NPAS4 regulates the transcriptional response of the suprachiasmatic nucleus to light and circadian behavior.

The suprachiasmatic nucleus (SCN) is the master circadian pacemaker in mammals and is entrained by environmental light. However, the molecular basis of the response of the SCN to light is not fully understood. We used RNA/chromatin immunoprecipitation/single-nucleus sequencing with circadian behavioral assays to identify mouse SCN cell types and explore their responses to light. We identified three peptidergic cell types that responded to light in the SCN: arginine vasopressin (AVP), vasoactive intestinal peptide (VIP), and cholecystokinin (CCK). In each cell type, light-responsive subgroups were enriched for expression of neuronal Per-Arnt-Sim (PAS) domain protein 4 (NPAS4) target genes. Further, mice lacking Npas4 had a longer circadian period under constant conditions, a damped phase response curve to light, and reduced light-induced gene expression in the SCN. Our data indicate that NPAS4 is necessary for normal transcriptional responses to light in the SCN and critical for photic phase-shifting of circadian behavior.

A genome-wide microRNA screen identifies the microRNA-183/96/182 cluster as a modulator of circadian rhythms.

The regulatory mechanisms of circadian rhythms have been studied primarily at the level of the transcription-translation feedback loops of protein-coding genes. Regulatory modules involving noncoding RNAs are less thoroughly understood. In particular, emerging evidence has revealed the important role of microRNAs (miRNAs) in maintaining the robustness of the circadian system. To identify miRNAs that have the potential to modulate circadian rhythms, we conducted a genome-wide miRNA screen using U2OS luciferase reporter cells. Among 989 miRNAs in the library, 120 changed the period length in a dose-dependent manner. We further validated the circadian regulatory function of an miRNA cluster, miR-183/96/182, both in vitro and in vivo. We found that all three members of this miRNA cluster can modulate circadian rhythms. Particularly, miR-96 directly targeted a core circadian clock gene, PER2. The knockout of the miR-183/96/182 cluster in mice showed tissue-specific effects on circadian parameters and altered circadian rhythms at the behavioral level. This study identified a large number of miRNAs, including the miR-183/96/182 cluster, as circadian modulators. We provide a resource for further understanding the role of miRNAs in the circadian network and highlight the importance of miRNAs as a genome-wide layer of circadian clock regulation.

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.

Double recording system of Period1 gene expression rhythm in the olfactory bulb and liver in freely moving mouse.

Clock genes express circadian rhythms in most organs. These rhythms are organized throughout the whole body, regulated by the suprachiasmatic nucleus (SCN) in the brain. Disturbance of these clock gene expression rhythms is a risk factor for diseases such as obesity and cancer. To understand the mechanism of regulating clock gene expression rhythms in vivo, multiple real time recording systems are required. In the present study, we developed a double recording system of Period1 expression rhythm in peripheral tissue (liver) and the brain. In peripheral tissue, quantification of gene expression in a steadily moving target was achieved by using a photomultiplier tube (PMT) attached to a tissue contact optical sensor (TCS). Using this technique, we were able to analyze circadian rhythms of clock gene expression over a prolonged period in the liver and olfactory bub (OB) of the brain. The present double recording system has no effect on behavioral activity or rhythm. Our novel system thus successfully quantifies clock gene expression in deep areas of the body in freely moving mice for a period sufficient to analyze circadian dynamics. In addition, our double recording system can be widely applied to many areas of biomedical research, as well as applications beyond medicine.

Vasoactive intestinal peptide controls the suprachiasmatic circadian clock network via ERK1/2 and DUSP4 signalling.

The suprachiasmatic nucleus (SCN) co-ordinates circadian behaviour and physiology in mammals. Its cell-autonomous circadian oscillations pivot around a well characterised transcriptional/translational feedback loop (TTFL), whilst the SCN circuit as a whole is synchronised to solar time by its retinorecipient cells that express and release vasoactive intestinal peptide (VIP). The cell-autonomous and circuit-level mechanisms whereby VIP synchronises the SCN are poorly understood. We show that SCN slices in organotypic culture demonstrate rapid and sustained circuit-level circadian responses to VIP that are mediated at a cell-autonomous level. This is accompanied by changes across a broad transcriptional network and by significant VIP-directed plasticity in the internal phasing of the cell-autonomous TTFL. Signalling via ERK1/2 and tuning by its negative regulator DUSP4 are critical elements of the VIP-directed circadian re-programming. In summary, we provide detailed mechanistic insight into VIP signal transduction in the SCN at the level of genes, cells and neural circuit.

A-to-I RNA editing enzyme ADAR2 regulates light-induced circadian phase-shift.

In mammals, the central circadian clock is located in the suprachiasmatic nucleus (SCN) of the hypothalamus and it orchestrates peripheral clocks in the whole body to organize physiological and behavioral rhythms. Light-induced phase-shift of the SCN clock enables synchronization of the circadian clock system with 24-h environmental light/dark cycle. We previously found that adenosine deaminase acting on RNA 2 (Adar2), an A-to-I RNA editing enzyme catalyzing rhythmic A-to-I RNA editing, governs a wide range of mRNA rhythms in the mouse liver and regulates the circadian behavior. In brain, ADAR2-mediated A-to-I RNA editing was reported to occur in various transcripts encoding ion channels and neurotransmitter receptors, which could influence neuronal function of the SCN. Here we show that ADAR2 plays a crucial role for light-induced phase-shift of the circadian clock. Intriguingly, exposure of Adar2-knockout mice to a light pulse at late night caused an aberrant phase-advance of the locomotor rhythms. By monitoring the bioluminescence rhythms of the mutant SCN slices, we found that a phase-advance induced by treatment with pituitary adenylyl cyclase-activating polypeptide (PACAP) was markedly attenuated. The present study suggests that A-to-I RNA editing in the SCN regulates a proper phase response to light in the mouse circadian system.

The Effect of Light Exposure at Night (LAN) on Carcinogenesis via Decreased Nocturnal Melatonin Synthesis.

In mammals, a master clock is located within the suprachiasmatic nucleus (SCN) of the hypothalamus, a region that receives input from the retina that is transmitted by the retinohypothalamic tract. The SCN controls the nocturnal synthesis of melatonin by the pineal gland that can influence the activity of the clock's genes and be involved in the inhibition of cancer development. On the other hand, in the literature, some papers highlight that artificial light exposure at night (LAN)-induced circadian disruptions promote cancer. In the present review, we summarize the potential mechanisms by which LAN-evoked disruption of the nocturnal increase in melatonin synthesis counteracts its preventive action on human cancer development and progression. In detail, we discuss: (i) the Warburg effect related to tumor metabolism modification; (ii) genomic instability associated with L1 activity; and (iii) regulation of immunity, including regulatory T cell (Treg) regulation and activity. A better understanding of these processes could significantly contribute to new treatment and prevention strategies against hormone-related cancer types.

Estimations of Retinal Blue-Light Irradiance Values and Melatonin Suppression Indices Through Clear and Yellow-Tinted Intraocular Lenses.

Spectral transmittance values in the wavelength range of 300 to 800 nanometers were measured using a spectrophotometer for 18 intraocular lenses (IOLs) including clear (ZCB00) and yellow-tinted (ZCB00V, both from AMO Japan) IOLs with three different lens powers. Also measured were the blue-light irradiance (BLI) values, which might reflect retinal damage caused by sunlight, and the melatonin suppression indices (MSIs), which might reflect the nonvisual photoreception function, through these IOLs. The BLIs (in mWcm-2) calculated were 7.62, 7.50, and 7.46 for the +10-diopter (D), +20-D, and +30-D ZCB00 IOLs, respectively; 4.10, 3.92, and 4.00 for the +10-D, +20-D, and +30-D ZCB00V IOLs, respectively; 5.76 for phakic eyes; and 15.00 for aphakic eyes. The MSIs (in mWcm-2sr-1) calculated were 1.18, 1.19, and 1.18 for the +10-D, +20-D, and +30-D ZCB00 IOLs, respectively; 0.98, 0.94, and 0.95 for the +10-D, +20-D, and +30-D ZCB00V IOLs, respectively; 1.03 for phakic eyes; and 1.21 for aphakic eyes. The data from the six clear IOLs (SA60AT, Alcon Japan; VA-60BBR, Hoya; AU6 K, Kowa, N4-18B, Nidek; X-60, Santen; KS-3Ai, Staar Japan) and seven yellow-tinted IOLs (SN60AT; YA-60BBR, Hoya; AU6N, Kowa; N4-18YG, Nidek; NX-60, Santen; KS-AiN, Staar Japan; XY-1, Hoya) reported previously also were discussed. Compared to aphakic eyes, ZCB00 and ZCB00V IOLs reduce the BLI values by 49-50% and 73-74%, respectively; and currently available ultraviolet-blocking clear and yellow-tinted IOLs reduce the BLI values by 43-82%, respectively. Yellow-tinted IOLs absorb more circadian rhythm-associated light than clear IOLs. Although the data presented in this study cannot be applied directly to IOL implanted in patients, the balance between photoprotection and photoreception must be considered when using IOLs in a clinical setting.

Constant light during lactation programs circadian and metabolic systems.

Exposure to light at night is a disruptive condition for the adult circadian system, leading to arrhythmicity in nocturnal rodents. Circadian disruption is a risk factor for developing physiological and behavioral alterations, including weight gain and metabolic disease. During early stages of development, the circadian system undergoes a critical period of adjustment, and it is especially vulnerable to altered lighting conditions that may program its function, leading to long-term effects. We hypothesized that during lactation a disrupted light-dark cycle due to light at night may disrupt the circadian system and in the long term induce metabolic disorders. Here we explored in pups, short- and long-term effects of constant light (LL) during lactation. In the short term, LL caused a loss of rhythmicity and a reduction in the immunopositive cells of VIP, AVP, and PER1 in the suprachiasmatic nucleus (SCN). In the short term, the affection on the circadian clock in the pups resulted in body weight gain, loss of daily rhythms in general activity, plasma glucose and triglycerides (TG). Importantly, the DD conditions during development also induced altered daily rhythms in general activity and in the SCN. Exposure to LD conditions after lactation did not restore rhythmicity in the SCN, and the number of immunopositve cells to VIP, AVP, and PER1 remained reduced. In the long term, daily rhythmicity in general activity was restored; however, daily rhythms in glucose and TG remained disrupted, and daily mean levels of TG were significantly increased. Present results point out the programming role played by the LD cycle during early development in the function of the circadian system and on metabolism. This study points out the risk represented by exposure to an altered light-dark cycle during early stages of development. ABBREVIATIONS: AVP: arginine vasopressin peptide; CRY: cryptochrome; DD: constant darkness; DM: dorsomedial; LD: light-dark cycle; LL: constant light; NICUs: neonatal intensive care units; P: postnatal days; PER: period; S.E.M.: standard error of the mean; SCN: suprachiasmatic nucleus; TG: triglycerides; VIP: vasointestinal peptide; VL: ventrolateral; ZT: zeitgeber time.

Scheduled feeding restores memory and modulates c-Fos expression in the suprachiasmatic nucleus and septohippocampal complex.

Disruptions in circadian timing impair spatial memory in humans and rodents. Circadian-arrhythmic Siberian hamsters (Phodopus sungorus) exhibit substantial deficits in spatial working memory as assessed by a spontaneous alternation (SA) task. The present study found that daily scheduled feeding rescued spatial memory deficits in these arrhythmic animals. Improvements in memory persisted for at least 3 weeks after the arrhythmic hamsters were switched back to ad libitum feeding. During ad libitum feeding, locomotor activity resumed its arrhythmic state, but performance on the SA task varied across the day with a peak in daily performance that corresponded to the previous daily window of food anticipation. At the end of scheduled feeding, c-Fos brain mapping revealed differential gene expression in entrained versus arrhythmic hamsters in the suprachiasmatic nucleus (SCN) that paralleled changes in the medial septum and hippocampus, but not in other neural structures. These data show that scheduled feeding can improve cognitive performance when SCN timing has been compromised, possibly by coordinating activity in the SCN and septohippocampal pathway.

Melanopsin Regulates Both Sleep-Promoting and Arousal-Promoting Responses to Light.

Light plays a critical role in the regulation of numerous aspects of physiology and behaviour, including the entrainment of circadian rhythms and the regulation of sleep. These responses involve melanopsin (OPN4)-expressing photosensitive retinal ganglion cells (pRGCs) in addition to rods and cones. Nocturnal light exposure in rodents has been shown to result in rapid sleep induction, in which melanopsin plays a key role. However, studies have also shown that light exposure can result in elevated corticosterone, a response that is not compatible with sleep. To investigate these contradictory findings and to dissect the relative contribution of pRGCs and rods/cones, we assessed the effects of light of different wavelengths on behaviourally defined sleep. Here, we show that blue light (470 nm) causes behavioural arousal, elevating corticosterone and delaying sleep onset. By contrast, green light (530 nm) produces rapid sleep induction. Compared to wildtype mice, these responses are altered in melanopsin-deficient mice (Opn4-/-), resulting in enhanced sleep in response to blue light but delayed sleep induction in response to green or white light. We go on to show that blue light evokes higher Fos induction in the SCN compared to the sleep-promoting ventrolateral preoptic area (VLPO), whereas green light produced greater responses in the VLPO. Collectively, our data demonstrates that nocturnal light exposure can have either an arousal- or sleep-promoting effect, and that these responses are melanopsin-mediated via different neural pathways with different spectral sensitivities. These findings raise important questions relating to how artificial light may alter behaviour in both the work and domestic setting.

Ras Activity Oscillates in the Mouse Suprachiasmatic Nucleus and Modulates Circadian Clock Dynamics.

Circadian rhythms, generated in the mouse suprachiasmatic nucleus (SCN), are synchronized to the environmental day-night changes by photic input. The activation of the extracellular signal-regulated kinases 1 and 2 (ERK1,2) and cAMP response element-binding protein (CREB)-mediated transcription play a critical role in this photoentrainment. The small GTPase Ras is one of the major upstream regulators of the ERK1,2/CREB pathway. In contrast to the well-described role of Ras in structural and functional synaptic plasticity in the adult mouse brain, the physiological regulation of Ras by photic sensory input is yet unknown. Here, we describe for the first time a circadian rhythm of Ras activity in the mouse SCN. Using synRas transgenic mice, expressing constitutively activated V12-Ha-Ras selectively in neurons, we demonstrate that enhanced Ras activation causes shortening of the circadian period length. We found upregulated expression and decreased inhibitory phosphorylation of the circadian period length modulator, glycogen synthase kinase-3 beta (GSK3ß), in the SCN of synRas mice. Conversely, downregulation of Ras activity by blocking its function with an antibody in oscillating cell cultures reduced protein levels and increased phosphorylation of GSK3ß and lengthened the period of BMAL1 promoter-driven luciferase activity. Furthermore, enhanced Ras activity in synRas mice resulted in a potentiation of light-induced phase delays at early subjective night, and increased photic induction of pERK1,2/pCREB and c-Fos. In contrast, at late subjective night, photic activation of Ras/ERK1,2/CREB in synRas mice was not sufficient to stimulate c-Fos protein expression and phase advance the clock. Taken together, our results demonstrate that Ras activity fine tunes the period length and modulates photoentrainment of the circadian clock.

Constant light enhances synchrony among circadian clock cells and promotes behavioral rhythms in VPAC2-signaling deficient mice.

Individual neurons in the suprachiasmatic nuclei (SCN) contain an intracellular molecular clock and use intercellular signaling to synchronize their timekeeping activities so that the SCN can coordinate brain physiology and behavior. The neuropeptide vasoactive intestinal polypeptide (VIP) and its VPAC2 receptor form a key component of intercellular signaling systems in the SCN and critically control cellular coupling. Targeted mutations in either the intracellular clock or intercellular neuropeptide signaling mechanisms, such as VIP-VPAC2 signaling, can lead to desynchronization of SCN neuronal clocks and loss of behavioral rhythms. An important goal in chronobiology is to develop interventions to correct deficiencies in circadian timekeeping. Here we show that extended exposure to constant light promotes synchrony among SCN clock cells and the expression of ~24 h rhythms in behavior in mice in which intercellular signaling is disrupted through loss of VIP-VPAC2 signaling. This study highlights the importance of SCN synchrony for the expression of rhythms in behavior and reveals how non-invasive manipulations in the external environment can be used to overcome neurochemical communication deficits in this important brain system.

Temporal changes of light-induced proteins in the SCN following treatment with the serotonin mixed agonist/antagonist BMY7378.

The 5-HT1A mixed agonist/antagonist BMY7378 has been shown to greatly potentiate photic phase advances in hamsters. The underlying mechanism and intracellular changes in the suprachiasmatic nucleus (SCN) by which this potentiation is accomplished have yet to be fully determined. Here, we examine the effect of BMY7378 on temporal activation patterns of a number of proteins and enzymes in the SCN following light exposure in the late subjective night. BMY7378 administration increased the amount of several photo-inducible proteins in the SCN at specific time points following light exposure in the late subjective night. Relative to animals given saline before a light pulse, the number of cells immunoreactive for cFos, JunB and PER1 was all significantly greater 360 min following the light pulse in BMY7378 pretreated animals, indicating an extended action of these light-induced proteins in the SCN following BMY7378 pretreatment. Aside from a modest, nonsignificant increase in P-ERK levels at 60 min, BMY7378 did not affect light-induced P-ERK levels. The levels of light-induced P-CREB were similarly unaffected by BMY7378. Also unaffected by BMY7378 treatment were cFos expression and JunB expression at 120 and 180 min following light exposure. These findings suggest that BMY7378 may potentiate photic phase shifts at least partly by prolonging the activity of some, but not all, light-induced proteins and biochemical pathways involved in coupling the light signal to the output of the circadian clock, particularly those which are active many hours after the light signal reaches the SCN.

CRTC2 activation in the suprachiasmatic nucleus, but not paraventricular nucleus, varies in a diurnal fashion and increases with nighttime light exposure.

Entrainment of the intrinsic suprachiasmatic nucleus (SCN) molecular clock to the light-dark cycle depends on photic-driven intracellular signal transduction responses of SCN neurons that converge on cAMP response element-binding protein (CREB)-mediated regulation of gene transcription. Characterization of the CREB coactivator proteins CREB-regulated transcriptional coactivators (CRTCs) has revealed a greater degree of differential activity-dependent modulation of CREB transactivational function than previously appreciated. In confirmation of recent reports, we found an enrichment of crtc2 mRNA and prominent CRTC2 protein expression within the SCN of adult male rats. With use of a hypothalamic organotypic culture preparation for initial CRTC2-reactive antibody characterization, we found that CRTC2 immunoreactivity in hypothalamic neurons shifted from a predominantly cytoplasmic profile under basal culture conditions to a primarily nuclear localization (CRTC2 activation) 30 min after adenylate cyclase stimulation. In adult rat SCN, we found a diurnal variation in CRTC2 activation (peak at zeitgeber time of 4 h and trough at zeitgeber time of 16-20 h) but no variation in the total number of CRTC2-immunoreactive cells. There was no diurnal variation of CRTC2 activation in the hypothalamic paraventricular nucleus, another site of enriched CRTC2 expression. Exposure of rats to light (50 lux) for 30 min during the second half of their dark (nighttime) phase produced CRTC2 activation. We observed in the SCN a parallel change in the expression of a CREB-regulated gene (FOS). In contrast, nighttime light exposure had no effect on CRTC2 activation or FOS expression in the paraventricular nucleus, nor did it affect corticosterone hormone levels. These results suggest that CRTC2 participates in CREB-dependent photic entrainment of SCN function.

Melatonin and the circadian system: contributions to successful female reproduction.

OBJECTIVE: To summarize the role of melatonin and circadian rhythms in determining optimal female reproductive physiology, especially at the peripheral level. DESIGN: Databases were searched for the related English-language literature published up to March 1, 2014. Only papers in peer-reviewed journals are cited. SETTING: Not applicable. PATIENT(S): Not applicable. INTERVENTION(S): Melatonin treatment, alterations of the normal light:dark cycle and light exposure at night. MAIN OUTCOME MEASURE(S): Melatonin levels in the blood and in the ovarian follicular fluid and melatonin synthesis, oxidative damage and circadian rhythm disturbances in peripheral reproductive organs. RESULT(S): The central circadian regulatory system is located in the suprachiasmatic nucleus (SCN). The output of this master clock is synchronized to 24 hours by the prevailing light-dark cycle. The SCN regulates rhythms in peripheral cells via the autonomic nervous system and it sends a neural message to the pineal gland where it controls the cyclic production of melatonin; after its release, the melatonin rhythm strengthens peripheral oscillators. Melatonin is also produced in the peripheral reproductive organs, including granulosa cells, the cumulus oophorus, and the oocyte. These cells, along with the blood, may contribute melatonin to the follicular fluid, which has melatonin levels higher than those in the blood. Melatonin is a powerful free radical scavenger and protects the oocyte from oxidative stress, especially at the time of ovulation. The cyclic levels of melatonin in the blood pass through the placenta and aid in the organization of the fetal SCN. In the absence of this synchronizing effect, the offspring may exhibit neurobehavioral deficits. Also, melatonin protects the developing fetus from oxidative stress. Melatonin produced in the placenta likewise may preserve the optimal function of this organ. CONCLUSION(S): Both stable circadian rhythms and cyclic melatonin availability are critical for optimal ovarian physiology and placental function. Because light exposure after darkness onset at night disrupts the master circadian clock and suppresses elevated nocturnal melatonin levels, light at night should be avoided.

Introduction: circadian rhythm and its disruption: impact on reproductive function.

Almost all forms of life have predictable daily or circadian rhythms in molecular, endocrine, and behavioral functions. In mammals, a central pacemaker located in the suprachiasmatic nuclei coordinates the timing of these rhythms. Daily light exposure that affects the retina of the eye directly influences this area, which is required to align endogenous processes to the appropriate time of day. The present "Views and Reviews" articles discuss the influence of circadian rhythms, especially nightly secretion of melatonin, on reproductive function and parturition. In addition, an examination is made of problems that arise from recurrent circadian rhythm disruption associated with changes in light exposure patterns common to modern day society. Finally, a possible solution to prevent disruptions in circadian phase markers by filtering out short wavelengths from nocturnal light is reviewed.

Modulation of mammalian circadian rhythms by tumor necrosis factor-α.

UNLABELLED: Systemic low doses of the endotoxin lipopolysaccharide (LPS, 100 µg/kg) administered during the early night induce phase-delays of locomotor activity rhythms in mice. Our aim was to evaluate the role of tumor necrosis factor (Tnf)-alpha and its receptor 1/p55 (Tnfr1) in the modulation of LPS-induced circadian effects on the suprachiasmatic nucleus (SCN). We observed that Tnfr1-defective mice (Tnfr1 KO), although exhibiting similar circadian behavior and light response to that of control mice, did not show LPS-induced phase-delays of locomotor activity rhythms, nor LPS-induced cFos and Per2 expression in the SCN and Per1 expression in the paraventricular hypothalamic nucleus (PVN) as compared to wild-type (WT) mice. We also analyzed Tnfr1 expression in the SCN of WT mice, peaking during the early night, when LPS has a circadian effect. Peripheral inoculation of LPS induced an increase in cytokine/chemokine levels (Tnf, Il-6 and Ccl2) in the SCN and in the PVN. In conclusion, in this study, we show that LPS-induced circadian responses are mediated by Tnf. Our results also suggest that this cytokine stimulates the SCN after LPS peripheral inoculation; and the time-related effect of LPS (i.e. phase shifts elicited only at early night) might depend on the increased levels of Tnfr1 expression. We also confirmed that LPS modulates clock gene expression in the SCN and PVN in WT but not in Tnfr1 KO mice. HIGHLIGHTS: We demonstrate a fundamental role for Tnf and its receptor in circadian modulation by immune stimuli at the level of the SCN biological clock.

Photic sensitivity for circadian response to light varies with photoperiod.

The response of the circadian system to light varies markedly depending on photic history. Under short day lengths, hamsters exhibit larger maximal light-induced phase shifts as compared with those under longer photoperiods. However, effects of photoperiod length on sensitivity to subsaturating light remain unknown. Here, Syrian hamsters were entrained to long or short photoperiods and subsequently exposed to a 15-min light pulse across a range of irradiances (0-68.03 µW/cm(2)) to phase shift activity rhythms. Phase advances exhibited a dose response, with increasing irradiances eliciting greater phase resetting in both conditions. Photic sensitivity, as measured by the half-saturation constant, was increased 40-fold in the short photoperiod condition. In addition, irradiances that generated similar phase advances under short and long days produced equivalent phase delays, and equal photon doses produced larger delays in the short photoperiod condition. Mechanistically, equivalent light exposure induced greater pERK, PER1, and cFOS immunoreactivity in the suprachiasmatic nuclei of animals under shorter days. Patterns of immunoreactivity in all 3 proteins were related to the size of the phase shift rather than the intensity of the photic stimulus, suggesting that photoperiod modulation of light sensitivity lies upstream of these events within the signal transduction cascade. This modulation of light sensitivity by photoperiod means that considerably less light is necessary to elicit a circadian response under the relatively shorter days of winter, extending upon the known seasonal changes in sensitivity of sensory systems. Further characterizing the mechanisms by which photoperiod alters photic response may provide a potent tool for optimizing light treatment for circadian and affective disorders in humans.

Effect of experimental glaucoma on the non-image forming visual system.

Glaucoma is a leading cause of blindness worldwide, characterized by retinal ganglion cell degeneration and damage to the optic nerve. We investigated the non-image forming visual system in an experimental model of glaucoma in rats induced by weekly injections of chondroitin sulphate (CS) in the eye anterior chamber. Animals were unilaterally or bilaterally injected with CS or vehicle for 6 or 10 weeks. In the retinas from eyes injected with CS, a similar decrease in melanopsin and Thy-1 levels was observed. CS injections induced a similar decrease in the number of melanopsin-containing cells and superior collicular retinal ganglion cells. Experimental glaucoma induced a significant decrease in the afferent pupil light reflex. White light significantly decreased nocturnal pineal melatonin content in control and glaucomatous animals, whereas blue light decreased this parameter in vehicle- but not in CS-injected animals. A significant decrease in light-induced c-Fos expression in the suprachiasmatic nuclei was observed in glaucomatous animals. General rhythmicity and gross entrainment appear to be conserved, but glaucomatous animals exhibited a delayed phase angle with respect to lights off and a significant increase in the percentage of diurnal activity. These results indicate the glaucoma induced significant alterations in the non-image forming visual system.