Dynamic regulation of polysialylated neural cell adhesion molecule in the suprachiasmatic nucleus

The suprachiasmatic nucleus (SCN) prominently expresses polysialic acid (PSA), a carbohydrate polymer that is attached to neural cell adhesion molecule (NCAM) and promotes changes in cell interactions. Previous studies have shown that expression of PSA is important for circadian rhythm stability under constant darkness, and for photic entrainment of the SCN circadian clock. In the present study, immunoblot analyses of the Syrian hamster SCN revealed marked diurnal fluctuations in PSA under a 24-h light/dark cycle. PSA levels were reduced by >90% during the mid-to-late dark phase, and were elevated to maximal daytime levels approximately 1 h after lights-on. A similar pattern of PSA fluctuation persisted under constant darkness. Exposure of animals under a 24-h light/dark cycle to a 30-min light pulse during the late dark phase dramatically increased SCN contents of PSA within 60 min, and these values returned to basal levels 1-2 h later. There was no effect of light-on expression of PSA in the hippocampus. Parallel studies revealed changes in the NCAM-180 isoform that carries PSA in the brain, suggesting that regulation of PSA may include protein as well as carbohydrate-associated mechanisms. Immunohistological analysis revealed light-induced enhancement of PSA in the SCN subregion containing calbindin D(28K) cells. PSA staining was also closely associated with the majority of SCN cells expressing light-inducible Fos protein. This rhythmic, light-inducible expression of PSA within the SCN suggests that dynamic cell interactions are important for the photic regulation of circadian clock phase.     

Circadian rhythm in inhibitory synaptic transmission in the mouse suprachiasmatic nucleus

It is widely accepted that most suprachiasmatic nucleus (SCN) neurons express the neurotransmitter GABA and are likely to use this neurotransmitter to regulate excitability within the SCN. To evaluate the possibility that inhibitory synaptic transmission varies with a circadian rhythm within the mouse SCN, we used whole cell patch-clamp recording in an acute brain slice preparation to record GABA-mediated spontaneous inhibitory postsynaptic currents (sIPSCs). We found that the sIPSC frequency in the dorsal SCN (dSCN) exhibited a TTX-sensitive daily rhythm that peaked during the late day and early night in mice held in a light:dark cycle. We next evaluated whether vasoactive intestinal peptide (VIP) was responsible for the observed rhythm in IPSC frequency. Pretreatment of SCN slices with VPAC(1)/VPAC(2)- or VPAC(2)-specific receptor antagonists prevented the increase in sIPSC frequency in the dSCN. The rhythm in sIPSC frequency was absent in VIP/peptide histidine isoleucine (PHI)-deficient mice. Finally, we were able to detect a rhythm in the frequency of inhibitory synaptic transmission in mice held in constant darkness that was also dependent on VIP and the VPAC(2) receptor. Overall, these data demonstrate that there is a circadian rhythm in GABAergic transmission in the dorsal region of the mouse SCN and that the VIP is required for expression of this rhythm.     

The chronobiology of the Natal mole-rat, Cryptomys hottentotus natalensis

The Natal mole-rat, Cryptomys hottentotus natalensis, rarely, if ever, is exposed to external light cues because it occurs in completely sealed tunnel systems. As a result, their classical visual system is regressed, and therefore, their circadian system is expected proportionally to be expanded. Locomotor activity was investigated under a number of different photic regimes. Nine of the 12 mole-rats exhibited endogenous circadian rhythms of locomotor activity under constant darkness, with a mean free run period of 24.13 h (range 23.93-24.13 h), with these animals entrained to a light-dark cycle (12 L:12 D). Because C. hottentotus natalensis are able to entrain their locomotor activity to an external light source, light must reach the suprachiasmatic nucleus (SCN), suggesting a functional circadian clock. A clear day-night rhythm of melatonin secretion in animals housed under a neutral photoperiod (12 L:12 D) was observed, with higher melatonin concentrations in the dark compared with the light phase. The rhythm was maintained after the animals were transferred to either continuous light (LL) or dark (DD), suggesting that the endogenous rhythm was maintained under acute exposure to light and dark. However, under DD, the rhythm appeared to shift slightly, potentially as a result of the rhythm free running. These results show that C. hottentotus natalensis has endogenous rhythms of both locomotor activity and melatonin secretion, which are modulated by light.     

Neuropeptide Y: Role in light-dark cycle entrainment of hamster circadian rhythms

Microinjection of neuropeptide Y (NPY) into the suprachiasmatic region of the hypothalamus (SCN) phase shifted the circadian activity rhythm of hamsters housed in constant light. NPY advanced the phase when injected during the 12 h that preceded the daily onset of activity and tended to phase delay the activity cycle when injected during the 12 h after activity onset. In contrast, injection of saline into the SCN or NPY into the ventricular system had no effect on circadian phase. These and other data suggest that NPY functions as a chemical messenger important for the light-dark cycle entrainment of circadian rhythms.     

Changes in and dorsal to the rat suprachiasmatic nucleus during early pregnancy

Circadian rhythms in behavior and physiology change as female mammals transition from one reproductive state to another. The mechanisms responsible for this plasticity are poorly understood. The suprachiasmatic nucleus (SCN) of the hypothalamus contains the primary circadian pacemaker in mammals, and a large portion of its efferent projections terminate in the ventral subparaventricular zone (vSPZ), which also plays important roles in rhythm regulation. To determine whether these regions might mediate changes in overt rhythms during early pregnancy, we first compared rhythms in Fos and Per2 protein expression in the SCN and vSPZ of diestrous and early pregnant rats maintained in a 12:12-h light/dark (LD) cycle. No differences in the Fos rhythm were seen in the SCN core, but in the SCN shell, elevated Fos expression was maintained throughout the light phase in pregnant, but not diestrous, rats. In the vSPZ, the Fos rhythm was bimodal in diestrous rats, but this rhythm was lost in pregnant rats. Peak Per2 expression was phase-advanced by 4 h in the SCN of pregnant rats, and some differences in Per2 expression were found in the vSPZ as well. To determine whether differences in Fos expression were due to altered responsivity to light, we next characterized light-induced Fos expression in the SCN and vSPZ of pregnant and diestrous rats in the mid-subjective day and night. We found that the SCN core of the two groups responded in the same way at each time of day, whereas the rhythm of Fos responsivity in the SCN shell and vSPZ differed between diestrous and pregnant rats. These results indicate that the SCN and vSPZ are functionally re-organized during early pregnancy, particularly in how they respond to the photic environment. These changes may contribute to changes in overt behavioral and physiological rhythms that occur at this time.     

A novel quantitative trait locus on mouse chromosome 18, "era1," modifies the entrainment of circadian rhythms

STUDY OBJECTIVE: The mammalian circadian clock in the suprachiasmatic nuclei (SCN) of the hypothalamus conveys 24-h rhythmicity to sleep-wake cycles, locomotor activity, and other behavioral and physiological processes. The timing of rhythms relative to the light/dark (LD12:12) cycle is influenced in part by the endogenous circadian period and the time of day specific sensitivity of the clock to light. We now describe a novel circadian rhythm phenotype, and a locus influencing that phenotype, in a segregating population of mice. METHODS: By crossbreeding 2 genetically distinct nocturnal strains of mice (Cast/Ei and C57BL/6J) and backcrossing the resulting progeny to Cast/Ei, we have produced a novel circadian phenotype, called early runner mice. RESULTS: Early runner mice entrain to a light/dark cycle at an advanced phase, up to 9 hours before dark onset. This phenotype is not significantly correlated with circadian period in constant darkness and is not associated with disruption of molecular circadian rhythms in the SCN, as assessed by analysis of period gene expression. We have identified a genomic region that regulates this phenotype-a major quantitative trait locus on chromosome 18 (near D18Mit184) that we have named era1 for Early Runner Activity locus one. Phase delays caused by light exposure early in the subjective night were of smaller magnitude in backcross offspring that were homozygous Cast/Ei at D18Mit184 than in those that were heterozygous at this locus. CONCLUSION: Genetic variability in the circadian response to light may, in part, explain the variance in phase angle of entrainment in this segregating mouse population.     

Circadian change in tryptophan hydroxylase protein levels within the rat intergeniculate leaflets and raphe nuclei

Serotonin (5-HT) is involved in the synchronisation of the mammalian circadian clock located in the suprachiasmatic nuclei of the hypothalamus (SCN). This clock is synchronised by light (photic cues) and by non-photic cues. Non-photic cues are notably conveyed to the SCN by a direct 5-HT pathway arising from the mesencephalic median raphe nucleus (MRN). Furthermore, an indirect projection conveys non-photic inputs by 5-HT fibres from the mesencephalic dorsal raphe nucleus (DRN) to the intergeniculate leaflets of the thalamus (IGL) which project to the SCN. In the rat, the quantitative distribution of tryptophan hydroxylase (TpH), used as an index of 5-HT synthesis, was studied by in situ immunoautoradiography in both the serotoninergic cell bodies area of the raphe nuclei and the serotoninergic terminal field of the IGL. Under a 12 h light: 12 h dark (LD 12:12), TpH protein amount exhibited a rhythmic variation within the IGL. The maximum levels were reached at the day/night transition. In both MRN and the lateral groups of the DRN, TpH variations were opposite to those observed in the IGL. Such phase opposition was reported previously in the MRN/SCN pathway and was correlated with a rhythmic release of 5-HT within the SCN [Eur J Neurosci 15 (2002) 833]. Thus, the daily rhythmicity of TpH levels observed in DRN-IGL pathway may be correlated with a rhythmic release of 5-HT in the IGL at the beginning of the night. Under constant darkness, TpH rhythmic variations in the two serotoninergic pathways were maintained and similar to those observed under light/dark cycle. These results demonstrate the existence of a circadian endogenous functioning in the 5-HT neurones projecting to the rat circadian system.     

Phase dependent response of vasoactive intestinal polypeptide to light and darkness in the suprachiasmatic nucleus

Responsiveness of the vasoactive intestinal polypeptide (VIP) content to light and darkness in the rat suprachiasmatic nucleus (SCN) was examined by enzyme immunoassay of micropunched tissues. VIP content in the SCN has been shown to decrease monotonically in animals maintained in illumination. Decreases in VIP content in the SCN in response to both 6-h light and dark pulses depended on the phase of the circadian cycle when the pulses were applied. Light imposed at circadian time (CT) 18 or CT 22 was more effective in suppressing VIP levels than light exposure of the same intensity imposed at CT 0 or CT 6. Darkness interrupting continuous light was more effective at around CT 0 and less effective at around CT 12. These results suggest that VIP responsiveness to light and darkness in the SCN is regulated by the circadian clock in different ways and are correlated with phase-dependent phase shifts in the activity rhythm after light and dark pulses.     

Dark pulse resetting of the suprachiasmatic clock in Syrian hamsters: behavioral phase-shifts and clock gene expression

In mammals, the circadian clock in the suprachiasmatic nuclei (SCN) is mainly synchronized to photic cues provided by the daily light/dark cycle. Phase-shifts produced by light exposure during the night are correlated with rapid induction of two clock genes, Per1 and Per2, in the SCN. Nonphotic stimuli such as behavioral and pharmacological cues, when presented during the subjective day, induce behavioral phase-advances and a down-regulation of Per1 and Per2 expression in the SCN. When applied during the subjective day, dark pulses in continuous light also produce phase-advances. These phase-shifting effects have been interpreted as reflecting either a photic image mirror, nonphotic cues, or a combination of both. Here we evaluated in Syrian hamsters housed in constant light how dark pulses applied in late subjective day affect levels of Per1, Per2 and Cry1 mRNA. Four-hour dark pulses with no access to a wheel produced 1.2+/-0.4 h phase-advances of locomotor activity rhythm while control manipulation induced non-significant shifts (0.1+/-0.2 h). Dark pulses transiently down-regulated Per1 and Per2 mRNA levels in the SCN by 40 and 20% respectively, while the levels of Cry1 mRNA remained unaffected. In behaviorally split hamsters in which Per oscillations were asymmetric between the left and right sides of the SCN, dark pulses reduced Per expression in the half-SCN with high Per. This study shows that exposure during the late subjective day to dark pulses independent of wheel-running have nonphotic-like effects on the SCN clock at both behavioral and molecular levels.     

Daily rhythms in PER1 within and beyond the suprachiasmatic nucleus of female grass rats (Arvicanthis niloticus)

Although circadian rhythms of males and females are different in a variety of ways in many species, their mechanisms have been primarily studied in males. Furthermore, rhythms are dramatically different in diurnal and nocturnal animals but have been studied predominantly in nocturnal ones. In the present study, we examined rhythms in one element of the circadian oscillator, the PER1 protein, in a variety of cell populations in brains of diurnal female grass rats. Every 4 h five adult female grass rats kept on a 12-h light/dark (LD) cycle were perfused and their brains were processed for immunohistochemical detection of PER1. Numbers of PER1-labeled cells were rhythmic not only within the suprachiasmatic nucleus (SCN), the locus of the primary circadian clock in mammals, but also in the peri-suprachiasmatic region, the oval nucleus of the bed nucleus of the stria terminalis, the central amygdala, and the nucleus accumbens. In addition, rhythms were detected within populations of neuroendocrine cells that contain tyrosine hydroxylase. The phase of the rhythm within the SCN was advanced compared with that seen previously in male grass rats. Rhythms beyond the SCN were varied and different from those seen in most nocturnal species, suggesting that signals originating in the SCN are modified by its direct and/or indirect targets in different ways in nocturnal and diurnal species.     

Differences in the suprachiasmatic nucleus and lower subparaventricular zone of diurnal and nocturnal rodents

Diurnal and nocturnal species are profoundly different in terms of the temporal organization of daily rhythms in physiology and behavior. The neural bases for these divergent patterns are at present unknown. Here we examine functional differences in the suprachiasmatic nucleus (SCN) and one of its primary targets in a diurnal rodent, the unstriped Nile grass rat (Arvicanthis niloticus) and in a nocturnal one, the laboratory rat (Rattus norvegicus). Grass rats and laboratory rats were housed in a 12:12 light:dark cycle, and killed at six time points. cFos-immunoreactive rhythms in the SCN of grass rats and laboratory rats were similar to those reported previously, with peaks early in the light phase and troughs in the dark phase. However, cFos-immunoreactivity in the lower subparaventricular zone (LSPV) of grass rats rose sharply 5 h into the dark phase, and remained high through the first hour after light onset, whereas in laboratory rats it peaked 1 h after light onset and was low at all other sampling times. Daily cFos rhythms in both the SCN and the LSPV persisted in grass rats, but not in laboratory rats, after extended periods in constant darkness. In grass rats, the endogenous cFos rhythm in the LSPV, but not the SCN, was present both in calbindin-positive and in calbindin-negative cells. Cells that expressed cFos at night in the region of the LSPV in grass rats were clearly outside of the boundaries of the SCN as delineated by Nissl stain and immunoreactivity for vasopressin and vasoactive intestinal peptide. The LSPV of the grass rat, a region that receives substantial input from the SCN, displays a daily rhythm in cFos expression that differs from that of laboratory rats with respect to its rising phase, the duration of the peak and its dependence on a light/dark cycle. These characteristics may reflect the existence of mechanisms in the LSPV that enable it to modulate efferent SCN signals differently in diurnal and nocturnal species.     

The role of calcium ions in circadian rhythm of suprachiasmatic nucleus neuron activity in rat hypothalamic slices

The role of calcium ions in maintaining the circadian rhythm of suprachiasmatic nucleus (SCN) neuron activity was investigated using rat hypothalamic slice preparations. In normal Krebs solution, the firing rate of SCN neurons was higher in the light period than in the dark period. In Ca2+-free Krebs solution, SCN neuron activity was low during all periods and did not show diurnal rhythm. These results suggest that the disappearance of circadian rhythmic change of SCN neuron activity in Ca2+-free Krebs solution may be due to the disappearance of synaptic transmission in the SCN.     

Direction-dependent effects of chronic "jet-lag" on hippocampal neurogenesis

Disruptions in circadian rhythms, as seen in human shift workers, are often associated with many health consequences including impairments in cognitive functions. However, the mechanisms underlying these affects are not well understood. The objective of the present study is to explore the effects of circadian disruption on hippocampal neurogenesis, which has been implicated in learning and memory and could serve as a potential pathway mediating the cognitive consequences associated with rhythm disruption. Circadian rhythm disruptions were introduced using a weekly 6 h phase shifting paradigm, in which male Wistar rats were subjected to either 6 h phase advances (i.e. traveling eastbound from New York to Paris) or 6 h phase delays (i.e. traveling westbound from Paris to New York) in their light/dark schedule every week. The effects of chronic phase shifts on hippocampal neurogenesis were assessed using doublecortin (DCX), a microtubule binding protein expressed in immature neurons. The results revealed that chronic disruption in circadian rhythms inhibits hippocampal neurogenesis, and the degree of reduction in neurogenesis depends upon the direction and duration of the shifts. In two cohorts of animals that experienced phase shifts for either 4 or 8 weeks, a greater decrease in neurogenesis was observed when the phase was advanced versus delayed in both groups. The direction-dependent effect mirrors the findings on clock gene expression in the SCN, suggesting a causal link between the reduction in hippocampal neurogenesis and a disrupted SCN circadian clock.     

Circadian rhythm of histamine release from the hypothalamus of freely moving rats.

Using an in vivo microdialysis technique coupled with HPLC-fluorometry, the release of neuronal histamine from the anterior hypothalamic area was monitored continuously in conscious, freely moving rats under a 12:12 h light:dark cycle. Spontaneous locomotor activity of the rats was measured simultaneously using a locomotor activity counter. Histamine release gradually increased in the second half of the light period (1400-2000) and the average histamine release during the dark period (2000-0800, 0.20 +/- 0.02 pmol/30 min) was significantly higher than that during the light period (0.12 +/- 0.01 pmol/30 min). This clear circadian change in the release suggests that the central histaminergic system is related to the circadian rhythm of rats.     

Constant light housing attenuates circadian rhythms of mPer2 mRNA and mPER2 protein expression in the suprachiasmatic nucleus of mice

Constant light (LL) or constant dark (DD) environmental lighting conditions cause a free-running period and activity reduction in the rodent behavioral circadian rhythm. In order to understand the molecular process underlying behavioral rhythms in LL or DD housing conditions, we examined the circadian profile of mPer2 mRNA and mPER2 in the suprachiasmatic nucleus (SCN), a main oscillator, of free-running mice. The circadian expression rhythm of mPer2 in the SCN was dampened under 7-day LL conditions, whereas that of mPER2 protein was moderately attenuated and its expression peak delayed. The circadian expression of mPer2 and its product was slightly attenuated and advanced by 7-day DD conditions. With arrhythmic behavioral activity caused by long-term LL housing, mPER2 protein lost its rhythmicity in the SCN. On the other hand, LL or DD housing did not affect the mPer2 gene and its product in the cerebral cortex. The present results suggest that mPER2 circadian expression in the SCN corresponds well with behavioral circadian oscillation under LL or DD conditions. Thus, the behavioral circadian rhythm seems to correlate with molecular clock works in the SCN.     

Attenuation of circadian light induced phase advances and delays by neuropeptide Y and a neuropeptide Y Y1/Y5 receptor agonist

Circadian rhythms can be synchronised to photic and non-photic stimuli. The circadian clock, anatomically defined as the suprachiasmatic nucleus in mammals, can be phase shifted by light during the night. Non-photic stimuli reset the circadian rhythm during the day. Photic and non-photic stimuli have been shown to interact during the day and night. Precise mechanisms for these complex interactions are unknown. A possible pathway for non-photic resetting of the clock is thought to generate from the intergeniculate leaflet, which conveys information to the suprachiasmatic nucleus (SCN) through the geniculohypothalamic tract and utilises neuropeptide Y (NPY) as its primary neurotransmitter.Interactions between light and NPY were investigated during the early (2 h after activity onset) and late (6 h after activity onset) night in male Syrian hamsters. NPY microinjections into the region of the SCN significantly attenuated light-induced phase delay, during the early subjective night. Phase advances to light were completely inhibited by the administration of NPY during the late night.The precise mechanism by which NPY attenuates or blocks photic phase shifts is unclear, but the NPY Y5 receptor has been implicated in the mediation of this inhibitory effect. The NPY Y1/Y5 receptor agonist, [Leu(31),Pro(34)]NPY, was administered via cannula microinjections following light exposure during the early and late night. [Leu(31),Pro(34)]NPY significantly attenuated phase delays to light during the early night and blocked phase advances during the late night, in a manner similar to NPY.These results show the ability of NPY to attenuate phase shifts to light during the early night and block light-induced phase advances during the late night. Furthermore, this is the first in vivo study implicating the involvement of the NPY Y1/Y5 receptors in the complex interaction of photic and non-photic stimuli during the night. The alteration of photic phase shifts by NPY may influence photic entrainment within the circadian system.     

Restricted feeding entrains liver clock without participation of the suprachiasmatic nucleus

BACKGROUND: There are two main stimuli that entrain the circadian rhythm, the light-dark cycle (LD) and restricted feeding (RF). Light-induced entrainment requires induction of the Per1 and Per2 genes in the suprachiasmatic nucleus (SCN), the locus of a main oscillator. In this experiment, we determined whether RF resets the expression of circadian clock genes in the mouse liver with or without participation of the SCN. RESULTS: Mice were allowed access to food for 4 h during the daytime (7 h advance of feeding time) under LD or constant darkness (DD). The peaks of mPer1, mPer2, D-site-binding protein (Dbp) and cholesterol 7alpha-hydroxylase (Cyp7A) mRNA in the liver were advanced 6-12 h after 6 days of RF, whereas those in SCN were unaffected. The advance of mPer expression in the liver by RF was still observed in SCN-lesioned mice. A 7 h advance in the LD cycle advanced the peaks of clock gene expression in both the liver and SCN, whereas, a shift in the LD did not move the phase of the liver clock when the shift was carried out under a fixed RF schedule during the night-time. CONCLUSIONS: These results suggest that restricted feeding strongly entrained the expression of circadian clock genes in the liver without the participation of an SCN clock function.     

Nighttime dim light exposure alters the responses of the circadian system

The daily light dark cycle is the most salient entraining factor for the circadian system. However, in modern society, darkness at night is vanishing as light pollution steadily increases. The impact of brighter nights on wild life ecology and human physiology is just now being recognized. In the present study, we tested the possible detrimental effects of dim light exposure on the regulation of circadian rhythms, using CD1 mice housed in light/dim light (LdimL, 300 lux:20 lux) or light/dark (LD, 300 lux:1 lux) conditions. We first examined the expression of clock genes in the suprachiasmatic nucleus (SCN), the locus of the principal brain clock, in the animals of the LD and LdimL groups. Under the entrained condition, there was no difference in PER1 peak expression between the two groups, but at the trough of the PER 1 rhythm, there was an increase in PER1 in the LdimL group, indicating a decrease in the amplitude of the PER1 rhythm. After a brief light exposure (30 min, 300 lux) at night, the light-induced expression of mPer1 and mPer2 genes was attenuated in the SCN of LdimL group. Next, we examined the behavioral rhythms by monitoring wheel-running activity to determine whether the altered responses in the SCN of LdimL group have behavioral consequence. Compared to the LD controls, the LdimL group showed increased daytime activity. After being released into constant darkness, the LdimL group displayed shorter free-running periods. Furthermore, following the light exposure, the phase shifting responses were smaller in the LdimL group. The results indicate that nighttime dim light exposure can cause functional changes of the circadian system, and suggest that altered circadian function could be one of the mechanisms underlying the adverse effects of light pollution on wild life ecology and human physiology.     

Effects of methamphetamine upon circadian rhythms in multiple unit activity inside and outside the suprachiasmatic nucleus in the golden hamster (Mesocricetus auratus)

To investigate the circadian system of the golden hamster, multiple unit activity (MUA) was recorded inside and outside the suprachiasmatic nucleus (SCN). MUA inside the SCN showed a daily rhythm with a daytime peak during a 24 h light-dark cycle (LD, 12:12), whereas MUA outside the SCN revealed a nighttime peak. The phase reversal of MUA between inside and outside the SCN in the golden hamster was similar to the rat which is also a nocturnal rodent. MUA rhythms to the lighting cycle started to freerun after methamphetamine administration. This result indicates that methamphetamine may affect directly the neural circadian oscillator to induce behavioral change.     

Thermoregulation in the cold changes depending on the time of day and feeding condition: physiological and anatomical analyses of involved circadian mechanisms

The circadian rhythm of body temperature (T_b) is a well-known phenomenon. However, it is unknown how the circadian system including the suprachiasmatic nucleus (SCN) and clock genes affects thermoregulation. Food deprivation in mice induces a greater reduction of T_b particularly in the light phase. We examined the role of Clock, one of key clock genes and the SCN during induced hypothermia. At 20 °C with fasting, mice increased their metabolic heat production in the dark phase and maintained T_b, whereas in the light phase, heat production was less, resulting in hypothermia. Under these conditions, neuronal activity in the SCN, assessed by cFos expression, increased only in the light phase. However, such differences in thermoregulatory and neural responses between the phases in Clock mutant mice were less marked. The neural network between the SCN and paraventricular nucleus appeared to be important in hypothermia. These findings suggest that the circadian system per se is influenced by both the feeding condition and environmental temperature and that it modulates thermoregulation.