Peripheral Oscillators Research Articles

Social stress and glucocorticoids alter PERIOD2 rhythmicity in the liver, but not in the suprachiasmatic nucleus

Circadian (~24 h) rhythms in behavior and physiological functions are under control of an endogenous circadian pacemaker in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN directly drives some of these rhythms or serves as a coordinator of peripheral oscillators residing in other tissues and organs. Disruption of the circadian organization may contribute to disease, including stress-related disorders. Previous research indicates that the master clock in the SCN is resistant to stress, although it is unclear whether stress affects rhythmicity in other tissues, possibly mediated by glucocorticoids, released in stressful situations. In the present study, we examined the effect of uncontrollable social defeat stress and glucocorticoid hormones on the central and peripheral clocks, respectively in the SCN and liver. Transgenic PERIOD2::LUCIFERASE knock-in mice were used to assess the rhythm of the clock protein PERIOD2 (PER2) in SCN slices and liver tissue collected after 10 consecutive days of social defeat stress. The rhythmicity of PER2 expression in the SCN was not affected by stress exposure, whereas in the liver the expression showed a delayed phase in defeated compared to non-defeated control mice. In a second experiment, brain slices and liver samples were collected from transgenic mice and exposed to different doses of corticosterone. Corticosterone did not affect PER2 rhythm of the SCN samples, but caused a phase shift in PER2 expression in liver samples. This study confirms earlier findings that the SCN is resistant to stress and shows that clocks in the liver are affected by social stress, which might be due to the direct influence of glucocorticoids released from the adrenal gland.

SIRT7 couples light-driven body temperature cues to hepatic circadian phase coherence and gluconeogenesis

The central pacemaker in the hypothalamic suprachiasmatic nucleus (SCN) synchronizes peripheral oscillators to coordinate physiological and behavioural activities throughout the body. How circadian phase coherence between the SCN and the periphery is controlled is not well understood. Here, we identify hepatic SIRT7 as an early responsive element to light that ensures circadian phase coherence in the mouse liver. The SCN-driven body temperature (BT) oscillation induces rhythmic expression of HSP70, which promotes SIRT7 ubiquitination and proteasomal degradation. Acute temperature challenge dampens the BT oscillation and causes an advanced liver circadian phase. Further, hepatic SIRT7 deacetylates CRY1, promotes its FBXL3-mediated degradation and regulates the hepatic clock and glucose homeostasis. Loss of Sirt7 in mice leads to an advanced liver circadian phase and rapid entrainment of the hepatic clock upon daytime-restricted feeding. These data identify a BT-HSP70-SIRT7-CRY1 axis that couples the mouse hepatic clock to the central pacemaker and ensures circadian phase coherence and glucose homeostasis.

Desipramine restores the alterations in circadian entrainment induced by prenatal exposure to glucocorticoids

Alterations in circadian rhythms are closely linked to depression, and we have shown earlier that progressive alterations in circadian entrainment precede the onset of depression in mice exposed in utero to excess glucocorticoids. The aim of this study was to investigate whether treatment with the noradrenaline reuptake inhibitor desipramine (DMI) could restore the alterations in circadian entrainment and prevent the onset of depression-like behavior. C57Bl/6 mice were exposed to dexamethasone (DEX—synthetic glucocorticoid analog, 0.05 mg/kg/day) between gestational day 14 and delivery. Male offspring aged 6 months (mo) were treated with DMI (10 mg/kg/day in drinking water) for at least 21 days before behavioral testing. We recorded spontaneous activity using the TraffiCage™ system and found that DEX mice re-entrained faster than controls after an abrupt advance in light-dark cycle by 6 h, while DMI treatment significantly delayed re-entrainment. Next we assessed the synchronization of peripheral oscillators with the central clock (located in the suprachiasmatic nucleus—SCN), as well as the mechanisms required for entrainment. We found that photic entrainment of the SCN was apparently preserved in DEX mice, but the expression of clock genes in the hippocampus was not synchronized with the light-dark cycle. This was associated with downregulated mRNA expression for arginine vasopressin (AVP; the main molecular output entraining peripheral clocks) in the SCN, and for glucocorticoid receptor (GR; required for the negative feedback loop regulating glucocorticoid secretion) in the hippocampus. DMI treatment restored the mRNA expression of AVP in the SCN and enhanced GR-mediated signaling by upregulating GR expression and nuclear translocation in the hippocampus. Furthermore, DMI treatment at 6 mo prevented the onset of depression-like behavior and the associated alterations in neurogenesis in 12-mo-old DEX mice. Taken together, our data indicate that DMI treatment enhances GR-mediated signaling and restores the synchronization of peripheral clocks with the SCN and support the hypothesis that altered circadian entrainment is a modifiable risk factor for depression.

Dormancy, Diapause, and the Role of the Circadian System in Insect Photoperiodism.

Whole-animal experiments devised to investigate possible association between photoperiodic time measurement and the circadian system (Bünning's hypothesis) are compared with more recent molecular investigations of circadian clock genes. In Sarcophaga argyrostoma and some other species, experimental cycles of light and darkness revealed a photoperiodic oscillator, set to constant phase at dusk and measuring night length repeatedly during extended periods of darkness. In some species, however, extreme dampening revealed an unrepetitive (i.e., hourglass-like) response. Rhythms of clock gene transcript abundance may also show similar phase relationships to the light cycle, and gene silencing of important clock genes indicates that they play a crucial role in photoperiodism either alone or in concert. However, the multiplicity of peripheral oscillators in the insect circadian system indicates that more complex mechanisms might also be important.

Modeling Brain Cognitive Functions by Oscillatory Neural Networks

We describe an oscillatory neural network designed as a system of generalized phase oscillators with a central element. It is shown that a winner-take-all principle can be realized in this system in terms of the competition of peripheral oscillators for the synchronization with a central oscillator. Several examples illustrate how this network can be used for the simulation of various cognitive functions: consecutive selection of objects in the image, visual search, and multiple object tracking.

Effect of melatonin on monochromatic light-induced changes in clock gene circadian expression in the chick liver

Light is the most prominent zeitgeber of the circadian system, which contains central and peripheral oscillators. Our previous studies found that light wavelength could influence the rhythms of melatonin synthesis and clock gene expression in the central oscillator of chicks. However, the effect of monochromatic light on the peripheral oscillator and the role of melatonin have yet to be clarified. In this study, 216 newly hatched chicks were divided into three groups (intact, sham operation and pinealectomy) and were raised under white (WL), red (RL), green (GL) or blue (BL) light for 14 days. Their plasma and livers were sampled at 6 time points with 4-h intervals. Plasma melatonin concentration and liver clock gene expression (cClock, cBmal1, cBmal2, cCry1, cCry2, cPer2, cPer3) were measured for circadian rhythm analysis. In intact and sham operation chicks under WL, all liver clock genes showed circadian expression along with oscillations in plasma melatonin. However, positive clock genes peaked at subjective night along with melatonin, while negative clock genes peaked at subjective day or the shifting time of day-night. Chick exposure to monochromatic light led to an unaltered circadian rhythmicity in plasma melatonin and liver clock genes; however, their rhythmic parameters were notably influenced. Compared to WL, GL enhanced the mesor and amplitude of melatonin and all kinds of clock genes, whereas RL had the opposite effect. Pinealectomy significantly decreased expression of liver clock genes, which was consistent with the reduction in plasma melatonin concentration, especially for the GL group, and resulted in the expression of liver clock genes showing low-mesor and low-amplitude oscillations as well as no statistically significant differences among the monochromatic light groups. Thus, we speculated that melatonin plays a key role in the effects of light wavelength on clock gene rhythm in the chick liver.

A reductionist, in vitro model of environmental circadian disruption demonstrates SCN-independent and tissue-specific dysregulation of inflammatory responses.

Environmental circadian disruption (ECD), characterized by repeated or long-term disruption in environmental timing cues which require the internal circadian clock to change its phase to resynchronize with the environment, is associated with numerous serious health issues in humans. While animal and isolated cell models exist to study the effects of destabilizing the relationship between the circadian system and the environment, neither approach provides an ideal solution. Here, we developed an in vitro model which incorporates both elements of a reductionist cellular model and disruption of the clock/environment relationship using temperature as an environmental cue, as occurs in vivo. Using this approach, we have demonstrated that some effects of in vivo ECD can be reproduced using only isolated peripheral oscillators. Specifically, we report exaggerated inflammatory responses to endotoxin following repeated environmental circadian disruption in explanted spleens. This effect requires a functional circadian clock but not the master brain clock, the suprachiasmatic nucleus (SCN). Further, we report that this is a result of cumulative, rather than acute, circadian disruption as has been previously observed in vivo. Finally, such effects appear to be tissue specific as it does not occur in lung, which is less sensitive to the temperature cycles employed to induce ECD. Taken together, the present study suggests that this model could be a valuable tool for dissecting the causes and effects of circadian disruption both in isolated components of physiological systems as well as the aggregated interactions of these systems that occur in vivo.

Systems Biology Approaches and Precision Oral Health: A Circadian Clock Perspective.

A vast majority of the pathophysiological and metabolic processes in humans are temporally controlled by a master circadian clock located centrally in the hypothalamic suprachiasmatic nucleus of the brain, as well as by specialized peripheral oscillators located in other body tissues. This circadian clock system generates a rhythmical diurnal transcriptional-translational cycle in clock genes and protein expression and activities regulating numerous downstream target genes. Clock genes as key regulators of physiological function and dysfunction of the circadian clock have been linked to various diseases and multiple morbidities. Emerging omics technologies permits largescale multi-dimensional investigations of the molecular landscape of a given disease and the comprehensive characterization of its underlying cellular components (e.g., proteins, genes, lipids, metabolites), their mechanism of actions, functional networks and regulatory systems. Ultimately, they can be used to better understand disease and interpatient heterogeneity, individual profile, identify personalized targetable key molecules and pathways, discover novel biomarkers and genetic alterations, which collectively can allow for a better patient stratification into clinically relevant subgroups to improve disease prediction and prevention, early diagnostic, clinical outcomes, therapeutic benefits, patient's quality of life and survival. The use of “omics” technologies has allowed for recent breakthroughs in several scientific domains, including in the field of circadian clock biology. Although studies have explored the role of clock genes using circadiOmics (which integrates circadian omics, such as genomics, transcriptomics, proteomics and metabolomics) in human disease, no such studies have investigated the implications of circadian disruption in oral, head and neck pathologies using multi-omics approaches and linking the omics data to patient-specific circadian profiles. There is a burgeoning body of evidence that circadian clock controls the development and homeostasis of oral and maxillofacial structures, such as salivary glands, teeth and oral epithelium. Hence, in the current era of precision medicine and dentistry and patient-centered health care, it is becoming evident that a multi-omics approach is needed to improve our understanding of the role of circadian clock-controlled key players in the regulation of head and neck pathologies. This review discusses current knowledge on the role of the circadian clock and the contribution of omics-based approaches toward a novel precision health era for diagnosing and treating head and neck pathologies, with an emphasis on oral, head and neck cancer and Sjögren's syndrome.

Delayed daily activity and reduced NREM slow-wave power in the APPswe/PS1dE9 mouse model of Alzheimer's disease

Alzheimer's disease (AD) is associated with disrupted circadian rhythms and sleep, which are thought to reflect an impairment of internal circadian timekeeping that contribute to clinical symptoms and disease progression. To evaluate these hypotheses, a suitable preclinical model of AD is needed. We performed a comprehensive assessment of circadian rhythms and sleep in the APPswe/PS1dE9 (APP/PS1) mouse model using long-term in vivo electroencephalogram (EEG) monitoring and behavioral assays from 5 to 22 months of age. APP/PS1 mice were crossed with a PERIOD2::LUCIFERASE (PER2::LUC) mouse model to evaluate synchrony among peripheral circadian oscillators. The APP/PS1 mice exhibited a mild but persistent phase delay of nocturnal activity onset in 12:12h light:dark conditions, as well as a shift toward higher frequencies in the EEG power spectra compared to littermate controls. Our results suggest that APP/PS1 mice may not be the optimal preclinical model for studying the specific circadian changes associated with AD but that quantitative EEG may offer a sensitive measure of AD-associated changes in sleep quality that can be modeled in APP/PS1 mice.

Circadian clock genes' overexpression in Drosophila alters diet impact on lifespan.

Diet restriction is one of the most accurately confirmed interventions which extend lifespan. Genes coding circadian core clock elements are known to be the key controllers of cell metabolism especially in aging aspect. The molecular mechanisms standing behind the phenomenon of diet-restriction-mediated life extension are connected to circadian clock either. Here we investigate the effects of protein-rich and low-protein diets on lifespan observed in fruit flies overexpressing core clock genes (cry, per, Clk, cyc and tim). The majority of core clock genes being upregulated in peripheral tissues (muscles and fat body) on protein-rich diet significantly decrease the lifespan of male fruit flies from 5 to 61%. Nevertheless, positive increments of median lifespan were observed in both sexes, males overexpressing cry in fat body lived 20% longer on poor diet. Overexpression of per also on poor medium resulted in life extension in female fruit flies. Diet restriction reduces mortality caused by overexpression of core clock genes. Cox-regression model revealed that diet restriction seriously decreases mortality risks of flies which overexpress core clock genes. The hazard ratios are lower for flies overexpressing clock genes in fat body relatively to muscle-specific overexpression. The present work suggests a phenomenological view of how two peripheral circadian oscillators modify effects of rich and poor diets on lifespan and hazard ratios.

Nighttime activities and peripheral clock oscillations depend on Wolbachia endosymbionts in flies

Wolbachia are ubiquitous bacterial endosymbionts of arthropods and affect host gene expression. Although Wolbachia infections were suggested to modulate sleep in flies, their influence on the circadian clock remained obscure. Here, we screened bacterial symbionts in a laboratory Drosophila melanogaster colony, and observed widespread infections of wMel strain Wolbachia. We established a Wolbachia-free strain from a clock gene reporter strain, period-luciferase (per-luc). Temperature (19–29 °C)-compensated free-running periods were detected regardless of infections which may reflect the lack of wMel infections in central circadian pacemaker neurons. However, locomotor activity levels during the night or subjective night were significantly amplified in uninfected flies. Moreover, the behavioral phenotype of F1 offspring of an uninfected female and infected male resembled that of uninfected flies. This trait is consistent with maternal transmission of Wolbachia infection. Interestingly, per-luc activities in headless bodies, as an index of peripheral circadian oscillators, were severely damped in uninfected flies. Additionally, circadian amplitudes of PER immunoreactivities in Malpighian tubules were reduced in uninfected flies. These results demonstrate that Wolbachia boost fly peripheral clock oscillations and diurnal behavioral patterns. Genetic mechanisms underlying behavioral rhythms have been widely analyzed using mutant flies whereas screening of Wolbachia will be necessary for future studies.

Insights into the Role of Circadian Rhythms in Bone Metabolism: A Promising Intervention Target?

Numerous physiological processes of mammals, including bone metabolism, are regulated by the circadian clock system, which consists of a central regulator, the suprachiasmatic nucleus (SCN), and the peripheral oscillators of the BMAL1/CLOCK-PERs/CRYs system. Various bone turnover markers and bone metabolism-regulating hormones such as melatonin and parathyroid hormone (PTH) display diurnal rhythmicity. According to previous research, disruption of the circadian clock due to shift work, sleep restriction, or clock gene knockout is associated with osteoporosis or other abnormal bone metabolism, showing the importance of the circadian clock system for maintaining homeostasis of bone metabolism. Moreover, common causes of osteoporosis, including postmenopausal status and aging, are associated with changes in the circadian clock. In our previous research, we found that agonism of the circadian regulators REV-ERBs inhibits osteoclast differentiation and ameliorates ovariectomy-induced bone loss in mice, suggesting that clock genes may be promising intervention targets for abnormal bone metabolism. Moreover, osteoporosis interventions at different time points can provide varying degrees of bone protection, showing the importance of accounting for circadian rhythms for optimal curative effects in clinical treatment of osteoporosis. In this review, we summarize current knowledge about circadian rhythms and bone metabolism.

Time zones of pancreatic islet metabolism.

Most living beings possess an intrinsic system of circadian oscillators, allowing anticipation of the Earth's rotation around its own axis. The mammalian circadian timing system orchestrates nearly all aspects of physiology and behaviour. Together with systemic signals originating from the central clock that resides in the hypothalamic suprachiasmatic nucleus, peripheral oscillators orchestrate tissue-specific fluctuations in gene transcription and translation, and posttranslational modifications, driving overt rhythms in physiology and behaviour. There is accumulating evidence of a reciprocal connection between the circadian oscillator and most aspects of physiology and metabolism, in particular as the circadian system plays a critical role in orchestrating body glucose homeostasis. Recent reports imply that circadian clocks operative in the endocrine pancreas regulate insulin secretion, and that islet clock perturbation in rodents leads to the development of overt type 2 diabetes. While whole islet clocks have been extensively studied during the last years, the heterogeneity of islet cell oscillators and the interplay between α- and β-cellular clocks for orchestrating glucagon and insulin secretion have only recently gained attention. Here, we review recent findings on the molecular makeup of the circadian clocks operative in pancreatic islet cells in rodents and in humans, and focus on the physiologically relevant synchronizers that are resetting these time-keepers. Moreover, the implication of islet clock functional outputs in the temporal coordination of metabolism in health and disease will be highlighted.

Approaches to the Pharmacological Management of Jet Lag.

For many years now a treatment mitigating the debilitating effects of jet lag has been sought. Rapid travel across time zones leads, in most people, to temporary symptoms, in particular poor sleep, daytime alertness and poor performance. Mis-timed circadian rhythms are considered to be the main factor underlying jet-lag symptoms, together with the sleep deprivation from long haul flights. Virtually all aspects of physiology are rhythmic, from cells to systems, and circadian rhythms are coordinated by a central pacemaker or clock in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN adapts slowly to changes in time zone, and peripheral clocks or oscillators adapt at different rates, such that the organism is in a state of desynchrony from the external environment and internally. Light exposure is the main factor controlling the circadian system and needs to be considered together with any pharmacological interventions. This review covers the relatively new chronobiotic drugs, which can hasten adaptation of the circadian system, together with drugs directly affecting alertness and sleep propensity. No current treatment can instantly shift circadian phase to a new time zone; however, adaptation can be hastened. The melatoninergic drugs are promising but larger trials in real-life situations are needed. For short stopovers it is recommended to preserve sleep and alertness without necessarily modifying the circadian system. New research suggests that modification of clock function via genetic manipulation may one day have clinical applications.

The relationship between night eating symptoms and disordered eating attitudes via insomnia and chronotype differences

Humans' sleep timing and the psychological construct “diurnal preference” determines their “chronotype” (i.e., morning or evening type). Diurnal preferences can affect sleep-awake rhythms and eating behaviors. Our aim in this study was to examine the relationship between night eating symptoms and disordered eating attitudes by evaluating insomnia and chronotype differences in university students. The participants, 383 university students, filled out a package of psychological tools, including the Morningness-Eveningness Questionnaire, the Insomnia Severity Index, the Night Eating Questionnaire, and the Eating Attitude Test. One way analysis of variance was used to investigate the relationship of chronotypes with scale scores, and mediation regression analysis was used to investigate the indirect effects of night eating symptoms on disordered eating attitudes. Insomnia and night eating scores of the participants varied statistically according to chronotypes, and both insomnia and night eating scores were associated with the evening type. Findings show that night eating symptoms have a direct effect on the chronotype differences and insomnia and an indirect effect on disordered eating attitudes, by increasing insomnia scores. In conclusion, night eating syndrome may represent the misalignment of food intake and may shift the circadian rhythm to delayed sleep phase, acting as a peripheral oscillator in human.

Separation of circadian- and behavior-driven metabolite rhythms in humans provides a window on peripheral oscillators and metabolism

Misalignment between internal circadian rhythmicity and externally imposed behavioral schedules, such as occurs in shift workers, has been implicated in elevated risk of metabolic disorders. To determine underlying mechanisms, it is essential to assess whether and how peripheral clocks are disturbed during shift work and to what extent this is linked to the central suprachiasmatic nuclei (SCN) pacemaker and/or misaligned behavioral time cues. Investigating rhythms in circulating metabolites as biomarkers of peripheral clock disturbances may offer new insights. We evaluated the impact of misaligned sleep/wake and feeding/fasting cycles on circulating metabolites using a targeted metabolomics approach. Sequential plasma samples obtained during a 24-h constant routine that followed a 3-d simulated night-shift schedule, compared with a simulated day-shift schedule, were analyzed for 132 circulating metabolites. Nearly half of these metabolites showed a 24-h rhythmicity under constant routine following either or both simulated shift schedules. However, while traditional markers of the circadian clock in the SCN-melatonin, cortisol, and PER3 expression-maintained a stable phase alignment after both schedules, only a few metabolites did the same. Many showed reversed rhythms, lost their rhythms, or showed rhythmicity only under constant routine following the night-shift schedule. Here, 95% of the metabolites with a 24-h rhythmicity showed rhythms that were driven by behavioral time cues externally imposed during the preceding simulated shift schedule rather than being driven by the central SCN circadian clock. Characterization of these metabolite rhythms will provide insight into the underlying mechanisms linking shift work and metabolic disorders.

Normalization of disrupted clock gene expression in males with tetraplegia: a crossover randomized placebo-controlled trial of melatonin supplementation.

Crossover double blind, randomized placebo-controlled trial. Circadian oscillators are located both in the brain and in peripheral organs. Melatonin, the main brain-derived hormone governing circadian variations, is highly associated with daylight patterns. However, in subjects with tetraplegia the melatonin levels are blunted. Here we studied peripheral oscillators in peripheral blood mononuclear cells (PBMCs) in males with tetraplegia by examining how exogenous melatonin may influence the expression of clock gene mRNAs. Sunnaas Rehabilitation Hospital, Nesoddtangen, Norway. Six males with tetraplegia received 2 mg of melatonin or placebo 4 days before the study period. We also included six able-bodied men sleeping or kept awake during the night. Plasma samples were collected four times during a 24-h period. The mRNA expression levels of the clock genes PER1, PER2, BMAL1, and REV-ERBα were quantified in PBMCs using quantitative RT-PCR. The mRNA expression levels of PER-1 and -2 and REV-ERBα were increased at 04:00 h compared with the able-bodied controls (p < 0.05). Melatonin supplementation changed mRNA peak-time toward the time of supplementation. Several peripheral clock genes displayed distorted expression levels in tetraplegia. Supplementation with melatonin changed the mRNA expression levels of these genes toward those observed among able-bodied. Financial support was provided from the Throne Holst Foundation, Sunnaas Rehabilitation hospital and the University of Ferrara (FAR2016).

Circadian period of luciferase expression shortens with age in human mature adipocytes from obese patients

Daily rhythms in physiology and behavior change with age. An unresolved question is to what extent such age-related alterations in circadian organization are driven by the central clock in the suprachiasmatic nucleus (SCN), modifying timing signals to contributing peripheral tissue oscillators, and are mediated by underlying changes in the local cellular oscillators themselves. Using a bioluminescence reporter approach, we sought to determine whether circadian clock function in human adipocytes from subcutaneous (SAT) and visceral (VAT) adipose tissues changes with age. SAT and VAT biopsies were obtained from obese individuals during gastric bypass surgeries [ n = 16; body mass index: 44.8 ± 11.4 kg/m2; age: 44 ± 9 yr (range: 30-58)]. Cells were isolated and transduced with a lentiviral circadian reporter construct [brain and muscle aryl hydrocarbon receptor nuclear translocator-like:luciferase ( BMAL:LUC)], and bioluminescence was recorded over a period of 3 d. Human BMAL1:LUC adipocytes displayed a robust luminescence rhythm with comparable within-individual periods in mature and preadipocytes ( P > 0.05). With increasing age, the circadian period decreased in mature adipocytes ( P = 0.005) (β = 4 min/yr; P < 0.05). Our ex vivo approach indicated that ageing changes the organization of endogenous circadian oscillators in human adipocytes, independent of SCN signaling.-Kolbe, I., Carrasco-Benso, M. P., López-Mínguez, J., Luján, J., Scheer, F. A. J. L., Oster, H., Garaulet, M. Circadian period of luciferase expression shortens with age in human mature adipocytes from obese patients.

Neural Network Interactions Modulate CRY-Dependent Photoresponses in Drosophila.

Light is one of the chief environmental cues that reset circadian clocks. In Drosophila, CRYPTOCHROME (CRY) mediates acute photic resetting of circadian clocks by promoting the degradation of TIMELESS in a cell-autonomous manner. Thus, even circadian oscillators in peripheral organs can independently perceive light in Drosophila However, there is substantial evidence for nonautonomous mechanisms of circadian photoreception in the brain. We have previously shown that the morning (M) and evening (E) oscillators are critical light-sensing neurons that cooperate to shift the phase of circadian behavior in response to light input. We show here that light can efficiently phase delay or phase advance circadian locomotor behavior in male Drosophila even when either the M- or the E-oscillators are ablated, suggesting that behavioral phase shifts and their directionality are largely a consequence of the cell-autonomous nature of CRY-dependent photoreception. Our observation that the phase response curves of brain and peripheral oscillators are remarkably similar further supports this idea. Nevertheless, the neural network modulates circadian photoresponses. We show that the M-oscillator neurotransmitter pigment dispersing factor plays a critical role in the coordination between M- and E-oscillators after light exposure, and we uncover a potential role for a subset of dorsal neurons in the control of phase advances. Thus, neural modulation of autonomous light detection might play an important role in the plasticity of circadian behavior.SIGNIFICANCE STATEMENT Input pathways provide circadian rhythms with the flexibility needed to harmonize their phase with environmental cycles. Light is the chief environmental cue that synchronizes circadian clocks. In Drosophila, the photoreceptor CRYPTOCHROME resets circadian clocks cell-autonomously. However, recent studies indicate that, in the brain, interactions between clock neurons are critical to reset circadian locomotor behavior. We present evidence supporting the idea that the ability of flies to advance or delay their rhythmic behavior in response to light input essentially results from cell-autonomous photoreception. However, because of their networked organization, we find that circadian neurons have to cooperate to reset the phase of circadian behavior in response to photic cues. Our work thus helps to reconcile cell-autonomous and non-cell-autonomous models of circadian entrainment.

MTOR signaling regulates central and peripheral circadian clock function.

The circadian clock coordinates physiology and metabolism. mTOR (mammalian/mechanistic target of rapamycin) is a major intracellular sensor that integrates nutrient and energy status to regulate protein synthesis, metabolism, and cell growth. Previous studies have identified a key role for mTOR in regulating photic entrainment and synchrony of the central circadian clock in the suprachiasmatic nucleus (SCN). Given that mTOR activities exhibit robust circadian oscillations in a variety of tissues and cells including the SCN, here we continued to investigate the role of mTOR in orchestrating autonomous clock functions in central and peripheral circadian oscillators. Using a combination of genetic and pharmacological approaches we show that mTOR regulates intrinsic clock properties including period and amplitude. In peripheral clock models of hepatocytes and adipocytes, mTOR inhibition lengthens period and dampens amplitude, whereas mTOR activation shortens period and augments amplitude. Constitutive activation of mTOR in Tsc2–/–fibroblasts elevates levels of core clock proteins, including CRY1, BMAL1 and CLOCK. Serum stimulation induces CRY1 upregulation in fibroblasts in an mTOR-dependent but Bmal1- and Period-independent manner. Consistent with results from cellular clock models, mTOR perturbation also regulates period and amplitude in the ex vivo SCN and liver clocks. Further, mTOR heterozygous mice show lengthened circadian period of locomotor activity in both constant darkness and constant light. Together, these results support a significant role for mTOR in circadian timekeeping and in linking metabolic states to circadian clock functions.