Suprachiasmatic Nucleus Lesions Research Articles

Dysregulation of the Suprachiasmatic Nucleus Disturbs the Circadian Rhythm and Aggravates Epileptic Seizures by Inducing Hippocampal GABAergic Dysfunction in C57BL/6 Mice.

The interplay between circadian rhythms and epilepsy has gained increasing attention. The suprachiasmatic nucleus (SCN), which acts as the master circadian pacemaker, regulates physiological and behavioral rhythms through its complex neural networks. However, the exact role of the SCN and its Bmal1 gene in the development of epilepsy remains unclear. In this study, we utilized a lithium-pilocarpine model to induce epilepsy in mice and simulated circadian disturbances by creating lesions in the SCN and specifically knocking out the Bmal1 gene in the SCN neurons. We observed that the pilocarpine-induced epileptic mice experienced increased daytime seizure frequency, irregular oscillations in core body temperature, and circadian gene alterations in both the SCN and the hippocampus. Additionally, there was enhanced activation of GABAergic projections from the SCN to the hippocampus. Notably, SCN lesions intensified seizure activity, concomitant with hippocampal neuronal damage and GABAergic signaling impairment. Further analyses using the Gene Expression Omnibus database and gene set enrichment analysis indicated reduced Bmal1 expression in patients with medial temporal lobe epilepsy, potentially affecting GABA receptor pathways. Targeted deletion of Bmal1 in SCN neurons exacerbated seizures and pathology in epilepsy, as well as diminished hippocampal GABAergic efficacy. These results underscore the crucial role of the SCN in modulating circadian rhythms and GABAergic function in the hippocampus, aggravating the severity of seizures. This study provides significant insights into how circadian rhythm disturbances can influence neuronal dysfunction and epilepsy, highlighting the therapeutic potential of targeting SCN and the Bmal1 gene within it in epilepsy management.

Circadian Rhythm and Leptin Hormone Responses to Nutritional Restriction in Gerbils (Meriones unguiculatus) with Suprachiasmatic Nucleus Lesions

Rhythmically-regulated feeding behavior is in harmony with physiologic and metabolic activities in mammals. This rhythmic regulation is orchestrated by Suprachiasmatic Nucleus (SCN). However, it is not entirely clear how the SCN, which generates endogenous (internal) rhythms, influences body weight and serum leptin profile with activity rhythms in relation to feeding timing. In this study, animals in long photoperiod (14L:10D) were divided into two groups as control (sham-SCNx) and SCN lesions (SCNx). Then, these groups were split into four separate subgroups: a) ad libitum feeding; b) feeding only during the dark phase; c) feeding only during the light phase; and d) feeding during a specific period of the day (11:00-14:00 h). Locomotor activity and leptin hormone changes were observed in animals fed in cages attached to activity wheels for one month. Under the conditions of food restriction, the locomotor activities of the groups with SCNx and sham-SCNx demonstrated a phase shift toward the time of feeding. Serum leptin level did not change with feeding conditions but decreased in lesioned groups (SCNx). In conclusion, nutritional restriction caused phase shifts in activity rhythms and it was found that the SCN in gerbils was in charge of these rhythmic changes in the presence and absence of nutrients.

Circadian Clock Genes Are Regulated by Rhythmic Corticosterone at Physiological Levels in the Rat Hippocampus

Introduction: In the hippocampus, clock gene expression is important for memory and mood; however, the signaling mechanism controlling clock gene expression in the hippocampus is unknown. Recent findings suggest that circadian glucocorticoid rhythms driven by the suprachiasmatic nucleus (SCN) control rhythmic clock gene expression in neurons; in addition, dexamethasone modulates hippocampal clock gene expression. We therefore hypothesized that oscillations of clock genes in the hippocampus could be driven by SCN-controlled circadian rhythms in glucocorticoids. Methods: Temporal profiles of hippocampal clock gene expression were established by quantitative reverse-transcription real-time PCR on rat hippocampi, while cellular distribution was established by in situ hybridization. To determine the effect of rhythmic glucocorticoids on hippocampal clock gene expression, the SCN was lesioned, adrenal glands removed and a 24 h exogenous corticosterone rhythm at physiological levels was reestablished by use of a programmable infusion pump. Results: Daily rhythms were detected for Per1, Per2, Bmal1, Nr1d1, and Dbp, while clock gene products were confirmed in both the hippocampus proper and the dentate gyrus. In sham controls, differential hippocampal expression of Per1 and Dbp between ZT3 and ZT15 was detectable. This rhythm was abolished by SCN lesion; however, reestablishing the natural rhythm in corticosterone restored differential rhythmic expression of both Per1 and Dbp. Further, a 6 h phase delay in the corticosterone profile caused a predictable shift in expression of Nr1d1. Conclusion: Our data show that rhythmic corticosterone can drive hippocampal clock gene rhythms suggesting that the SCN regulates the circadian oscillator of the hippocampus by controlling the circadian rhythm in circulating glucocorticoids.

The Circadian Oscillator of the Cerebellum: Triiodothyronine Regulates Clock Gene Expression in Granule Cells in vitro and in the Cerebellum of Neonatal Rats in vivo.

The central circadian clock resides in the suprachiasmatic nucleus (SCN) of the hypothalamus, but an SCN-dependent molecular circadian oscillator is present in the cerebellar cortex. Recent findings suggest that circadian release of corticosterone is capable of driving the circadian oscillator of the rat cerebellum. To determine if additional neuroendocrine signals act to shape cerebellar clock gene expression, we here tested the role of the thyroid hormone triiodothyronine (T3) in regulation of the cerebellar circadian oscillator. In cultured cerebellar granule cells from mixed-gender neonatal rats, T3 treatment affected transcript levels of the clock genes Per2, Arntl, Nr1d1, and Dbp, suggesting that T3 acts directly on granule cells to control the circadian oscillator. We then used two different in vivo protocols to test the role of T3 in adult female rats: Firstly, a single injection of T3 did not influence clock gene expression in the cerebellum. Secondly, we established a surgical rat model combining SCN lesion with a programmable micropump infusing circadian physiological levels of T3; however, rhythmic infusion of T3 did not reestablish differential clock gene expression between day and night in SCN lesioned rats. To test if the effects of T3 observed in vitro were related to the developmental stage, acute injections of T3 were performed in mixed-gender neonatal rats in vivo; this procedure significantly affected cerebellar expression of the clock genes Per1, Per2, Nr1d1, and Dbp. Developmental comparisons showed rhythmic expression of all clock genes analyzed in the cerebellum of adult rats only, whereas T3 responsiveness was limited to neonatal animals. Thus, T3 shapes cerebellar clock gene profiles in early postnatal stages, but it does not represent a systemic circadian regulatory mechanism linking the SCN to the cerebellum throughout life.

Loss of Circadian Timing Disrupts Theta Episodes during Object Exploration.

This study examined whether theta oscillations were compromised by the type of circadian disruption that impairs hippocampal-dependent memory processes. In prior studies on Siberian hamsters, we developed a one-time light treatment that eliminated circadian timing in the central pacemaker, the suprachiasmatic nucleus (SCN). These arrhythmic animals had impaired hippocampal-dependent memory whereas animals made arrhythmic with SCN lesions did not. The current study examined whether theta oscillations are compromised by the same light treatment that produced memory impairments in these animals. We found that both methods of inducing circadian-arrhythmia shortened theta episodes in the EEG by nearly 50%. SCN-lesioned animals, however, exhibited a 3-fold increase in the number of theta episodes and more than doubled the total time that theta dominated the EEG compared to SCN-intact circadian-arrhythmic animals. Video tracking showed that changes in theta were paralleled by similar changes in exploration behavior. These results suggest that the circadian-arrhythmic SCN interferes with hippocampal memory encoding by fragmenting theta oscillations. SCN-lesioned animals can, however, compensate for the shortened theta episodes by increasing their frequency. Implications for rhythm coherence and theta sequence models of memory formation are discussed.

Monochromatic Blue Light Activates Suprachiasmatic Nucleus Neuronal Activity and Promotes Arousal in Mice Under Sevoflurane Anesthesia.

Background: Monochromatic blue light (MBL), with a wavelength between 400–490 nm, can regulate non-image-forming (NIF) functions of light in the central nervous system. The suprachiasmatic nucleus (SCN) in the brain is involved in the arousal-promoting response to blue light in mice. Animal and human studies showed that the responsiveness of the brain to visual stimuli is partly preserved under general anesthesia. Therefore, this study aimed to investigate whether MBL promotes arousal from sevoflurane anesthesia via activation of the SCN in mice.Methods: The induction and emergence time of sevoflurane anesthesia under MBL (460 nm and 800 lux) exposure was measured. Cortical electroencephalograms (EEGs) were recorded and the burst-suppression ratio (BSR) was calculated under MBL during sevoflurane anesthesia. The EEGs and local field potential (LFP) recordings with or without locally electrolytic ablated bilateral SCN were used to further explore the role of SCN in the arousal-promoting effect of MBL under sevoflurane anesthesia. Immunofluorescent staining of c-Fos was conducted to reveal the possible downstream mechanism of SCN activation.Results: Unlike the lack of effect on the induction time, MBL shortened the emergence time and the EEG recordings showed cortical arousal during the recovery period. MBL resulted in a significant decrease in BSR and a marked increase in EEG power at all frequency bands except for the spindle band during 2.5% sevoflurane anesthesia. MBL exposure under sevoflurane anesthesia enhances the neuronal activity of the SCN. These responses to MBL were abolished in SCN lesioned (SCNx) mice. MBL evoked a high level of c-Fos expression in the prefrontal cortex (PFC) and lateral hypothalamus (LH) compared to polychromatic white light (PWL) under sevoflurane anesthesia, while it exerted no effect on c-Fos expression in the ventrolateral preoptic area (VLPO) and locus coeruleus (LC) c-Fos expression.Conclusions: MBL promotes behavioral and electroencephalographic arousal from sevoflurane anesthesia via the activation of the SCN and its associated downstream wake-related nuclei. The clinical implications of this study warrant further study.

Free-running circadian breathing rhythms are eliminated by suprachiasmatic nucleus lesion.

It is widely agreed that breathing is subject to circadian regulation. Circadian differences in respiratory physiology significantly impact a number of diseases including sleep apnea, asthma, and seizure-induced death. The effect of time of day on breathing has been previously characterized; however, an endogenous free-running respiratory rhythm in mammals has not previously been described. Furthermore, it is assumed that circadian rhythms in breathing are dependent on the hypothalamic suprachiasmatic nucleus (SCN), the home of the mammalian central circadian oscillator, but this has not been shown experimentally. The breathing of mice was monitored during wakefulness using whole body plethysmography at six times of day while housed under light-dark conditions and at six circadian phases while housed under constant darkness. Respiratory frequency and minute ventilation, but not tidal volume, were significantly higher during the active phase in both entrained and free-running conditions. To determine whether circadian regulation of breathing requires the SCN, in separate sets of animals this structure was electrolytically lesioned bilaterally or a sham surgery was performed, and breathing was measured at six different time points. Time-dependent oscillations in breathing were lost in SCN-lesioned animals, but not those subjected to sham surgery. These results suggest that breathing is subject to circadian regulation via the SCN. Mechanistic insights into the circadian regulation of breathing may lead to targeted interventions to reduce the morbidity and mortality associated with diseases with respiratory pathophysiology.NEW & NOTEWORTHY It has long been appreciated that breathing is altered by time of day. This study demonstrates that rhythmicity in breathing persists in constant darkness but is dependent on the suprachiasmatic nucleus in the hypothalamus. Understanding circadian rhythms in breathing may be important for the treatment and prevention of diseases such as sleep apnea and sudden unexpected death in epilepsy.

Biological clocks and incremental growth line formation in dentine.

Dentine- and enamel-forming cells secrete matrix in consistent rhythmic phases, resulting in the formation of successive microscopic growth lines inside tooth crowns and roots. Experimental studies of various mammals have proven that these lines are laid down in subdaily, daily (circadian), and multidaily rhythms, but it is less clear how these rhythms are initiated and maintained. In 2001, researchers reported that lesioning the so-called master biological clock, the suprachiasmatic nucleus (SCN), halted daily line formation in rat dentine, whereas subdaily lines persisted. More recently, a key clock gene (Bmal1) expressed in the SCN in a circadian manner was also found to be active in dentine- and enamel- secretory cells. To probe these potential neurological and local mechanisms for the production of rhythmic lines in teeth, we reexamined the role of the SCN in growth line formation in Wistar rats and investigated the presence of daily lines in Bmal1 knockout mice (Bmal1-/- ). In contrast to the results of the 2001 study, we found that both daily and subdaily growth lines persisted in rat dentine after complete or partial SCN lesion in the majority of individuals. In mice, after transfer into constant darkness, daily rhythms continued to manifest as incremental lines in the dentine of each Bmal1 genotype (wild-type, Bmal+/- , and Bmal1-/- ). These results affirm that the manifestation of biological rhythms in teeth is a robust phenomenon, imply a more autonomous role of local biological clocks in tooth growth than previously suggested, and underscore the need further to elucidate tissue-specific circadian biology and its role in incremental line formation. Investigations of this nature will strengthen an invaluable system for determining growth rates and calendar ages from mammalian hard tissues, as well as documenting the early lives of fossil hominins and other primates.

Impaired function of the suprachiasmatic nucleus rescues the loss of body temperature homeostasis caused by time-restricted feeding

The suprachiasmatic nucleus (SCN) is the master circadian pacemaker that drives body temperature rhythm. Time-restricted feeding (TRF) has potential as a preventative or therapeutic approach against many diseases. The potential side effects of TRF remain unknown. Here we show that daily 4-h TRF treatment in mice can severely impair body temperature homeostasis and can result in lethality. Nearly half of the mice died at 21 °C, and all mice died at 18 °C during 4-h TRF. This effect was modulated by the circadian clock and was associated with severe hypothermia due to loss of body temperature homeostasis. Remarkably, disrupting the circadian clock by SCN lesions or constant light rescued lethality during TRF. Analysis of circadian gene expression in the dorsomedial hypothalamus (DMH) demonstrated that TRF reprograms rhythmic transcriptome in DMH and suppresses expression of genes, such as Ccr5 and Calcrl, which are involved in thermoregulation. Together, our results demonstrated a potent effect of TRF on body temperature homeostasis and the role of SCN function in this process. Our results further suggested that circadian arrhythmicity can have potential beneficial effects under certain stress conditions.

The suprachiasmatic nucleus drives day-night variations in postprandial triglyceride uptake into skeletal muscle and brown adipose tissue.

What is the central question of this study? What are the factors influencing day-night variations in postprandial triglycerides? What is the main finding and its importance? Rats show low postprandial plasma triglyceride concentrations early in the active period that are attributable to a higher uptake by skeletal muscle and brown adipose tissue. We show that these day-night variations in uptake are driven by the suprachiasmatic nucleus, probably via a Rev-erbα-mediated mechanism and independent of locomotor activity. These findings highlight that the suprachiasmatic nucleus has a major role in day-night variations in plasma triglycerides and that disturbances in our biological clock might be an important risk factor contributing to development of postprandial hyperlipidaemia. Energy metabolism follows a diurnal pattern, mainly driven by the suprachiasmatic nucleus (SCN), and disruption of circadian regulation has been linked to metabolic abnormalities. Indeed, epidemiological evidence shows that night work is a risk factor for cardiovascular disease, and postprandial hyperlipidaemia is an important contributor. Therefore, the aim of this work was to investigate the factors that drive day-night variations in postprandial triglycerides (TGs). Intact and SCN-lesioned male Wistar rats were subjected to an oral fat challenge during the beginning of the rest phase (day) or the beginning of the active phase (night). The plasma TG profile was evaluated and tissue TG uptake assayed. After the fat challenge, intact rats showed lower postprandial plasma TG concentrations early in the night when compared with the day. However, no differences were observed in the rate of intestinal TG secretion between day and night. Instead, there was a higher uptake of TG by skeletal muscle and brown adipose tissue early in the active phase (night) when compared with the rest phase (day), and these variations were abolished in rats bearing bilateral SCN lesions. Rev-erbα gene expression suggests this as a possible mediator of the mechanism linking the SCN and day-night variations in TG uptake. These findings show that the SCN has a major role in day-night variations in plasma TGs by promoting TG uptake into skeletal muscle and brown adipose tissue. Consequently, disturbance of the biological clock might be an important risk factor contributing to the development of hyperlipidaemia.

Circadian clock-dependent increase in salivary IgA secretion modulated by sympathetic receptor activation in mice

The salivary gland is rhythmically controlled by sympathetic nerve activation from the suprachiasmatic nucleus (SCN), which functions as the main oscillator of circadian rhythms. In humans, salivary IgA concentrations reflect circadian rhythmicity, which peak during sleep. However, the mechanisms controlling this rhythmicity are not well understood. Therefore, we examined whether the timing of parasympathetic (pilocarpine) or sympathetic (norepinephrine; NE) activation affects IgA secretion in the saliva. The concentrations of saliva IgA modulated by pilocarpine activation or by a combination of pilocarpine and NE activation were the highest in the middle of the light period, independent of saliva flow rate. The circadian rhythm of IgA secretion was weakened by an SCN lesion and Clock gene mutation, suggesting the importance of the SCN and Clock gene on this rhythm. Adrenoceptor antagonists blocked both NE- and pilocarpine-induced basal secretion of IgA. Dimeric IgA binds to the polymeric immunoglobulin receptor (pIgR) on the basolateral surface of epithelial cells and forms the IgA-pIgR complex. The circadian rhythm of Pigr abundance peaked during the light period, suggesting pIgR expression upon rhythmic secretion of IgA. We speculate that activation of sympathetic nerves during sleep may protect from bacterial access to the epithelial surface through enhanced secretion of IgA.

Suprachiasmatic Nucleus Interaction with the Arcuate Nucleus; Essential for Organizing Physiological Rhythms

The suprachiasmatic nucleus (SCN) is generally considered the master clock, independently driving all circadian rhythms. We recently demonstrated the SCN receives metabolic and cardiovascular feedback adeptly altering its neuronal activity. In the present study, we show that microcuts effectively removing SCN-arcuate nucleus (ARC) interconnectivity in Wistar rats result in a loss of rhythmicity in locomotor activity, corticosterone levels, and body temperature in constant dark (DD) conditions. Elimination of these reciprocal connections did not affect SCN clock gene rhythmicity but did cause the ARC to desynchronize. Moreover, unilateral SCN lesions with contralateral retrochiasmatic microcuts resulted in identical arrhythmicity, proving that for the expression of physiological rhythms this reciprocal SCN-ARC interaction is essential. The unaltered SCN c-Fos expression following glucose administration in disconnected animals as compared to a significant decrease in controls demonstrates the importance of the ARC as metabolic modulator of SCN neuronal activity. Together, these results indicate that the SCN is more than an autonomous clock, and forms an essential component of a larger network controlling homeostasis. The present novel findings illustrate how an imbalance between SCN and ARC communication through circadian disruption could be involved in the etiology of metabolic disorders.

Stereotaxic Surgery for Suprachiasmatic Nucleus Lesions in Mice.

Site-specific lesions are invaluable methods for investigating the function of brain regions within the central nervous system and can be used to study neural mechanisms of behaviors. Precise stereotaxic surgery is required to lesion small regions of the brain such as the suprachiasmatic nucleus (SCN), which harbors the master circadian clock. In this protocol, we describe stereotaxic surgery optimized for bilateral lesion of the mouse SCN by loading electric current. Success of the SCN lesion is verified histologically and behaviorally by monitoring arrhythmic locomotor activity. The SCN-lesioned mouse allows for the evaluation of behavioral, biochemical, and physiological consequences of ablation of the master circadian clock.

The SCN Clock Governs Circadian Transcription Rhythms in Murine Epididymal White Adipose Tissue.

The circadian master pacemaker in the suprachiasmatic nucleus (SCN) orchestrates peripheral clocks in various organs and synchronizes them with external time, including those in adipose tissue, which displays circadian oscillations in various metabolic and endocrine outputs. Because our knowledge about the instructive role of the SCN clock on peripheral tissue function is based mainly on SCN lesion studies, we here used an alternative strategy employing the Cre/ loxP system to functionally delete the SCN clock in mice. We performed whole-genome microarray hybridizations of murine epididymal white adipose tissue (eWAT) RNA preparations to characterize the role of the SCN clock in eWAT circadian transcriptome regulation. Most of the rhythmic transcripts in control animals were not rhythmic in SCN mutants, but a significant number of transcripts were rhythmic only in mutant eWAT. Core clock genes were rhythmic in both groups, but as was reported before for other tissues, rhythms were dampened and phase advanced in mutant animals. In SCN-mutant mice, eWAT lost the rhythm of metabolic pathway-related transcripts, while transcripts gaining rhythms in SCN-mutant mice were associated with various immune functions. These data reveal a complex interaction of SCN-derived and local circadian signals in the regulation of adipose transcriptome programs.

Circadian expression of connexins in the mouse heart

Significant circadian variations exist in the frequency of cardiac arrhythmia; however, the underlying mechanism is largely unknown. Connexins are essential in the propagation of electrical activity throughout the heart and are an important determinant of conduction velocity. Their dysfunction is related to the genesis of cardiac arrhythmia. In this study, we investigated if cx40 and cx43 expressed circadianly in the mouse heart using suprachiasmatic nuclei (SCN) lesion and pharmacological autonomic nervous system block mouse. Significant 24-h variations were observed in the expression of cx40 and cx43 in the sham-operated mice. In the SCNX mouse, all genes examined lost circadian rhythm. In the ANSB mouse, the expressions of Bmal1 was dampened significantly but still had circadian rhythm, whereas the two connexin gene expressions lost rhythm. These results suggest that cx40 and cx43 gene expressions have clear circadian rhythm and might be regulated by the central clock in the SCN through the ANS.

Suprachiasmatic Nucleus and Subparaventricular Zone Lesions Disrupt Circadian Rhythmicity but Not Light-Induced Masking Behavior in Nile Grass Rats.

The ventral subparaventricular zone (vSPVZ) receives direct retinal input and influences the daily patterning of activity in rodents, making it a likely candidate for the mediation of acute behavioral responses to light (i.e., masking). We performed chemical lesions aimed at the vSPVZ of diurnal grass rats (Arvicanthis niloticus) using N-methyl-D,L-aspartic acid (NMA), a glutamate agonist. Following NMA lesions, we placed grass rats in various lighting conditions (e.g., 12:12 light-dark, constant dark, constant light); presented a series of light pulses at circadian times (CT) 6, 14, 18, and 22; and placed them in a 7-h ultradian cycle to assess behavioral masking. Extensive bilateral lesions of the vSPVZ disrupted the expression of circadian rhythms of activity and abolished the circadian modulation of masking responses to light, without affecting light-induced masking behavior per se. We also found that in diurnal grass rats, NMA was capable of destroying not only neurons of the vSPVZ but also those of the suprachiasmatic nucleus (SCN), even though excitotoxins have been ineffective at destroying cells within the SCN of nocturnal rodents. The vulnerability of the grass rat's SCN to NMA toxicity raises the possibility of a difference in density of receptors for glutamate between nocturnal and diurnal species. In cases in which damage extended to the SCN, masking responses to light were present and similar to those displayed by animals with damage restricted to the vSPVZ. Thus, extensive bilateral lesions of the SCN and vSPVZ disrupted the expression of circadian rhythms without affecting acute responses to light in a diurnal species. Our present and previous results suggest that retinorecipient brain areas other than the SCN or vSPVZ, such as the intergeniculate leaflet or olivary pretectal nucleus, may be responsible for the mediation of masking responses to light in the diurnal grass rat.

Circadian PER2::LUC rhythms in the olfactory bulb of freely moving mice depend on the suprachiasmatic nucleus but not on behaviour rhythms

The temporal order of physiology and behaviour in mammals is regulated by the coordination of the master circadian clock in the suprachiasmatic nucleus (SCN) and peripheral clocks in various tissues outside the SCN. Because the circadian oscillator(s) in the olfactory bulb (OB) is regarded as SCN independent, we examined the relationship between the SCN master clock and the circadian clock in the OB. We also examined the role of vasoactive intestinal peptide receptor 2 in the circadian organization of the OB. We continuously monitored the circadian rhythms of a clock gene product PER2 in the SCN and OB of freely moving mice by means of a bioluminescence reporter and an optical fibre implanted in the brain. Robust circadian rhythms were detected in the OB and SCN for up to 19 days. Bilateral SCN lesions abolished the circadian behaviour rhythms and disorganized the PER2 rhythms in the OB. The PER2 rhythms in the OB showed more than one oscillatory component of a similar circadian period, suggesting internal desynchronization of constituent oscillators. By contrast, significant circadian PER2 rhythms were detected in the vasoactive intestinal peptide receptor 2-deficient mice, despite the substantial deterioration or abolition of circadian behavioural rhythms. These findings indicate that the circadian clock in the OB of freely moving mice depends on the SCN master clock but not on the circadian behavioural rhythms. The circadian PER2::LUC rhythm in the cultured OB was as robust as that in the cultured SCN but reset by slice preparation, suggesting that culturing of the slice reinforces the circadian rhythm.

Effect of Suprachiasmatic Nucleus Lesion on Period2 and C-fos Expression in Habenular Nucleus

The habenular nucleus (Hb) located in the epithalamus is an important connection between the limbic forebrain and midbrain pathway and it regulates physiological activities including reproduction, sleep-wake, body temperature. Hb is anatomically close to the suprachiasmatic nucleus (SCN), the pacemaker of circadian rhythms in mammals. In vivo data showed circadian oscillation of Period2 (Per2) expression in Hb. However, it remains unknown whether the circadian oscillation of Per2 expression is regulated by SCN. We used semi-quantity RT-PCR methods to examine the relative expression of Per2 and c-fos mRNA in the Hb at 13:00 and 1:00 in the LD (light-dark) cycle either SCN was intact or destroyed. The expression of Per2 and c-fos mRNA in the Hb was higher at 13:00 than at 1:00. However, damaging the SCN abolished the difference of Per2 and c-fos mRNA expression at the two time ponits in the Hb, suggesting circadian oscillation of Per2 and c-fos gene in the Hb may be regulated by SCN under in vivo condition.

In Vivo Imaging of the Central and Peripheral Effects of Sleep Deprivation and Suprachiasmatic Nuclei Lesion on PERIOD-2 Protein in Mice.

That sleep deprivation increases the brain expression of various clock genes has been well documented. Based on these and other findings we hypothesized that clock genes not only underlie circadian rhythm generation but are also implicated in sleep homeostasis. However, long time lags have been reported between the changes in the clock gene messenger RNA levels and their encoded proteins. It is therefore crucial to establish whether also protein levels increase within the time frame known to activate a homeostatic sleep response. We report on the central and peripheral effects of sleep deprivation on PERIOD-2 (PER2) protein both in intact and suprachiasmatic nuclei-lesioned mice. In vivo and in situ PER2 imaging during baseline, sleep deprivation, and recovery. Mouse sleep-recording facility. Per2::Luciferase knock-in mice. N/A. Six-hour sleep deprivation increased PER2 not only in the brain but also in liver and kidney. Remarkably, the effects in the liver outlasted those observed in the brain. Within the brain the increase in PER2 concerned the cerebral cortex mainly, while leaving suprachiasmatic nuclei (SCN) levels unaffected. Against expectation, sleep deprivation did not increase PER2 in the brain of arrhythmic SCN-lesioned mice because of higher PER2 levels in baseline. In contrast, liver PER2 levels did increase in these mice similar to the sham and partially lesioned controls. Our results stress the importance of considering both sleep-wake dependent and circadian processes when quantifying clock-gene levels. Because sleep deprivation alters PERIOD-2 in the brain as well as in the periphery, it is tempting to speculate that clock genes constitute a common pathway mediating the shared and well-known adverse effects of both chronic sleep loss and disrupted circadian rhythmicity on metabolic health.

The light-dark cycle controls peripheral rhythmicity in mice with a genetically ablated suprachiasmatic nucleus clock.

The mammalian circadian timing system consists of a master pacemaker in the suprachiasmatic nucleus (SCN), which is thought to synchronize peripheral clocks in various organs with each other and with external time. Our knowledge about the role of the SCN clock is based mainly on SCN lesion and transplantation studies. We have now directly deleted the SCN clock using the Cre/LoxP system and investigated how this affects synchronization of peripheral rhythms. Impaired locomotor activity and arrhythmic clock gene expression in the SCN confirm that the SCN clockwork was efficiently abolished in our mouse model. Nonetheless, under light-dark (LD) conditions, peripheral clocks remained rhythmic and synchronized to the LD cycle, and phase relationships between peripheral clocks were sustained. Adaptation to a shifted LD cycle was accelerated in SCN clock-deficient mice. Moreover, under zeitgeber-free conditions, rhythmicity of the peripheral clock gene expression was initially dampened, and after several days peripheral clocks were desynchronized. These findings suggest that the SCN clock is dispensable for the synchronization of peripheral clocks to the LD cycle. A model describing an SCN clock-independent pathway that synchronizes peripheral clocks with the LD cycle is discussed.