fasting versus nonfasting melatonin responses.

RESULTS There were no differences between the groups (fasted vs. fed or follicular phase vs. simulated luteal phase) in age or %IBW. Ovulatory women and those on oral contraceptives had similar ages (mean ⫾ SE: 24.8 ⫾ 1.5 years and 25.0 ⫾ 1.8 years, respectively) and %IBWs (100 ⫾ 1.2% vs. 102 ⫾ 1.5%, respectively). The mean age of the women who fasted in the follicular phase (FP-fast) was 24.0 ⫾ 2.1 years, and their %IBW was 99.4 ⫾ 1.7. The mean age for the women in the follicular phase (FP) who did not fast (FP-fed) was 26.2 ⫾ 2.0 years and %IBW was 101.0 ⫾ 1.8. The mean age of the women with the simulated luteal phase (SLP) who fasted (SLP-fast) was 23.8 ⫾ 2.7 years, and mean %IBW was 103.1 ⫾ 1.8. The mean age of the women in the simulated luteal phase who did not fast (SLP-fed) was 27.0 ⫾ 1.5 years, and mean %IBW was 100.5 ⫾ 2.9. The biochemical and endocrine responses to fasting are shown in Table 1. As expected, women who fasted while in the follicular and simulated luteal phases had consistent decrements in fT3 and profound increases in ␤-hydroxybutyric acid. Free T4 levels were unchanged during the 72-hour fast. In women in the follicular phase, progesterone concentrations in the cycle in which the fast occurred were indistinguishable from those in a preceding cycle. Estradiol levels rose similarly in women in the follicular phase, regardless of whether they fasted during the study. Gonadotropin levels differed significantly between women in the follicular phase and those taking oral contraVol. 75, No. 5, May 2001

TABLE 1 Mean biochemical and hormone levels before (Fed) and during (Fast) the last 24 hours of a 72-hour fast in four groups of women. fT3, pg/mL (⫾SE) Group FP-fast FP-fed SLP-fast SLP-fed

␤OH-BA, ␮m/L (⫾SE)

fT4, ng/dL (⫾SE)

Estradiol, pmol/L (⫾SE)

Progesterone, nmol/L (⫾SE)

Fed

Fast

Fed

Fast

Fed

Fast

Fed

Fast

Fed

Fast

1.6 (0.1) 1.5 (0.2) 1.9 (0.1) 2.0 (0.2)

0.8a (0.1) 1.6 (0.2) 1.1a (0.1) 1.9 (0.3)

0.8 (0.1) 0.8 (0.1) 0.9 (0.1) 1.0 (0.1)

0.8 (0.1) 0.9 (0.1) 0.9 (0.1) 1.0 (0.1)

28.0 (8.0) 17.3 (2.1) 67.4 (21.3) 42.0 (9.9)

3,365a (649) 13.1b (2.5) 4,803a (1,243) 63.5 (26.5)

164 (24) 110 (10) ⬍70 90.1 (20.1)

312b (61) 157 (17) ⬍70 ⬍70

34.4 (6.6) 38.1 (7.4) NA NA

33.5 (5.1) 40.1 (7.7) NA NA

Note: FP-fast ⫽ women who fasted in the follicular phase; FP-fed ⫽ women who did not fast in the follicular phase; SLP-fast ⫽ women who fasted in a simulated luteal phase; SLP-fed ⫽ women studied during a simulated luteal phase while not fasting; NA ⫽ not applicable. a Paired P⬍.01. b Paired P⬍.05. Berga. Endocrine effects of fasting in women. Fertil Steril 2001.

melatonin level was stable in those who did not fast, it was somewhat lower in those who did. In the women who fasted, the peak (mean of the three highest values) was 131.4 ⫾ 15.2 before and 123.3 ⫾ 15.3 pg/mL during the fast (P⬍.05). In those who did not fast, it was 122.5 ⫾ 26.0 and 128.4 ⫾ 22.2 pg/mL at the two times measured (P⫽.7).

ceptives. Mean FSH and LH levels were suppressed in women in the simulated luteal phase, and LH pulse frequency was so low that the LH responses to fasting could not be analyzed. As shown in Table 2, however, LH pulses of women in the follicular phase were easily detected, but LH pulse frequency and pulse amplitude were unaltered by fasting.

Cortisol concentrations rose dramatically in response to fasting in both the FP and SLP groups, as shown in Table 3. Before the initiation of the fast, mean cortisol concentrations were higher in steroid-treated women (P⬍.001), presumably because of increased binding globulins. Diurnal rhythmicity in cortisol secretion was obscured in this group, making the determination of acrophase less reliable. The acrophase of the cortisol rhythm was advanced by about 75 min during fasting in the FP group, but no change was detected in the SLP group.

The circadian profile of melatonin in women who fasted and those who did not is shown in Figure 2. The nocturnal duration of melatonin secretion was extended by 90 minutes in those women who fasted, from a mean of 10.5 ⫾ 0.4 hours to 12.0 ⫾ 0.4 hours (P⬍.005). The duration was unaltered in those who did not fast, 11.6 ⫾ 0.2 versus 12.1 ⫾ 0.8 hours (P⫽.5). The increase in duration in the women who fasted was entirely due to an 81-minute advance in the onset of nocturnal melatonin secretion from 2243 ⫾ 0.17 to 2122 ⫾ 0.50 clock hours (P⬍.004). Onset of nocturnal melatonin secretion in the women who did not fast was unaltered: onset occurred at 2100 ⫾ 0.15 versus 2022 ⫾ 0.58 clock hours (P⫽.3). Offset of nocturnal melatonin secretion was stable in both fasted and nonfasted subjects. Midpoint was advanced by approximately 38 min in those who fasted (P⬍.01) and was stable in those who did not (P⫽.4). Although the peak

DISCUSSION Undernutrition presents a metabolic challenge that induces a constellation of endocrine and neuroendocrine responses designed to promote adaptation and preserve homeostasis. In some instances, metabolic challenge also may

TABLE 2 Mean (⫾SE) gonadotropin levels and LH pulse characteristics before (Fed) and during (Fast) a 72-hour fast. LH pulse No. per 24 h (⫾SE)

Mean LH, IU/L (⫾SE)

Amplitude, IU/L (⫾SE)

Mean FSH, IU/L (⫾SE)

Group

Fed

Fast

Fed

Fast

Fed

Fast

Fed

Fast

FP-fast FP-fed

15.3 (0.8) 16.2 (0.7)

14.7 (1.0) 13.8 (1.8)

1.7 (0.2) 2.1 (0.5)

3.6 (1.6) 2.2 (0.7)

4.2 (0.4) 4.6 (0.9)

7.4 (3.3) 5.6 (1.7)

4.5 (0.5) 5.6 (0.6)

4.8 (0.7) 4.9 (0.8)

Note. FP-fast ⫽ women who fasted in the follicular phase; FP-fed ⫽ women who did not fast in the follicular phase. Berga. Endocrine effects of fasting in women. Fertil Steril 2001.

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FIGURE 2 Mean ⫾ SE of melatonin concentrations (picograms per milliliter) in 14 women before (open circles) and during (closed circles) the last 24 hours of a 72-hour fast (top panel) and in 8 women who were studied on separate days but who did not fast (bottom panel). Note the phase advance of melatonin onset in those who fasted as compared with the nearly identical circadian melatonin profiles in those who did not fast.

Berga. Endocrine effects of fasting in women. Fertil Steril 2001.

engender reproductive compromise. Fasting has been utilized as an experimental paradigm for elucidating the mechanisms and conditions linking undernutrition and reproductive compromise. In men, a 48-hour fast elicited a 50% decline in LH pulse frequency and a 20% decline in testosterone secretion (2). In contrast, in the present study, a 72-hour fast in the midfollicular phase did not suppress central reproductive drive and ovulation appeared to be unperturbed. Given the other endocrine responses that fast930

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Endocrine effects of fasting in women

ing provoked in these same women, this is surprising. Other investigators, too, have found similar results in women when fasting was undertaken in the follicular phase (5, 19). On the other hand, Loucks et al. observed a small but documentable decrease in LH pulse frequency in response to 5 days of subtotal calorie deprivation initiated during the midfollicular phase, but the impact of this decline in GnRH/LH drive upon follicular performance was not monitored (20). In another study, Alvero et al. demonstrated that a 72-hour fast supVol. 75, No. 5, May 2001

TABLE 3 Mean ⫾ SE integrated cortisol levels (ng/mL) and cortisol acrophase (in circadian hours) before (B) and during (D) a 72-h fast in four groups of women. 0800–0730 h, ng/mL (⫾SE) Group FP-fast FP-fed SLP-fast SLP-fed

Fed 83.9 (6.7) 76.3 (4.9) 155.2 (13.0) 212.2 (42.9)

Fast

0800–1530 h, ng/mL (⫾SE) Fed

1600–2330 h, ng/mL (⫾SE)

Fast

118.8a (8.6) 118.0 (12.6) 132.3 (16.1) 75.1 (2.7) 98.2 (4.4) 97.9 (3.3) 244.6a (17.9) 227.4 (29.3) 276.8a (24.4) 212.3 (31.3) 285.5 (57.0) 278.6 (33.7)

Fed

Fast

59.1 (3.7) 52.1 (7.0) 108.7 (10.9) 136.0 (37.3)

98.7a (9.5) 50.5 (4.5) 221.0a (16.6) 129.2 (30.1)

2400–0730 h, ng/mL (⫾SE) Fed

Fast

71.4 (11.2) 124.0a (7.5) 75.5 (7.0) 75.5 (6.7) 146.0 (24.1) 234.9a (18.8) 214.5 (36.4) 225.7 (31.2)

Acrophase, ng/mL (⫾SE) Fed

Fast

10.2 (0.7) 9.9 (0.6) 10.0 (0.6) 9.2 (0.4)

9.0b (0.9) 9.9 (0.4) 10.3 (0.7) 9.3 (0.3)

Note: FP-fast ⫽ women who fasted in the follicular phase; FP-fed ⫽ women who did not fast in the follicular phase; SLP-fast ⫽ women who fasted in a simulated luteal phase; SLP-fed ⫽ women studied during a simulated luteal phase while not fasting; NA ⫽ not applicable. a Paired P⬍.01. b Paired P⬍.05. Berga. Endocrine effects of fasting in women. Fertil Steril 2001.

pressed LH pulse frequency by about 20% in lean (% body fat ⬍20) but not normal-weight, eumenorrheic women (21). The fasting-induced decline in GnRH/LH drive in lean women either lengthened the follicular phase or abrogated ovulation. Overall, the GnRH pulse generator of normal weight women seems relatively resistant to the suppressive effects of fasting if initiated in the relatively low steroid milieu of the follicular phase. Preexisting characteristics, such as low body weight, heighten sensitivity to the endocrine effects of undernutrition. Despite our aim, we were unable to determine whether women are more sensitive to the effects of fasting in the luteal as opposed to the follicular phase. The relatively greater sensitivity of the GnRH pulse generator of males to fasting has been attributed to sensitizing effects of gonadal steroids (7). Similarly, in female monkeys, hypoglycemic stress interrupted the GnRH pulse generator in intact but not ovariectomized monkeys (22). However, the impact of a nonnutritional stress, endotoxin administration, upon reproductive function was greater in monkeys challenged in the follicular (23) as compared with in the luteal phase (24). Taken together, these and other data suggest that the central pathways mediating the interaction between challenge and reproductive function are specific to the type of stressor. Further, whether sex steroids enhance or ameliorate the impact of a given stressor upon reproductive function also varies according to stressor type (25). In all paradigms, the molecular mechanisms mediating the relative resistance (or sensitivity) of the reproductive axis to stress remain unknown, although sex steroids do impact upon a multitude of central neuronal systems capable of modulating GnRH function and thus likely gate neuroendocrine reactivity. In this study, women who fasted developed endocrine alterations similar to those seen in women with FHA (10, 14 –16). Similarities included decreased thyronine levels, increased cortisol secretion, and amplified nocturnal secreFERTILITY & STERILITY威

tion of melatonin. Any or all of these endocrine alterations have been hypothesized to modulate central input to the reproductive axis. One important difference between eumenorrheic women who are fasted and women with FHA, however, is that the experimental fasting paradigm employed an acute challenge, whereas the triggers and behaviors linked to the development of FHA are both multiple and chronic in nature (25). Women with FHA report a combination of metabolic and psychogenic stresses and rarely display only undernutrition. An unexpected finding of this study was that a brief fast induced a phase advance in the diurnal rhythms of melatonin and cortisol. Although appetite is diurnally driven, it has not been previously shown that metabolic challenge alters the body’s clock outputs. Although the magnitude of the phase advance might seem modest, it is important to remember that the most potent zeitgeber is day length. Remarkably, the phase advance we observed occurred despite a stable light– dark signal. Further, none of the subjects who were fed displayed any change in circadian phase position. Although it is recommended that international travelers reduce caloric intake when flying east, the rationale for this recommendation has not been substantiated. The present data would buttress the claim that caloric deprivation can phase advance the human clock. Previous studies of the impact of fasting upon melatonin secretion have been discordant, but none observed a phase advance (27–29). However, in none of these studies were melatonin levels characterized so carefully. It would be of interest to know whether other metabolic challenges, such as exercise, also induce a similar phase advance. Interestingly, in this study, the shift in cortisol acrophase during fasting in the follicular and simulated luteal phases was discordant, whereas the shift in melatonin onset and midpoint to fasting in the two phases was concordant. This 931

concordance confirms that melatonin profiles are a more reliable marker of circadian phase position than cortisol profiles because they are less masked by other factors. In this setting, the circadian rhythm of cortisol was likely masked by the steroid-induced concomitant increase in cortisol-binding globulin and total cortisol concentrations. If the melatonin profiles are taken as the best estimate of the central clock, then one can conclude that fasting advances the central clock comparably in both a follicular and luteal milieu. In summary, brief periods of undernutrition initiated in women in the midfollicular phase caused the expected suppression of thyronine and rises in cortisol and ␤-hydroxybutyric acid but no documentable decline in central drive to the reproductive axis. The basis for the relative resistance of the GnRH pulse generator of women to acute nutritional deprivation remains unexplained, but it is not attributable to a lack of metabolic perturbation. A recent study suggested that under conditions of matched glycemia, there was sexual dimorphism in the autonomic, metabolic, and cardiovascular responses to prolonged submaximal exercise in healthy men and women, with men displaying greater autonomic and cardiovascular responses and women manifesting greater lipolytic and ketogenic responses (30). The present observations do not obviate an effect of more prolonged nutritional deprivation upon reproductive function in women, but they do suggest that the GnRH pulse generator of men is more sensitive to acute caloric deprivation than that of women. An unexpected finding was that circadian phase position was advanced by fasting in women. The impact of other metabolic challenges such as exercise or sleep deprivation upon circadian outputs merits investigation because simple lifestyle changes may afford a convenient, economical, and relatively safe strategy for coping with jet lag. It remains to be determined, however, whether fasting will induce a comparable phase advance in men or whether strategies for coping with jet lag will need to be gender specific.

6. 7.

8. 9. 10. 11.

12.

13. 14. 15. 16. 17. 18. 19.

20.

21. 22.

23.

24. Acknowledgments: The investigators thank the nurses of the General Clinical Research Center, Anne McGeary, and Chrissie Contis for technical assistance.

25.

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Vol. 75, No. 5, May 2001