ASSOCIATION OF THE ANTIDIABETIC EFFECTS OF BROMOCRIPTINE WITH A SHIFT IN THE DAILY RHYTHM OF MONOAMINE METABOLISM WITHIN THE SUPRACHIASMATIC NUCLEI OF THE SYRIAN HAMSTER

Bromocriptine, a dopamine D₂ agonist, inhibits seasonal fattening and improves seasonal insulin resistance in Syrian hamsters. Alterations in daily rhythms of neuroendocrine activities are involved in the regulation of seasonal metabolic changes. Changes in circadian neuroendocrine activities that regulate metabolism are believed to be modulated by central circadian oscillators within the hypothalamic suprachiasmatic nuclei (SCN) of seasonal animals. We examined the association of metabolic responses to bromocriptine with its effects on the daily rhythms of metabolic hormones and daily monoamine profiles within the SCN, a primary circadian pacemaker known to regulate metabolism, in Syrian hamsters. Obese glucose-intolerant male Syrian hamsters (body weight [BW] 185 ± 10 g) held on 14h daily photoperiods were treated at light onset with bromocriptine (800 μg/animal/day, ip) or vehicle for 2 weeks. Animals were then subjected to a glucose tolerance test (GTT) (3 g/kg BW, ip). Different subsets of animals (n = 6) from each treatment group were sacrificed at 0h/24h, 5h, 10h, 15h, or 20h after light onset for analyses of SCN monoamines, plasma insulin, prolactin, cortisol, thyroxin (T₄), triiodothyronine (T₃), glucose, and free fatty acids (FFAs). Compared with control values, bromocriptine treatment significantly reduced weight gain (14.9 vs. ?2.9 g, p <. 01) and the areas under the GTT glucose and insulin curves by 29% and 48%, respectively (p <. 05). Basal plasma insulin concentration was markedly reduced throughout the day in bromocriptine-treated animals without influencing plasma glucose levels. Bromocriptine reduced the daily peak in FFA by 26% during the late light span(p <. 05). Bromocriptine significantly shifted the daily plasma cortisol peak from the early dark to the light period of the day, reduced the plasma prolactin (mean 1.8 vs. 39.4 ng/dL) and T₄ throughout the day (mean 1.6 vs. 3.8 μg/dL), and selectively reduced T₃ during the dark period of the day (p <. 01). Concurrently, bromocriptine treatment significantly reduced SCN dopamine turnover during the light period and shifted daily peaks of SCN serotonin and 5-hydroxy-indoleacetic acid (5-HIAA) content by 12h from the light to the dark period of the day (p <. 05). This was confirmed by a further in vivo microdialysis study in which bromocriptine increased SCN extracellular 5-HIAA of glucose-intolerant hamsters during the dark phase (47% increase, p <. 05) toward levels observed in normal glucose-tolerant hamsters. Thus, bromocriptine-induced resetting of daily patterns of SCN neurotransmitter metabolism is associated with the effects of bromocriptine on attenuation of the obese insulin-resistant and glucose-intolerant condition. A large body of corroborating evidence suggests that such bromocriptine-induced changes in SCN monoamine metabolism may be functional in its effects on metabolism. (Chronobiology International, 17(2), 155–172, 2000)     

Circadian and Seasonal Variations of Plasma Insulin and Cortisol Concentrations in the Syrian Hamster, Mesocricetus Auratus

Circadian rhythms of plasma insulin, Cortisol, and glucose concentrations were examined in scotosensitive (reproductively sensitive to inhibitory effects of short daylengths) and scotorefractory male and female Syrian hamsters (Mesocricetus auratus) maintained on short (LD 10:14) and long (LD 14:10) daylengths. The baseline concentration (mean of all values obtained every 4 hr six times of day) of insulin was much greater in female than in male scotosensitive hamsters kept on short daylengths. These differences in insulin concentration may account for the observed heavy fat stores in female and low fat stores in male scotosensitive hamsters kept on short daylengths. The baseline concentrations of Cortisol were approximately equal in both scotosensitive and scotorefractory males held on short and long daylengths, but were relatively low in females held on short daylengths and especially high in scotorefractory females held on long daylengths.

The plasma concentrations of both cortisol and insulin varied throughout the day in many of the groups tested. However, the variations were not equivalent. The circadian variations of cortisol were similar irrespective of sex, seasonal condition and daylength. Peak concentrations generally occurred about 12 hr after light onset. In contrast, the circadian variations of insulin differed markedly. For example in male hamsters, robust daily variations were found in scotosensitive hamsters held on short daylengths but not on long daylengths and in scotorefractory hamsters held on long daylengths but not on short daylengths. Furthermore, the daily peak occurred during the light in the scotosensitive hamsters and during the dark in the scotorefractory animals. Neither the daily feeding pattern (about 60% consumed during dark) nor the daily variations of glucose concentration varied appreciably with seasonal condition or daylength. They do not appear to determine nor directly reflect the variations in cortisol and glucose concentrations. It is postulated that the daily rhythms of cortisol and insulin are regulated by different neural pacemaker systems and that changes in the phase relations of circadian systems account in part for seasonal changes in body fat stores.     

Reductions of body fat stores and total plasma cholesterol and triglyceride concentrations in several species by bromocriptine treatment

Administrations (injections and in feed) of bromocriptine, a dopamine agonist that inhibits prolactin secretion, reduced body fat stores and plasma total cholesterol and triglyceride concentrations in several rodent species (Syrian hamsters, Djungarian hamsters, Swiss Webster mice and obese Zucker rats). Body fat indices were reduced by at least 50% in the hamsters and mice within 10-15 days of treatment and by 29% in 8 weeks in the rats. Bromocriptine reduced total plasma cholesterol concentration by 17% in Syrian hamsters, 41% in mice and 30% in rats fasted before blood sampling. In nonfasted obese rats, bromocriptine dramatically reduced both cholesterol (from 440 to 130 mg/dl) and triglyceride (from 1570 to 540 mg/dl) levels compared with controls. These findings offer further evidence for a primary role of prolactin in lipid metabolism and indicate that bromocriptine may be useful for treating obesity and lipid-based cardiovascular disorders.     

Circadian rhythms of photorefractory siberian hamsters remain responsive to melatonin

Short day lengths increase the duration of nocturnal melatonin (Mel) secretion, which induces the winter phenotype in Siberian hamsters. After several months of continued exposure to short days, hamsters spontaneously revert to the spring-summer phenotype. This transition has been attributed to the development of refractoriness of Mel-binding tissues, including the suprachiasmatic nucleus (SCN), to long-duration Mel signals. The SCN of Siberian hamsters is required for the seasonal response to winter-like Mel signals, and becomes refractory to previously effective long-duration Mel signals restricted to this area. Acute Mel treatment phase shifts circadian locomotor rhythms of photosensitive Siberian hamsters, presumably by affecting circadian oscillators in the SCN. We tested whether seasonal refractoriness of the SCN to long-duration Mel signals also renders the circadian system of Siberian hamsters unresponsive to Mel. Males manifesting free-running circadian rhythms in constant dim red light were injected with Mel or vehicle for 5 days on a 23.5-h T-cycle beginning at circadian time 10. Mel injections caused significantly larger phase advances in activity onset than did the saline vehicle, but the magnitude of phase shifts to Mel did not differ between photorefractory and photosensitive hamsters. Similarly, when entrained to a 16-h light/8-h dark photocycle, photorefractory and photosensitive hamsters did not differ in their response to Mel injected 4 h before the onset of the dark phase. Activity onset in Mel-injected hamsters was masked by light but was revealed to be significantly earlier than in vehicle-injected hamsters upon transfer to constant dim red light. The acute effects of melatonin on circadian behavioral rhythms are preserved in photorefractory hamsters.     

Circadian Changes in Anxiolysis-Related Behavior of Syrian Hamsters. Correlation with Hypothalamic GABA Release

The diurnal variations in anxiolysis and exploratory behavior were examined in a plus-maze paradigm in Syrian hamsters exposed to 14 h light: 10 h dark photoperiods or to constant darkness for 3 days. The percent of time spent in open arms and the percent of entries to open arms (both indexes of anxiety-related behavior) as well as the number of crosses to both arms (an index of locomotor behavior) showed significant daily variations under the two lighting conditions, maxima being found at night (2400-0400 h). Flumazenil (5 mg/kg) injected at the middle of the light (at 1600 h) or dark period (at 0400 h) decreased by 39-54 % anxiolysis-related behavior without affecting locomotor activity significantly. 3 H-?-aminobutyric acid (GABA) release from preoptic-medial basal hypothalamic explants obtained from hamsters exposed to 14 h light: 10 h dark photoperiods attained maximal values at 2400-0400 h. The results further supported the existence of circadian changes in anxiolysis-related behavior in Syrian hamsters that correlated with an increased hypothalamic GABA release.     

c-Fos expression in the brains of behaviorally "split" hamsters in constant light: calling attention to a dorsolateral region of the suprachiasmatic nucleus and the medial division of the lateral habenula

"Splitting" of circadian activity rhythms in Syrian hamsters maintained in constant light appears to be the consequence of a reorganized SCN, with left and right halves oscillating in antiphase; in split hamsters, high mRNA levels characteristic of day and night are simultaneously expressed on opposite sides of the paired SCN. To visualize the splitting phenomenon at a cellular level, immunohistochemical c-Fos protein expression in the SCN and brains of split hamsters was analyzed. One side of the split SCN exhibited relatively high c-Fos levels, in a pattern resembling that seen in normal, unsplit hamsters during subjective day in constant darkness; the opposite side was labeled only within a central-dorsolateral area of the caudal SCN, in a region that likely coincides with a photo-responsive, glutamate receptor antagonist-insensitive, pERK-expressing cluster of cells previously identified by other laboratories. Outside the SCN, visual inspection revealed an obvious left-right asymmetry of c-Fos expression in the medial preoptic nucleus and subparaventricular zone of split hamsters killed during the inactive phase and in the medial division of the lateral habenula during the active phase (when the hamsters were running in their wheels). Roles for the dorsolateral SCN and the mediolateral habenula in circadian timekeeping are not yet understood.     

Light-induced c-Fos expression in the SCN and behavioural phase shifts of Djungarian hamsters with a delayed activity onset

C-Fos expression in the suprachiasmatic nucleus (SCN) and phase shifts of the activity rhythm following photic stimulation were investigated in Djungarian hamsters (Phodopus sungorus) of two different circadian phenotypes. Wild-type (WT) hamsters display robust daily patterns of locomotor activity according to the light/dark conditions. Hamsters of the DAO (delayed activity onset) phenotype, however, progressively delay the activity onset, whereas activity offset remains coupled to “light-on”. Although the exact reason for the delayed activity onset is not yet clarified, it is connected with a disturbed interaction between the light/dark cycle and the circadian clock. The aim was to test the link between photoreception and the behavioral output of the circadian system in hamsters of both phenotypes, to get further insight in the underlying mechanism of the DAO phenomenon. Animals were exposed to short light pulses at different times during the dark period to analyze phase shifts of the activity rhythm and expression of Fos protein in the SCN. The results indicate that the photosensitive phase in DAO hamsters is shifted like the activity onset. Also, phase shifts were significantly smaller in DAO hamsters. At the same time, levels of Fos expression did not differ between phenotypes regarding the circadian phase. The results provide evidence that the shifted photosensitivity of the circadian system in DAO hamsters does not differ from that of WT animals, and lead us to conclude that processes within the SCN that enable light information to reset the circadian pacemaker might offer an explanation for the DAO phenomenon.     

Activation of dopamine D2 receptors simultaneously ameliorates various metabolic features of obese women

The metabolic syndrome comprises a cluster of metabolic anomalies including insulin resistance, abdominal obesity, dyslipidemia, and hypertension. Previous studies suggest that impaired dopamine D2 receptor (D2R) signaling is involved in its pathogenesis. We studied the acute effects of bromocriptine (a D2R agonist) on energy metabolism in obese women; body weight and caloric intake remained constant. Eighteen healthy, obese women (BMI 33.2 +/- 0.6 kg/m(2), mean age 37.5 +/- 1.7, range 22-51 yr) were studied twice in the follicular phase of their menstrual cycle in a prospective, single-blind, crossover design. Subjects received both placebo (P; always first occasion) and bromocriptine (B; always second occasion) on separate occasions for 8 days. At each occasion blood glucose and insulin were assessed every 10 min for 24 h, and circadian plasma free fatty acid (FFA) and triglyceride (TG) levels were measured hourly. Fuel oxidation was determined by indirect calorimetry. Body weight and composition were not affected by the drug. Mean 24-h blood glucose (P < 0.01) and insulin (P < 0.01) were significantly reduced by bromocriptine, whereas mean 24 h FFA levels were increased (P < 0.01), suggesting that lipolysis was stimulated. Bromocriptine increased oxygen consumption (P = 0.03) and resting energy expenditure (by 50 kcal/day, P = 0.03). Systolic blood pressure was significantly reduced by bromocriptine. Thus these results imply that short-term bromocriptine treatment ameliorates various components of the metabolic syndrome while it shifts energy balance away from lipogenesis in obese humans.     

Involvement of p38 mitogen-activated protein kinase activation in bromocriptine-induced apoptosis in rat pituitary GH3 cells

Bromocriptine, a dopamine D(2) receptor agonist, is a therapeutic agent for patients with prolactinoma and hyperprolactinemia. In this study we demonstrated that bromocriptine induced activation of p38 mitogen-activated protein (MAP) kinase, with concomitant induction of apoptosis in rat pituitary adenoma cell line GH3 cells. Treatment of GH3 cells for 48 h with bromocriptine increased the p38 MAP kinase activity up to 3- to 5-fold and simultaneously increased the number of apoptotic cells. Inclusion in the medium of SB212090 or SB203580, specific p38 MAP kinase inhibitors, completely abolished the bromocriptine-induced activation of p38 MAP kinase and significantly reduced the number of apoptotic cells. The bromocriptine-induced p38 MAP kinase activation was not prevented by S(-)-eticropride hydrochloride, a specific D(2) receptor antagonist. Treatment with either epidermal growth factor (EGF) or thyrotropin-releasing hormone (TRH), which stimulates p44/42 MAP kinase, rescued cells from the bromocriptine-induced apoptosis, with concomitant inhibition of the bromocriptine-induced p38 MAP kinase activation. These results suggest that bromocriptine induces apoptosis in association with p38 MAP kinase activation, and that the p44/42 MAP kinase signaling through EGF and TRH receptors has an opposing effect on p38 MAP kinase activation as well as on apoptosis induced with bromocriptine in GH3 cells.     

Exotic photoperiods induce and entrain split circadian activity rhythms in hamsters

The split circadian activity rhythm that emerges in hamsters after prolonged exposure to constant light has been a theoretical cornerstone of a multioscillator view of the mammalian circadian pacemaker. The present study demonstrates a novel method for splitting hamster circadian rhythms and entraining them to exotic light:dark cycles. Male Syrian hamsters previously maintained on a 14-h day and 10-h night were exposed to a second 5-h dark phase in the afternoon. The 10-h night was progressively shortened until animals experienced two 5-h dark phases beginning 10 h apart. Most hamsters responded by splitting their activity rhythms into two components associated with the afternoon and nighttime dark phases, respectively. Each activity component was entrained to this light:dark:light:dark cycle. Transfer of split hamsters to constant darkness resulted in rapid joining of the two activity components with the afternoon component associated with onset of the fused rhythm. In constant light, the nighttime component corresponded to activity onset of the fused rhythm, but splitting emerged again at an interval characteristic for this species. The results place constraints on multi-oscillator models of circadian rhythms and offer opportunities to characterize the properties of constituent circadian oscillators and their interactions.     

Resetting the Annual Cycle with Timed Daily Injections of 5-Hydroxytryptophan and L-Dihydroxyphenylalanine in Syrian Hamsters

Daily injections of dihydroxyphenylalanine (DOPA), a precursor for dopamine synthesis, given 12 hr after daily injections of S-hydroxytryptophan (5-HTP), a serotonin precursor, induced uterine growth and increased serum thyroxine and luteinizing hormone (LH) concentrations in scotosensitive female hamsters maintained on short day lengths. On the other hand, daily injections of DOPA given at the same time as daily injections of S-HTP reduced uterine weights and serum concentrations of thyroxine and LH in scotorefractory female hamsters. These results indicate that the endogenous mechanism controlling seasonality (scotosensitivity and scotorefractoriness) in the Syrian hamster may be reset by drugs that influence serotonergic and catecholaminergic activity. This seasonal mechanism might involve two circadian systems that undergo seasonal changes in their phase relations.     

Effects of food deprivation on locomotor activity, plasma glucose, and circadian clock resetting in Syrian hamsters

Circadian rhythms in Syrian hamsters can be phase advanced by activity or arousal stimulated during the daily rest phase ("subjective day"). A widely used method for stimulating activity is confinement to a novel wheel. Some hamsters decline to run, and some procedures may reduce the probability of running. The authors evaluated food deprivation (FD) as a method to promote running. Given evidence that perturbations of cell metabolism or glucose availability may affect circadian clock function in some tissues or species, they also assessed the effects of FD on free-running circadian phase, resetting responses to photic and nonphotic stimuli and plasma glucose. In constant light, a 27-h fast significantly increased running in a novel wheel and marginally increased the average size of resulting phase shifts. FD, without novel wheel confinement, was associated with some very large phase shifts or disruption of rhythmicity in hamsters that spontaneously ran in their home wheels during the subjective day. Hamsters that ran only during the usual active phase (subjective night) or that were prevented from running did not exhibit phase shifts, despite refeeding in the mid-subjective day. Using an Aschoff Type II design for measuring shifts, a 27-h fast significantly increased the number of hamsters that ran continuously when confined to a novel wheel but did not affect the dose-response relation between the amount of running and the size of the resulting shift. A day of fasting also did not affect the size of phase delay or advance shifts to 30-min light pulses in the subjective night. Plasma glucose was markedly reduced by wheel running in combination with fasting but was increased by running in nonfasted hamsters. These results establish FD as a useful tool for stimulating activity in home cage or novel wheels and indicate that in Syrian hamsters, significant alterations in glucose availability, associated with running, fasting, and refeeding, have surprisingly little effect on circadian pacemaker function.     

Bromocriptine prevents the castration-induced rise in porphyrin concentration in the harderian glands of the male Syrian hamster, Mesocricetus auratus

The Harderian glands in Syrian hamsters exhibit a striking sexual dimorphism. Male Harderian glands show two cell types and low levels of porphyrins and melatonin. Of the enzymes involved in the synthesis of melatonin, N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT) show high and low activity levels, respectively. Female Harderian glands show but one cell type and have high porphyrin and melatonin levels, low NAT activity, and high HIOMT activity. In castrated males, the Harderian glands exhibit a female pattern of morphology, porphyrin levels, and indoleamine metabolism. In an attempt to determine whether prolactin in involved in this sexually dimorphic response of the Harderian glands, intact and castrated male and intact female hamsters were injected daily with 500 micrograms of bromocriptine, a dopamine agonist. Bromocriptine led to reduced serum prolactin levels in all groups. It had no apparent effect on the Harderian glands of intact males. In contrast, in castrated males bromocriptine prevented the postcastrational rise in porphyrin levels but had no effect on NAT or HIOMT activities. In females, bromocriptine treatment had no effect on porphyrin concentrations or HIOMT activity; it led to a statistically significant increase in NAT activity. We propose that testosterone inhibits Harderian porphyrin synthesis while dopamine or prolactin stimulates it.     

Dim nocturnal illumination alters coupling of circadian pacemakers in Siberian hamsters,<Emphasis Type="Italic"> Phodopus sungorus</Emphasis>

The circadian pacemaker of mammals comprises multiple oscillators that may adopt different phase relationships to determine properties of the coupled system. The effect of nocturnal illumination comparable to dim moonlight was assessed in male Siberian hamsters exposed to two re-entrainment paradigms believed to require changes in the phase relationship of underlying component oscillators. In experiment 1, hamsters were exposed to a 24-h light-dark-light-dark cycle previously shown to split circadian rhythms into two components such that activity is divided between the two daily dark periods. Hamsters exposed to dim illumination (<0.020 lx) during each scotophase were more likely to exhibit split rhythms compared to hamsters exposed to completely dark scotophases. In experiment 2, hamsters were transferred to winter photoperiods (10 h light, 14 h dark) from two different longer daylengths (14 h or 18 h light daily) in the presence or absence of dim nighttime lighting. Dim nocturnal illumination markedly accelerated adoption of the winter phenotype as reflected in the expansion of activity duration, gonadal regression and weight loss. The two experiments demonstrate substantial efficacy of light intensities generally viewed as below the threshold of circadian systems. Light may act on oscillator coupling through rod-dependent mechanisms.Abbreviations activity duration - DD constant dark or dim - E evening oscillator - ETV estimated testis volume - LDLD light-dark-light-dark cycle - LED light emitting diode - M morning oscillator - SCN suprachiasmatic nuclei - free-running period     

Entrainment of 2 subjective nights by daily light:dark:light:dark cycles in 3 rodent species

Recent work with exotic 24-h light:dark:light:dark (LDLD) cycles indicates surprising flexibility in the entrainment patterns of Syrian hamsters. Following exposure to an LDLD cycle, hamsters may adopt a form of rhythm splitting in which markers of subjective night (e.g., activity, melatonin) are expressed in each of the twice daily scotophases. This pattern contrasts markedly with that of conventionally entrained hamsters in which markers of subjective night are expressed once daily in only 1 of the 2 dark periods. The "split" entrainment pattern was examined further here in Syrian and Siberian hamsters and in mice exposed to LDLD 7:5:7:5, a condition that reliably induces split activity rhythms in all 3 species. The phase angle of entrainment and activity duration were generally similar comparing the 2 daily activity bouts in each species. The stability of this split entrainment state was assessed by deletions of photophases on individual days, by exposure to skeleton photoperiods, and by transfer to constant darkness. As in Syrian hamsters, the one-time substitution of darkness for one 7-h photophase did not grossly alter activity patterns of Siberian hamsters but acutely disrupted the split rhythms of mice. Skeleton light pulses of progressively shorter duration did not significantly alter split entrainment patterns of either Syrian or Siberian hamsters. Both species continued to exhibit stable entrainment with activity expressed in alternate scotophases of an LD 1:5 cycle presented 4 times daily. In contrast, the split activity rhythms of mice were not maintained under skeleton pulses. In constant darkness, rhythms of Siberian hamsters remained distinctly split for a minimum of 2 cycles. Split entrainment to these novel LDLD and 4-pulse skeleton lighting regimes demonstrates a marked degree of plasticity common to the circadian systems of several rodent species and identifies novel entrainment patterns that may be reliably elicited with simple environmental manipulations. Inter- and intraspecific differences in the stability of split activity rhythms likely reflect differences in coupling interactions between the component circadian oscillators, which, adopting separate phase relations to these novel LD cycles, yield a split entrainment pattern.     

Extracellular nitric oxide signaling in the hamster biological clock

Nocturnal light pulses induce phase shifts in circadian rhythms and activate cFos expression in the suprachiasmatic nuclei (SCN). We have studied the role of nitric oxide (NO) in the intercellular communication within the dorsal and ventral portions of the SCN in Syrian hamsters. Administration of the NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide blocked photic phase advances in a dose-dependent manner and inhibited light-induced cFos-ir, without affecting light-induced circadian phase delays. These results suggest that NO may act as an intercellular messenger in the SCN, mediating light-induced phase advances.     

Characterization of circadian function in Djungarian hamsters insensitive to short day photoperiod

Summary Djungarian hamsters (Phodopus sungorus sungorus) depend mainly on day length to cue seasonal adjustments. However, not all individuals respond to short day conditions. A previous study from this laboratory proposed that nonresponsiveness to short day conditions rests with a defect in the circadian organization of these hamsters.In this study we found pronounced differences between responsive and nonresponsive hamsters in the expression of circadian rhythmicity under constant darkness and under constant illumination. While responsive hamsters showed a free-running activity pattern with a period of 23.86+0.04 h and responded to brief light pulses with the expected phase delays and phase advances, nonresponsive hamsters exhibited a period of 24.04+0.05 h and responded to light pulses with phase advances. Furthermore, 9 out of 15 responsive hamsters showed a clear split in the activity pattern within 8 weeks under constant light (80–100 lux), while only 1 of the 7 nonresponsive hamsters exhibited a split activity pattern. As a result of these differences in circadian function, nonresponsive Djungarian hamsters are incapable of proper photoperiod time measurement and photoperiod-induced seasonality.Abbreviations PRC phase response curve - ct circadian time - DD constant dark - LL constant light     

Twice Daily Melatonin Peaks in Siberian but not Syrian Hamsters under 24 h Light:Dark:Light:Dark Cycles

The daily pattern of blood-borne melatonin varies seasonally under the control of a multi-oscillator circadian pacemaker. Here we examine patterns of melatonin secretion and locomotor activity in Siberian and Syrian hamsters entrained to bimodal LDLD8:4:8:4 and LD20:4 lighting schedules that facilitate novel temporal arrangements of component circadian oscillators. Under LDLD, both species robustly bifurcated wheel-running activity in distinct day scotophase (DS) and night scotophase (NS) bouts. Siberian hamsters displayed significant melatonin increases during each scotophase in LDLD, and in the single daily scotophase of LD20:4. The bimodal melatonin secretion pattern persisted in acutely extended 16 h scotophases. Syrian hamsters, in contrast, showed no significant increases in plasma melatonin during either scotophase of LDLD8:4:8:4 or in LD20:4. In this species, detectable levels were observed only when the DS of LDLD was acutely extended to yield 16 h of darkness. Established species differences in the phase lag of nocturnal melatonin secretion relative to activity onset may underlie the above contrast: In non-bifurcated entrainment to 24 h LD cycles, Siberian hamsters show increased melatonin secretion within ～2 h after activity onset, whereas in Syrian hamsters, detectable melatonin secretion phase lags activity onset and the L/D transition by at least 4?h. The present results provide new evidence indicating multi-oscillator regulation of the waveform of melatonin secretion, specifically, the circadian control of the onset, offset and duration of nocturnal secretion.     

A 15-minute light pulse during darkness prevents the antigonadotrophic action of afternoon melatonin injections in male hamsters

When adult male Syrian hamsters were maintained under 14 h light and 10 h darkness daily (lights on from 0600-2000 h), peak pineal melatonin levels (705 pg/gland) were attained at 0500 h. When the dark phase of the light:dark cycle was interrupted with a 15 min pulse of light from 2300–2315 h (3 h after lights out), the highest melatonin levels achieved was roughly 400 pg/gland. Finally, if the 15 min pulse of light was given at 0200–0215 h (6 h after lights out) the nocturnal rise in pineal melatonin was completely abolished. Having made these observations, a second experiment was designed to determine the ability of afternoon melatonin injections to inhibit reproduction in hamsters kept under an uninterrupted 1410 cycle or under the same lighting regimen where the dark phase was interrupted with a 15 min pulse of light (0200–0215 h). In the uninterrupted light:dark schedule the daily afternoon injection of 25 g melatonin caused the testes and the accessory sex organs to atrophy within 11 weeks. Conversely, if the dark phase was interrupted with light between 0200–0215 h, afternoon melatonin injections were incapable of inhibiting the growth of the reproductive organs. The findings suggest that exogenously administered melatonin normally synergizes with endogenously produced melatonin to cause gonadal involution in hamsters.     

Altered Entrainment to the Day/Night Cycle Attenuates the Daily Rise in Circulating Corticosterone in the Mouse

The suprachiasmatic nucleus (SCN) is a circadian oscillator entrained to the day/night cycle via input from the retina. Serotonin (5-HT) afferents to the SCN modulate retinal signals via activation of 5-HT_1B receptors, decreasing responsiveness to light. Consequently, 5-HT_1B receptor knockout (KO) mice entrain to the day/night cycle with delayed activity onsets. Since circulating corticosterone levels exhibit a robust daily rhythm peaking around activity onset, we asked whether delayed entrainment of activity onsets affects rhythmic corticosterone secretion. Wheel-running activity and plasma corticosterone were monitored in mice housed under several different lighting regimens. Both duration of the light∶dark cycle (T cycle) and the duration of light within that cycle was altered. 5-HT_1B KO mice that entrained to a 9.5L:13.5D (short day in a T = 23 h) cycle with activity onsets delayed more than 4 h after light offset exhibited a corticosterone rhythm in phase with activity rhythms but reduced 50% in amplitude compared to animals that initiated daily activity <4 h after light offset. Wild type mice in 8L:14D (short day in a T = 22 h) conditions with highly delayed activity onsets also exhibited a 50% reduction in peak plasma corticosterone levels. Exogenous adrenocorticotropin (ACTH) stimulation in animals exhibiting highly delayed entrainment suggested that the endogenous rhythm of adrenal responsiveness to ACTH remained aligned with SCN-driven behavioral activity. Circadian clock gene expression in the adrenal cortex of these same animals suggested that the adrenal circadian clock was also aligned with SCN-driven behavior. Under T cycles <24 h, altered circadian entrainment to short day (winter-like) conditions, manifest as long delays in activity onset after light offset, severely reduces the amplitude of the diurnal rhythm of plasma corticosterone. Such a pronounced reduction in the glucocorticoid rhythm may alter rhythmic gene expression in the central nervous system and in peripheral organs contributing to an array of potential pathophysiologies.