Central Circadian Clock Research Articles

The role of circadian clock in astrocytes: From cellular functions to ischemic stroke therapeutic targets.

Accumulating evidence suggests that astrocytes, the abundant cell type in the central nervous system (CNS), play a critical role in maintaining the immune response after cerebral infarction, regulating the blood-brain barrier (BBB), providing nutrients to the neurons, and reuptake of glutamate. The circadian clock is an endogenous timing system that controls and optimizes biological processes. The central circadian clock and the peripheral clock are consistent, controlled by various circadian components, and participate in the pathophysiological process of astrocytes. Existing evidence shows that circadian rhythm controls the regulation of inflammatory responses by astrocytes in ischemic stroke (IS), regulates the repair of the BBB, and plays an essential role in a series of pathological processes such as neurotoxicity and neuroprotection. In this review, we highlight the importance of astrocytes in IS and discuss the potential role of the circadian clock in influencing astrocyte pathophysiology. A comprehensive understanding of the ability of the circadian clock to regulate astrocytes after stroke will improve our ability to predict the targets and biological functions of the circadian clock and gain insight into the basis of its intervention mechanism.

Nutrition, Sleep, Circadian Rhythms, and Health Implications: “Come Together”

Over the last few years, novel and important aspects of nutrition that are often overlooked in nutritional epidemiology, experimental research, and recommendations for health maintenance and disease prevention concerning the circadian rhythmicity of feeding, as well as the bidirectional interaction of nutrition with central and peripheral endogenous circadian clocks, and with other rhythmic behaviors including the sleep-wake cycle have received increasing attention from the research community [...].

Reducing light exposure enhances the circadian rhythm of the biological clock through interactions with the gut microbiota

Light mainly synergistically regulates the central biological clock system. In farming, long-term light exposure may induce metabolic disorders and increase the load on the liver in laying hens. In contrast, intermittent photoperiods can reduce light exposure and increase rest time to improve the health of laying hens. The circadian rhythms of gut microbes are essential for the health of the host. However, the circadian rhythms of gut microbes and how those microbes interact with the host under intermittent photoperiods are not clear. We used laying hens as a model to evaluate the circadian rhythms of gut microbes and biological clock genes under different intermittent photoperiods. Intermittent photoperiod 1 (IP1, 16 [3 h -L/1 h -D]: 8 D) enhanced the circadian rhythms of cBmal1, cBmal2, cCry1, and cCry2 in the hypothalamus and increased the expression of cClock, cBmal1, and cCry2 in the liver and seven clock genes in the cecal wall. The intermittent photoperiod also significantly altered the composition and metabolic function of the cecal microbiota via the melatonin pathway. The concentrations of short-chain fatty acids (SCFAs) and the abundance of SCFA-producing genera such as Odoribacter significantly increased under the IP1 treatment and might have further fed back into and strengthened the peripheral and central rhythms by activating the SCFA receptor gene pathway in cecal wall. These findings clarify the mediation mechanisms for the circadian rhythms of the central circadian clock and highlight the role of intermittent photoperiod-induced regulation of the interaction between the host clock and the cecal microbial community.

PSUN111 Rationale and Design of the Dinner Time 2 Trial: A Randomized, Crossover Trial to Compare the Effects of Delayed Eating vs Delayed Sleeping on Overnight Metabolism in Healthy Volunteers

Abstract Background Obesity and its associated metabolic complications are leading causes of morbidity and mortality worldwide. Recent studies suggest that the timing of meals may be critically important for weight control and metabolic health. Consuming calories later in the day is associated with greater risks of obesity and metabolic syndrome. Interventional diet studies show more weight loss with early, rather than later, eating. Our team conducted a randomized, crossover study ("Dinner Time Study") that compared the metabolic effects of routine dinner (6pm) vs late dinner (10pm) with a fixed sleep period (11pm-7am) in young, healthy adults. We found that late dinner caused an 18% increase in post-prandial glucose and a 10% decrease in dietary fat oxidation. These metabolic consequences in the long term may lead to the development of obesity and type 2 diabetes. However, it remains unclear whether the adverse metabolic effects of late dinner are mediated by circadian misalignment (eating at the "wrong" time relative to the body's central circadian clock) or mediated by sleep (eating too close to bedtime, coinciding with the fall in metabolic rate induced by sleep). To address this question, we aim to examine the metabolic effects of early dinner, late dinner, and late dinner followed by delayed sleep, in healthy adults. Methods Dinner Time 2 Study is a randomized crossover trial with 3 treatment arms with a 3-4-week washout period: (1) early dinner + routine sleep; (2) late dinner + routine sleep; (3) late dinner + late sleep. Dinner times and bedtimes will be customized to each participant's central circadian rhythm (assessed by dim light melatonin onset, DLMO). The primary objectives of this study are to (1) examine the metabolic effects of early dinner (DLMO-3h) vs late dinner (DLMO+1h) with a fixed routine bedtime (DLMO+2h); (2) examine the metabolic effects of routine bedtime (DLMO+2h) vs delayed bedtime (DLMO+6h) with a fixed late dinner time (DLMO+1h). We will examine 24-h profiles of glucose, insulin, free fatty acids, triglycerides, and dietary fat oxidation using serial blood sampling and an ingested stable isotope ([2H31] palmitate) tracer. We aim to recruit 20 healthy adults, age 18-30 years old, with a BMI 18-29.9 kg/m2. Participants who perform night shift work or have any sleep disorders or metabolic diseases including diabetes are ineligible. Each participant will have 4 overnight admissions to our Clinical Research Unit (1 DLMO visit and 3 metabolic visits). A total of 3 participants have successfully completed the protocol since recruitment started in 2021. Conclusion Dinner Time 2 will greatly advance our understanding of the interactions between meal timing, sleep timing, and metabolism, which could inform the design and implementation of future studies that leverage chronobiology to combat diabetes and obesity. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

Males of Aedes aegypti show different clock gene expression profiles in the presence of conspecific females

BackgroundThe study of behavioral and physiological traits in mosquitoes has been mainly focused on females since males are not hematophagous and thus do not transfer the parasites that cause diseases in human populations. However, the performance of male mosquitoes is key for the expansion of populations and the perpetuation of mosquito species. Pre-copulatory communication between males and females is the initial and essential step for the success of copulation and studying the male facet of this interaction provides fertile ground for the improvement of vector control strategies. Like in most animals, reproduction, feeding, and oviposition are closely associated with locomotor activity in mosquitoes. Rhythmic cycles of locomotor activity have been previously described in Aedes aegypti, and in females, they are known to be altered by blood-feeding and arbovirus infection. In previous work, we found that males in the presence of females significantly change their locomotor activity profiles, with a shift in the phase of the activity peak. Here, we investigated whether this shift is associated with changes in the expression level of three central circadian clock genes.MethodsReal-time PCR reactions were performed for the gene period, cycle, and cryptochrome 2 in samples of heads, antennae, and abdominal tips of solitary males and males in the presence of females. Assays with antennae-ablated males were also performed, asking whether this is an essential organ mediating the communication and the variation in activity profiles.ResultsThe gene period showed a conserved expression pattern in all tissues and conditions, while the other two genes varied according to the male condition. A remarking pattern was observed in cry2, where the difference between the amplitude of expression at the beginning of photophase and the expression peak in the scotophase was greater when males were in the presence of females. Antennae ablation in males did not have a significant effect on the expression profiles, suggesting that female recognition may involve other senses besides hearing and olfaction.ConclusionOur results suggest that the expression of gene cryptochrome 2 varies in association with the interaction between males and females.Graphical abstract

Preliminary evidence that misalignment between sleep and circadian timing alters risk-taking preferences.

Decision-making has been shown to suffer when circadian preference is misaligned with time of assessment; however, little is known about how misalignment between sleep timing and the central circadian clock impacts decision-making. This study captured naturally occurring variation in circadian alignment (i.e., alignment of sleep-wake timing with the central circadian clock) to examine if greater misalignment predicts worse decision-making. Over the course of 2 weeks, 32 late adolescent drinkers (aged 18-22 years; 61% female; 69% White) continuously wore actigraphs and completed two overnight in-laboratory visits (Thursday and Sunday) in which both dim-light melatonin onset (DLMO) and behavioural decision-making (risk taking, framing, and strategic reasoning tasks) were assessed. Sleep-wake timing was assessed by actigraphic midsleep from the 2 nights prior to each in-laboratory visit. Alignment was operationalised as the phase angle (interval) between average DLMO and average midsleep. Multilevel modelling was used to predict performance on decision-making tasks from circadian alignment during each in-laboratory visit; non-linear associations were also examined. Shorter DLMO-midsleep phase angle predicted greater risk-taking under conditions of potential loss (B=-0.11, p=0.06), but less risk-taking under conditions of potential reward (B=0.14, p=0.03) in a curvilinear fashion. Misalignment did not predict outcomes in the framing and strategic reasoning tasks. Findings suggest that shorter alignment in timing of sleep with the central circadian clock (e.g., phase-delayed misalignment) may impact risky decision-making, further extending accumulating evidence that sleep/circadian factors are tied to risk-taking. Future studies will need to replicate findings and experimentally probe whether manipulating alignment influences decision-making.

Circadian clock and temporal meal pattern.

The central circadian clock in the brain controls the time-of-the-day variations in acute meal responses, with a low glycemic response but a high satiety/thermogenic response to meals consumed at waking compared to other time points. Consistently, studies show that consuming a significant proportion of calories, particularly carbohydrates, in breakfast is beneficial for the chronic management of obesity and its associated metabolic syndrome, compared to consuming identical meals at dinner. Conversely, breakfast skipping or/and late dinner can have unfavorable metabolic outcomes. It remains controversial how meal frequency affects metabolic health. In contrast, irregular meals, especially irregular breakfasts, show consistent adverse metabolic consequences. Time-restricted feeding (TRF), with all calories consumed within less than 12-h per day, can improve metabolism and extend lifespan. A major component of TRF in humans is caloric restriction, which contributes significantly to the beneficial effects of TRF in humans. By comparison, TRF effects in rodents can be independent of caloric restriction and show day/night phase specificity. TRF could alleviate metabolic abnormalities due to circadian disruption, but its effects appear independent of the circadian clock in rodents. Understanding neuroendocrine mechanisms underlying clock-mediated metabolic regulation will shed light on the metabolic effects of temporal meal patterns.

Research Progress on the Pathogenesis of Circadian Rhythm Sleep-Wake Disorders

Circadian rhythm fluctuates within 24 hours, affecting many physiological processes and aspects of daily life, including eating behavior, regulation of sleep wake cycle and metabolic homeostasis. Circadian rhythm sleep-wake disorders(CRSWDs) refer to the inconsistency between the human body's circadian behavioral activities and the needs of social life. Common classifications classify it as Delayed sleep phase type (DSP), Advanced sleep phase type (ASP), Free-running type and Irregular sleep-wake type. The potential pathogenesis of CRSWDs is closely related to human clock genes. Emens et al reported finding that N24H sleep-wake disorder is an important milestone in etiology. Light is the stronges inducement of circadian rhythm. CRSWDs may due to the long-term lack of light and other external timing factors, leading to circadian rhythm disorders and sleep homeostasis imbalance rather than being driven by a disorder of the central circadian clock. Regular exposure to strong light is often used to treat CRSWDs. This paper briefly introduces the historical background of understanding circadian rhythm, and then the common pathogenesis and mechanism of sleep wake disorder with different circadian rhythms and the corresponding treatment were discussed.

Prenatal androgen treatment impairs the suprachiasmatic nucleus arginine-vasopressin to kisspeptin neuron circuit in female mice.

Polycystic ovary syndrome (PCOS) is associated with elevated androgen and luteinizing hormone (LH) secretion and with oligo/anovulation. Evidence indicates that elevated androgens impair sex steroid hormone feedback regulation of pulsatile LH secretion. Hyperandrogenemia in PCOS may also disrupt the preovulatory LH surge. The mechanisms through which this might occur, however, are not fully understood. Kisspeptin (KISS1) neurons of the rostral periventricular area of the third ventricle (RP3V) convey hormonal cues to gonadotropin-releasing hormone (GnRH) neurons. In rodents, the preovulatory surge is triggered by these hormonal cues and coincident timing signals from the central circadian clock in the suprachiasmatic nucleus (SCN). Timing signals are relayed to GnRH neurons, in part, via projections from SCN arginine-vasopressin (AVP) neurons to RP3VKISS1 neurons. Because rodent SCN cells express androgen receptors (AR), we hypothesized that these circuits are impaired by elevated androgens in a mouse model of PCOS. In prenatally androgen-treated (PNA) female mice, SCN Ar expression was significantly increased compared to that found in prenatally vehicle-treated mice. A similar trend was seen in the number of Avp-positive SCN cells expressing Ar. In the RP3V, the number of kisspeptin neurons was preserved. Anterograde tract-tracing, however, revealed reduced SCNAVP neuron projections to the RP3V and a significantly lower proportion of RP3VKISS1 neurons with close appositions from SCNAVP fibers. Functional assessments showed, on the other hand, that RP3VKISS1 neuron responses to AVP were maintained in PNA mice. These findings indicate that PNA changes some of the neural circuits that regulate the preovulatory surge. These impairments might contribute to ovulatory dysfunction in PNA mice modeling PCOS.

Inhibitory responses to retinohypothalamic tract stimulation in the circadian clock of the diurnal rodent Rhabdomys pumilio.

In both diurnal and nocturnal mammals, the timing of activity is regulated by the central circadian clock of the suprachiasmatic nucleus (SCN). The SCN is synchronized to the external light cycle via the retinohypothalamic tract (RHT). To investigate potential differences in light processing between nocturnal mice and the diurnal rodent Rhabdomys pumilio, we mimicked retinal input by stimulation of the RHT ex vivo. Using Ca2+ imaging, we observed excitations as well as inhibitions of SCN neurons in response to electrical RHT stimulation. In mice, the vast majority of responses were excitatory (85%), whereas in Rhabdomys, the proportion of excitatory and inhibitory responses was similar (51% excitatory, 49% inhibitory). Glutamate blockers AP5 and CNQX blocked the excitatory responses to RHT stimulation but did not abolish the inhibitory responses in mice or Rhabdomys, indicating that the inhibitions were monosynaptically transmitted via the RHT. Simultaneous application of glutamate blockers with the GABAA antagonist gabazine blocked all inhibitory responses in mice, but not in Rhabdomys. Collectively, our results indicate that in Rhabdomys, considerably more inhibitory responses to light are present and that these responses are driven directly by the RHT. We propose that this increased proportion of inhibitory input could reflect a difference in the entrainment mechanism employed by diurnal rodents.

The Central Circadian Clock Protein TaCCA1 Regulates Seedling Growth and Spike Development in Wheat (Triticum aestivum L.).

The biological functions of the circadian clock on growth and development have been well elucidated in model plants, while its regulatory roles in crop species, especially the roles on yield-related traits, are poorly understood. In this study, we characterized the core clock gene CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1) homoeologs in wheat and studied their biological functions in seedling growth and spike development. TaCCA1 homoeologs exhibit typical diurnal expression patterns, which are positively regulated by rhythmic histone modifications including histone H3 lysine 4 trimethylation (H3K4me3), histone H3 lysine 9 acetylation (H3K9Ac), and histone H3 lysine 36 trimethylation (H3K36me3). TaCCA1s are preferentially located in the nucleus and tend to form both homo- and heterodimers. TaCCA1 overexpression (TaCCA1-OE) transgenic wheat plants show disrupted circadian rhythmicity coupling with reduced chlorophyll and starch content, as well as biomass at seedling stage, also decreased spike length, grain number per spike, and grain size at the ripening stage. Further studies using DNA affinity purification followed by deep sequencing [DNA affinity purification and sequencing (DAP-seq)] indicated that TaCCA1 preferentially binds to sequences similarly to “evening elements” (EE) motif in the wheat genome, particularly genes associated with photosynthesis, carbon utilization, and auxin homeostasis, and decreased transcriptional levels of these target genes are observed in TaCCA1-OE transgenic wheat plants. Collectively, our study provides novel insights into a circadian-mediated mechanism of gene regulation to coordinate photosynthetic and metabolic activities in wheat, which is important for optimal plant growth and crop yield formation.

The Circadian Clock of Polarized Microglia and Its Interaction with Mouse Brain Oscillators.

The activity of the immune system is controlled by circadian clocks present in different immune cells. The brain-resident subtype of immune cells, microglia, exhibits a wide range of functional phenotypes depending on the signaling molecules in their microenvironment. The exact role of microglia in the hypothalamic suprachiasmatic nuclei (SCN), the central circadian clock, has not been known. Therefore, the aim of this study was to determine (1) whether microenvironment-induced changes in microglial polarization affect circadian clocks in these cells and (2) whether the presence of microglia contributes to SCN clock function. Microglial and SCN clocks were monitored using PER2-driven bioluminescence rhythms at the tissue and single-cell levels. We found that polarization of resting microglia to a pro-inflammatory (M1) or anti-inflammatory (M2) state significantly altered the period and amplitude of their molecular circadian clock; importantly, the parameters changed plastically with the repolarization of microglia. This effect was reflected in specific modulations of the expression profiles of individual clock genes in the polarized microglia. Depletion of microglia significantly reduced the amplitude of the SCN clock, and co-cultivation of the SCN explants with M2-polarized microglia specifically improved the amplitude of the SCN clock. These results demonstrate that the presence of M2-polarized microglia has beneficial effects on SCN clock function. Our results provide new insight into the mutual interaction between immune and circadian systems in the brain.

The Influence of Dental Implants on the Circadian Clock and the Role of Melatonin in the Oral Cavity

The circadian rhythm regulates many physiological human health and disease processes. The central circadian clock is located in the suprachiasmatic nucleus in the hypothalamus and controls a whole range of circadian clock genes. During bone remodeling, circadian rhythm gene expression was observed in osteoblasts and osteoclasts. Moreover, circadian rhythm genes are an essential part of the circadian network in the osseointegration of titanium implants. Namely, titanium biomaterials significantly affect the expression of the circadian clock gene in the oral cavity and thus regulate the establishment of osseointegration. In addition, vitamin D has potential practical implications for patients with dental implants because sufficient vitamin D levels before surgery improve the osseointegration process. Also, the hormone melatonin, whose production depending on the circadian rhythm, affects bone homeostasis. Therefore, melatonin affects bone formation and could serve as a regenerative agent by increasing the process of osseointegration. This review highlights the impact of dental implants on circadian rhythm and osseointegration.

Universal Stress Protein regulates the circadian rhythm of central oscillator genes in Arabidopsis.

Environmental stresses restrict plant growth and development and decrease crop yield. The circadian clock is associated with the ability of a plant to adapt to daily environmental fluctuations and the production and consumption of energy. Here, we investigated the role of Arabidopsis Universal Stress Protein (USP; At3g53990) in the circadian regulation of nuclear clock genes. The Arabidopsis usp knockout mutant line exhibited critically diminished circadian amplitude of the central oscillator CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) but enhanced the amplitude of TIMING OF CAB EXPRESSION 1 (TOC1). However, the expression of USP under the control of its own promoter restored the circadian timing of both genes, suggesting that USP regulates the circadian rhythm of Arabidopsis central clock genes, CCA1 and TOC1.

192-OR: T-Type Calcium Channel Cav3.2 in GABAergic Neurons Regulates Rhythmic Glucose Metabolism

Systemic insulin sensitivity shows a diurnal rhythm with a peak upon waking. The GABAergic (γ-aminobutyric acid-producing) neurons in the suprachiasmatic nucleus (SCN) (SCNGABA neurons) were reported to control the diurnal rhythm of insulin-mediated suppression of hepatic glucose production in mice in earlier work1. SCNGABA neural firing activity displays a diurnal rhythm, with the intrinsic ionic mechanism underlying this temporal pattern not completely understood. Here we provided evidence supporting a role of the T type voltage-gated calcium ion channel subunit 3.2 (Cav3.2) in SCNGABA neurons in regulating rhythmic neural firing and glucose homeostasis. We found that Cav3.2 global knockout (Cav3.2KO) mice displayed disrupted diurnal rhythm of blood glucose levels. Selective deletion of Cav3.2 in the SCNGABA neurons abolished the oscillatory firing activity of SCNGABA neurons and led to glucose intolerance. Furthermore, our real-time calcium imaging data in freely moving mice showed that SCN GABA neuronal activity displayed a circadian rhythm pattern in WT control mice but not Cav3.2KO mice or mice with Cav3.2 selectively deleted in the SCNGABA neurons. These findings provide insights into how the calcium ion channel regulates the central circadian clock and glucose metabolism, with implications in the extended dawn phenomenon in type 2 diabetes. Keywords: Type 2 diabetes, calcium channel, Cav3.2, GABA neurons, circadian rhythm Reference: 1 Ding, G. et al. REV-ERB in GABAergic neurons controls diurnal hepatic insulin sensitivity. Nature 592, 763-767, doi:10.1038/s41586-021-03358-w (2021) . Disclosure P. Xu: None. W. Zhou: None. Z. Sun: None. Y. He: None. Funding NIH (P20 GM135002)

The central circadian regulator CCA1 functions in Glycine max during defense to a root pathogen, regulating the expression of genes acting in effector triggered immunity (ETI) and cell wall metabolism

Expression of the central circadian oscillator components CIRCADIAN CLOCK ASSOCIATED 1 (CCA1), TIMING OF CAB1 (TOC1), GIGANTEA (GI), and CONSTANS (CO) occurs in Glycine max root cells (syncytia) parasitized by the nematode Heterodera glycines while undergoing resistance, indicating a defense role. GmCCA1-1 relative transcript abundance (RTA) in roots experiencing overexpression (OE) or RNA interference (RNAi) is characterized by rhythmic oscillations, compared to a ribosomal protein gene (GmRPS21) control. A GmCCA1-1 RTA change, advancing by 12 h, exists in H. glycines-infected as compared to uninfected controls in wild-type, H. glycines-resistant, G. max[Peking/PI 548402]. The G. max[Peking/PI 548402] transgenic controls exhibit the RTA change by 4 h post infection (hpi), not consistently occurring in the H. glycines-susceptible G. max[Williams 82/PI 518671] until 56 hpi. GmCCA1-1 expression is observed to be reduced in H. glycines-infected GmCCA1-1-OE roots as compared to non-infected transgenic roots with no significant change observed among RNAi roots. The GmCCA1-1 expression in transgenic GmCCA1-1-OE roots remains higher than control and RNAi roots. Decreased GmCCA1-1 mRNA among infected roots shows the altered expression is targeted by H. glycines. Gene expression of proven defense genes including 9 different mitogen activated protein kinases (GmMAPKs), NON-RACE SPECIFIC DISEASE RESISTANCE 1 (GmNDR1-1), RPM1-INTERACTING PROTEIN 4 (GmRIN4-4), and the secreted xyloglucan endotransglycosylase/hydrolase 43 (GmXTH43) in GmCCA1-1-OE and GmCCA1-1-RNAi roots, compared to controls, reveal a significant role of GmCCA1-1 expression in roots undergoing defense to H. glycines parasitism. The observation that GmCCA1-1 regulates GmXTH43 expression links the central circadian oscillator to the functionality of the secretion system.

0003 On the Same Wavelength? Quantifying the Associations between Eating Timing and Rest-Activity Rhythms in Free-Living Adults

Abstract Introduction Misalignment between the central circadian clock and daily behaviors increases cardiometabolic morbidity and mortality risk, likely due to internal misalignment between central and peripheral circadian rhythms. Experimental studies suggest food intake may act as a time cue (‘zeitgeber’) for resetting circadian rhythms, representing a potential behavioral target to ameliorate circadian misalignment and associated health consequences. However, the extent to which eating timing relates to circadian rhythms in free-living adults is unclear. Therefore, we tested the associations between eating timing with 24-h rest-activity-rhythms (RAR), a free-living proxy for endogenous circadian rhythms, in non-shift-working adults. Methods Adults without chronic health conditions or sleep disorders completed 14 days of 24/7 wrist accelerometry to evaluate RAR variables of interdaily stability (IS; day-to-day stability in RAR), intradaily variability (IV; within-day fragmentation of RAR), relative amplitude (RA; difference between peak vs. trough activity), L5 onset time (5-h period with lowest activity), and M10 onset time (10-h period with highest activity). Concurrently, time-stamped image-assisted diet records were obtained to generate average eating timing variables, including daily eating onset (time of first caloric intake after awakening), offset (last caloric intake time), duration (time elapsed between eating onset and offset), and caloric midpoint (time at which 50% of daily kcals were consumed), and variables illustrating irregularity in eating timing (standard deviation of eating timing variables). Pearson’s correlations quantified the associations between RAR and eating timing variables. Results Participants (N=30) were 28.0±6.6 years, 57% female, with a BMI of 23.8±2.5 kg/m2. Higher IS was correlated with lower irregularity in both eating onset (r=-0.55, p<0.01) and duration (r=-0.51, p<0.01). Higher RA correlated with earlier eating onset (r=-0.47, p<0.01), longer eating duration (r=0.53, p<0.01), and lower eating onset irregularity (r=-0.37, p<0.05). Later L5 correlated with later eating onset (r=0.67, p<0.001), offset (r=0.58, p<0.001), caloric midpoint (r=0.56, p<0.01), and greater eating offset irregularity (r=0.53, p<0.01). Later M10 correlated with later eating offset (r=0.40, p<0.05). Conclusion Preliminary findings indicate that eating timing and RAR are moderately correlated in free-living adults. Earlier eating timing, increased eating regularity, and longer daily eating duration may represent behavioral targets for improving circadian rhythms and subsequent cardiometabolic outcomes. Support (If Any): Support provided by the American Heart Association (#831488) and a University of Delaware Research Fund-Strategic Initiative Award.

0214 Effects of Simulated Night-Shiftwork Induced Circadian Misalignment on the Human Plasma Metabolome

Abstract Introduction Circadian misalignment occurs when behaviors such as food intake and sleep happen at inappropriate endogenous circadian times, is common among shift-workers, and is a risk factor for cardiometabolic disease. Identifying mechanisms underlying adverse cardiometabolic risk linked to circadian misalignment could help develop new countermeasure strategies to mitigate health consequences of shiftwork. Thus, we analyzed 24-hour profiles of plasma metabolites during circadian alignment and misalignment in a night-shiftwork protocol. Methods 14 healthy adults (6M/8F), aged 26.4±1.2y (mean±SD), completed a 6 day in-laboratory simulated night-shiftwork study. The protocol comprised 2 baseline days (circadian alignment) followed by a transition day to shiftwork, and then 2 simulated night-shiftwork days (circadian misalignment). Participants consumed energy-balanced diets throughout the protocol, starting 3 days prior to laboratory admission. Plasma was collected every 4h during circadian alignment and misalignment conditions and was analyzed via untargeted metabolomics (liquid chromatography/mass-spectrometry). A model selection approach (best fit) with circadian and behavioral cosine models identified metabolites influenced by circadian (central circadian clock measured via dim-light melatonin onset) versus behavioral cycles (food intake/sleep). Results 5,171 metabolites were detected with average abundance of 424 (~8%) changing between conditions (false discovery rate (FDR)<0.05). 380 metabolites had significant (FDR<0.05) 24-h time-of-day patterns. Of these 380 metabolites, 248 had 24-h time-of-day patterns during circadian alignment, 283 had 24-h time-of-day patterns during misalignment, and 141 had 24-h time-of-day patterns during both conditions. Of the metabolites with 24-h time-of-day patterns in both conditions, 50 were circadian-influenced (e.g., arachidinoyl-serine) and 91 were behaviorally-influenced (e.g., propionylcarnitine). Conclusion Almost twice as many metabolites were influenced by the behavioral versus circadian cycle suggesting altered timing of behaviors like food intake and sleep have a significant impact on 24-h time-of-patterns in the human plasma metabolome. For example, propionylcarnitine is involved in energy metabolism and likely reflects 24h time-of-day patterns altered by food intake. Alternatively, arachidinoyl-serine was circadian influenced, is involved in vasodilation, and thus may contribute to circadian regulation of blood pressure. Further understanding circadian and behavioral regulation of the metabolome could have application in identifying mechanisms of physiological dysregulation and targets for intervention to mitigate health consequences of circadian misalignment. Support (If Any) NIH-DK092624, NIH-TR001082, NIH-DK048520, NIH-HL132150, NIH-HL145099, NIH-DK111161

0196 Retinal responsivity is associated with circadian phase and circadian alignment but not sleep timing

Abstract Introduction Light entrains the central circadian clock, with projections from the retina to the SCN through melanopsin-containing retinal ganglion cells (ipRGCs). Altered responsivity to light reflects ipRGC functioning and may be a physiological vulnerability for disrupted photoentrainment. Understanding how retinal responsivity relates to sleep and circadian timing may inform who might most benefit from sleep and circadian interventions. Methods 64 participants (85 observations) ages 20-66 years old were recruited during winter (n=35) and summer months (n=50), and included individuals with seasonal depression (n=33) and nonseasonal, never depressed controls (n=31). The post-illumination pupil response (PIPR) to red and blue light was used to measure the responsivity of ipRGCs (average 1:30pm; 10am-7pm). Circadian phase was assessed using Dim Light Melatonin Onset (DLMO), collected every 30-minutes on Friday evenings. Midsleep timing was measured using actigraphy (average number nights=4), and circadian alignment was calculated as the DLMO-midsleep phase angle. We performed a multilevel regression to determine the relationship between PIPR and markers of sleep and circadian timing, accounting for repeated seasonal assessments with a random intercept of participant. Covariates included age, gender, diagnostic group, and circadian time of PIPR assessments. We ran sensitivity analyses including photoperiod length on the day of PIPR assessment to account for potential light exposure on the PIPR. Results Greater retinal responsivity was associated with later DLMO (b=4.45; partially standardized b=0.28; SE=1.84; p=0.03), and shorter DLMO-midsleep phase angle (b= -7.33; partially standardized b= -0.32; SE=2.51; p=0.004), but not midsleep (b= -3.44; partially standardized b= -0.14; SE=2.35; p>0.05). Individuals with later DLMO had PIPR assessments at earlier circadian times (b= -0.12; SE=0.04; p=0.01). Older participants (b= -0.04; SE=0.02; p=0.04) and controls (b= -0.95; SE=0.44; p=0.04) had earlier sleep midpoints, but covariates were not associated with circadian markers. The association between circadian timing and PIPR became nonsignificant (b=4.15; partially standardized b=0.26; SE=1.88; p=0.06) when including photoperiod. Conclusion Retinal responsivity was associated with circadian but not behavioral sleep timing, suggesting ipRGC functioning may have downstream effects on circadian entrainment. Assessing circadian variation of retinal responsivity remains a crucial next step prior to testing whether retinal responsivity impacts response to circadian-focused interventions. Support (If Any) NIMH K.A.R.MH103303

Time Restricted Feeding to the Light Cycle Dissociates Canonical Circadian Clocks and Physiological Rhythms in Heart Rate.

Circadian rhythms are approximate 24-h biological cycles that optimize molecular and physiological functions to predictable daily environmental changes in order to maintain internal and organismal homeostasis. Environmental stimuli (light, feeding, activity) capable of altering the phase of molecular rhythms are important tools employed by circadian biologists to increase understanding of the synchronization of circadian rhythms to the environment and to each other within multicellular systems. The central circadian clock, located in the suprachiasmatic nucleus (SCN) of the hypothalamus is largely responsive to light and is thought to entrain the phase of peripheral clocks via neurohumoral signals. Mice are nocturnal and consume most of their food during the dark cycle. Early studies demonstrated that altered metabolic cues in the form of time restricted feeding, specifically, feeding mice during the light cycle, resulted in an uncoupling of molecular clocks in peripheral tissues with those from the SCN. These studies showed as much as a 12-h shift in gene expression in some peripheral tissues but not others. The shifts occurred without corresponding changes in the central clock in the brain. More recent studies have demonstrated that changes in cardiac physiology (heart rate, MAP) in response to time of food intake occur independent of the cardiac molecular clock. Understanding differences in the physiology/function and gene expression in other organs both independently and in relation to the heart in response to altered feeding will be important in dissecting the roles of the various clocks throughout the body, as well as, understanding their links to cardiovascular pathology.