Studying the transcriptional changes in the brain following sleep deprivation has provided insight into the molecular mechanisms that differ between sleep and wake. Individual studies are limited in their ability to detect differentially expressed genes due to small sample size. Here we performed a meta-analysis of published brain expression data, totalling 173 microarrays across 245 mice. 498 genes were identified as significantly changing with sleep-deprivation at q < 0.01, 96 of which were previously identified by the original studies. Of the remaining 402 novel candidate sleep genes, 14 were associated with human sleep traits and 3 with sleep phenotypes in knockout mice. Candidate gene validation showed significant upregulation of Rasd1 (Dexras1) following sleep deprivation, and phenotyping of Rasd1 KO mice revealed changes in the amount and distribution of behavioural sleep duration and sleep bout structure. These results provide a greater understanding of the molecular correlates of sleep and provide a resource for the sleep research community.
Alzheimer's disease (AD) is traditionally conceptualized as a disorder of protein aggregation and neurodegeneration, yet growing evidence indicates that fundamental temporal organization of brain physiology is also disrupted. In the healthy brain, circadian clocks coordinate sleep-wake behavior, glial immunometabolism, astrocytic aquaporin-4 polarity, and glymphatic-lymphatic clearance, aligning immune readiness and proteostasis with daily activity-rest cycles. In AD, this temporal coordination progressively deteriorates, manifesting as sleep fragmentation, instability of rest-activity rhythms, vulnerability of central clock structures, and loss of circadian gating of glial and clearance pathways. These disruptions create phase-inappropriate immune and metabolic states, impair protein clearance, and alter the fate of extracellular vesicles, which may shift from mediators of waste export to facilitators of proteopathic spread. Importantly, circadian failure also constrains therapeutic delivery and biomarker interpretation by modulating blood-brain barrier transport, brain fluid dynamics, and brain-to-blood signal export. We propose that AD can be reframed as a systems-level timing disorder, in which loss of temporal coherence integrates molecular pathology, glial dysfunction, clearance failure, therapeutic inefficacy, and biomarker variability. This framework highlights chrono-pharmacology, chrono-neurotherapeutics, and circadian-informed biomarkers as essential components of precision strategies for AD prevention and treatment.
Alzheimer's disease (AD) is traditionally conceptualized as a disorder of protein aggregation and neurodegeneration, yet growing evidence indicates that fundamental temporal organization of brain physiology is also disrupted. In the healthy brain, circadian clocks coordinate sleep–wake behavior, glial immunometabolism, astrocytic aquaporin-4 polarity, and glymphatic–lymphatic clearance, aligning immune readiness and proteostasis with daily activity–rest cycles. In AD, this temporal coordination progressively deteriorates, manifesting as sleep fragmentation, instability of rest–activity rhythms, vulnerability of central clock structures, and loss of circadian gating of glial and clearance pathways. These disruptions create phase-inappropriate immune and metabolic states, impair protein clearance, and alter the fate of extracellular vesicles, which may shift from mediators of waste export to facilitators of proteopathic spread. Importantly, circadian failure also constrains therapeutic delivery and biomarker interpretation by modulating blood–brain barrier transport, brain fluid dynamics, and brain-to-blood signal export. We propose that AD can be reframed as a systems-level timing disorder, in which loss of temporal coherence integrates molecular pathology, glial dysfunction, clearance failure, therapeutic inefficacy, and biomarker variability. This framework highlights chrono-pharmacology, chrono-neurotherapeutics, and circadian-informed biomarkers as essential components of precision strategies for AD prevention and treatment.
A circadian clock enables an organism to occupy a particular temporal niche, and the evolutionary pressures associated with that temporal niche in turn shape the organization of the circadian network in which the clock is embedded. Although circadian organization has often been dichotomized into centralized and distributed systems, our modern understanding of circadian networks reveals a more complex organization that cannot be captured by this simple dichotomy. In this review, we examine how coupling between nodes of the circadian network (from cells, to tissues, to organs) gives rise to coherent circadian organization in mammals. We further highlight how comparative research on non-mammalian organisms reveals conserved and divergent strategies for circadian coupling that inform general principles of circadian network function across species.
The suprachiasmatic nucleus (SCN), a central clock in the hypothalamus of mammals, consists of heterogeneous populations of neurons. The SCN expresses various neurotransmitters/neuropeptides and hundreds of other genes as detected by cell census studies including transcriptomics. Nonetheless, the SCN can sustain a precise ∼24 h rhythm and acts as a central pacemaker to control daily rhythms of behavior and physiological processes throughout a lifespan. How does the SCN achieve this regularity in a network of ∼20,000 diverse cellular identities? Are there unique roles in individual SCN neurons or in subsets of SCN neurons? How are they classified, connected to sustain synchrony, and entrained to external light-dark cycles at the level of cell types? Only recently, the functional significance of individual oscillators in the SCN is beginning to be uncovered through the development and advancement of neurotechniques to target specific cell types for genetic manipulation and imaging. This review will summarize recent conditional knockout studies, focusing particularly on genetic drivers utilized in various experimental approaches. We will also discuss what questions lie ahead to disentangle the complexity of cellular components in the central pacemaker network.
The daily sleep-wake cycle is a conserved behaviour defined by locomotion quiescence and enhanced responsive threshold to sensory stimuli. Both the circadian clock and sleep-homeostasis determine the daily sleep profile. Environmental light is a major sensory input and also regulates circadian clock and the balance between sleep and wakefulness. In Drosophila, the cellular mechanism and neural circuitry underlying light-mediated circadian synchronization are well-established, yet the direct relationship between light/visual input and sleep remains unclear. To address this knowledge gap, we measured sleep behaviour in Drosophila with mutations in genes involved in phototransduction and downstream neural transmission. We observed consistent day sleep fragmentation in flies with mutations in multiple phototransduction components. We also found that mutation that led to hyperpolarised Drosophila photoreceptors resulted in shorter day sleep. We found a severe reduction in locomotor speed in several visual mutants during normal waking time preventing assessment of their sleep-linked immobility. Taken together, our rigorous quantification of sleep in phototransduction genetic mutants reveals the key role of visual input in promoting sleep.
Sleep stage flagging is critical for diagnosing conditions like insomnia, sleep apnea, and narcolepsy. Traditional methods rely on time-intensive manual scoring by experts, limiting scalability and accessibility, especially in resource-limited settings. Automating sleep stage classification through signal processing and machine learning could improve diagnostic efficiency and reduce healthcare burdens. While prior studies have utilized multiple signals such as electroencephalogram (EEG), electromyogram (EMG), and electrocardiogram (ECG), this study focuses solely on ECG to provide a simpler, more accessible solution. By simplifying signal input, the approach enhances feasibility in resource-constrained environments. Features based on heart rate variability (HRV) and Poincaré plot descriptors were extracted and used to train machine learning models for five-stage sleep classification. The approach was evaluated using two publicly available datasets, the Haaglanden Medisch Centrum Sleep Staging Database and the MIT-BIH Polysomnographic Database, which were chosen for their varied recording environments and subject diversity. Neural Networks, K-Nearest Neighbors (KNN), XGBoost, and Random Forest were employed to assess performance. The highest classification accuracy of 67 % was achieved with long-duration ECG recordings, outperforming models trained on shorter segments by 12 %. These findings emphasize the impact of signal duration on classification performance and suggest opportunities to refine sleep stage prediction. The study demonstrates the feasibility of ECG-only systems for portable, low-cost, and scalable sleep monitoring. The insights gained may facilitate the development of more accessible and efficient sleep disorder detection, particularly in low-resource settings.
Objective Sleep disorders including insomnia, obstructive sleep apnea (OSA), narcolepsy and REM sleep behavior disorder (RBD), significantly impair cognition, emotional wellbeing and physical health. This review synthesizes Neuroimaging evidence across major sleep disorders. It also documents knowledge gaps in the literature and provides clear next steps towards better diagnostic and therapeutic steps for sleep disorders. Methods PRISMA 2020 guidelines were followed for the systematic review of Neuroimaging studies on sleep disorders, with a comprehensive search of PubMed, Scopus and Google Scholar between 1997 and 2024. Results A total of 93 Neuroimaging studies published between 1997 and 2024 were systematically reviewed. Insomnia is characterized by reduced gray matter volume in the prefrontal cortex and hippocampus, increased beta activity in EEG, and decreased frontal lobe metabolism, supporting the hyperarousal model. OSA is associated with cortical thinning, hippocampal atrophy, and disrupted Default Mode Network (DMN) connectivity, correlating with cognitive deficits. Narcolepsy exhibits hypothalamic atrophy, reduced orexin signaling, and abnormal thalamocortical connectivity, explaining excessive daytime sleepiness and cataplexy. Parasomnias, particularly REM sleep behavior disorder (RBD), show neurodegenerative changes in the brainstem and basal ganglia, serving as early markers for synucleinopathies like Parkinson's disease. Conclusion Multimodal Neuroimaging and electrophysiological findings provide critical insights into the pathophysiology of sleep disorders, highlighting distinct neural patterns that enhance diagnostic precision and guide targeted interventions. Multimodal imaging approaches are essential for advancing precision medicine in sleep disorder management.
Light, particularly blue-wavelength light exerts a broad range of non-image forming (NIF) effects including the stimulation of cognition and alertness and the regulation of mood, sleep and circadian rhythms. However, its underlying brain mechanisms are not fully elucidated. Likewise, whether adolescents show a different NIF sensitivity to light compared to adults is not established. Here, we investigated whether cortical excitability, a basic aspect of brain function that depends on sleep-wake regulation, is affected by blue light and whether the effect is similar in young adults and adolescents. We used transcranial magnetic stimulation coupled to high-density electroencephalography (TMS-EEG) in healthy young adults (N = 13, 24.2 ± 3.4 y) and in adolescents (N = 15, 16.9 ± 1.1 y). Our results showed that, in young adults, blue light affected cortical excitability following an apparent inverted-U relationship, while adolescents' cortical excitability was not significantly different under blue light compared to orange light. In addition, although light did not affect performance on a visuomotor vigilance task completed during the TMS-EEG recordings, cortical excitability was positively correlated to task performance in both age groups. This study provides valuable insights into the complex interplay between light, cortical excitability, and behavior. Our findings highlight the role of age in NIF effects of light, suggesting that brain responses to light differ during developmental periods.
Aging is a risk factor for various disorders, and age-dependent changes in sleep parameters are involved in the pathogenesis of many diseases. Senescence-accelerated mice-prone 8 (SAMP8) have a short lifespan and show a disrupted circadian rhythm. However, little is known about how sleep parameters change with age in SAMP8. In this study, we evaluated changes in sleep parameters with aging in SAMP8 compared with those in senescence-accelerated mouse resistant 1 (SAMR1) as the control. Sleep quantity and fragmentation of sleep were evaluated at 4, 36, and 56 weeks of age using a PiezoSleep® system. The average duration of sleep episodes, reflected as the sleep fragmentation, decreased in an age-dependent manner in both SAMR1 and SAMP8, especially under light phase conditions. Interestingly, while the difference between SAMR1 and SAMP8 was not evident at 4 weeks of age, it was significantly reduced in SAMP8 at 36 and 56 weeks of age. These results suggest that the reduction of average sleep episode duration associated with the aging process was accelerated in SAMP8. To explore the mechanisms underlying the accelerated sleep fragmentation in SAMP8, we performed RNA sequencing on hypothalamic specimens obtained from SAMR1 and SAMP8 at 49 weeks of age, which revealed upregulation of type I interferon (IFN)-responsive genes in the SAMP8 hypothalamus. Furthermore, serum IFN-α levels at 49 weeks of age were higher in SAMP8 compared with those in SAMR1, suggesting that elevated IFN-α production in SAMP8 could be associated with the sleep fragmentation.
The circadian clock component PER2 coordinates daily oscillations in gene expression across multiple tissues, yet its role in assembling multi-protein regulatory complexes remains incompletely understood. Here, we report that PER2 nucleates a ternary complex with the tumor suppressor BRCA1 and the transcription factor POU2F1(OCT-1) to impose circadian control on target gene promoters. Using bacterial two-hybrid screening, we identified BRCA1 as a novel PER2-interacting protein. Biochemical mapping revealed that PER2 engages BRCA1 through multiple discrete binding interfaces: PER2 spanning residues 356-574 and 683-872 interact with both the N-terminal (1-400) and C-terminal BRCT (1670-1863) domains of BRCA1. Structural modeling predicted 361 residue contacts between PER2 and BRCA1, substantially more than the 74 contacts predicted for PER2:POU2F1(OCT-1), indicating differential affinities that enable ordered complex assembly. Sequential pull-down assays demonstrated that PER2, BRCA1, and POU domain form a stable ternary complex in vitro, with POU2F1(OCT-1) serving as the DNA-binding platform. Electrophoretic mobility shift assays revealed that pre-assembly of PER2 with POU domain inhibits DNA binding, while BRCA1 is essential for stabilizing PER2 recruitment to DNA-bound POU2F1(OCT-1). Using ESR1 as a functional readout, we demonstrated that this ternary complex directly regulates promoter activity. Circadian transcriptome analysis revealed that Esr1 exhibits robust clock-dependent oscillations that are abolished in Per1/2 double-knockout mice, while Pou2f1 and Brca1 maintain constitutive expression. These findings establish PER2 as a circadian scaffold that assembles multivalent protein complexes to temporally gate transcription, providing mechanistic insight into how circadian disruption can influence target gene expression.
Transcranial alternating current stimulation (tACS) is a promising tool for research on oscillatory brain activity, yet both behavioral and electrophysiological outcome measures show high variability across studies. One source for this variability might be chronotype and an incidental mismatch between chronotype and the time of the measurement.14 evening type and 14 morning type participants performed a sustained attention task — once at their chronotypically optimal and once at a non-optimal time of day. TACS was applied for 20 min at the individual alpha frequency over two electrodes located at Cz and Oz. EEG was recorded for 10 min prior to and after stimulation. Sleep timing and quality were assessed with a sleep questionnaire. While planned analyses failed to find effects of stimulation and session timing on alpha power, exploratory analyses revealed that below average sleep quality in evening types in the morning was associated with no changes or unexpected decreases in alpha power after stimulation. Effects of sleep quality were present in the morning for evening types, but neither in the evening session nor in morning types. It is suggested that this effect of sleep quality reflects increased sleepiness, which could impede expected aftereffects of tACS. It is likely that effects of sleepiness might be especially relevant when people are stimulated at a chronotypically non-optimal time. Due to the exploratory nature of these sleep effects and their presence in only a small subgroup leading to low power and confidence, future systematic sham-controlled studies are needed to clarify the relationship between sleep, time of day and chronotype in α-tACS proposed here.
Photoperiod is the primary environmental cue that regulates changes in behavior across seasons. Previously, we have shown that photoperiod has sex-specific effects on synaptic dopamine dynamics in the nucleus accumbens (NAc). Further, evidence suggests that the dopamine transporter (DAT) is a potential locus of action for the sex-specific effects of photoperiod on NAc dopamine. The NAc is a critical node within the reward circuit that brings motivation to action, and changes to NAc dopamine dynamics at the synapse can result in robust changes in behaviors. Cocaine is a psychostimulant that targets monoamine transporters, including DAT, and generates robust behavioral effects. Thus, using cocaine-mediated behavior, we can determine whether photoperiod impacts DAT function and dopamine physiology. Here, using male and female mice we examined the effect of seasonally relevant photoperiods on DAT function in the NAc and dopamine-dependent behavior. We found that females raised in Short, winter-like photoperiod have blunted cocaine-induced hyperlocomotion. Conversely, females raised in Long, summer-like photoperiod exhibit greater DA release and cocaine-mediated DAT inhibition while we observe decreased sensitivity to cocaine-associated learning. The combined work presented here provides evidence that photoperiod has differential, female-specific effects on NAc DAT function and DAT-mediated behaviors.
The suprachiasmatic nucleus (SCN) of the hypothalamus is a principal light-responsive circadian clock that adjusts circadian rhythms in mammalian physiology and behavior to changes in external light signals. Although mechanisms underlying how light acutely resets the timing of circadian rhythms have been characterized, it remains elusive how light signals induce lasting changes in circadian period, known as period after-effects. Here we have found that the period after-effects on circadian behavior of changing photoperiods are blocked by application of the DNA methyltransferase inhibitor RG108 near the SCN. At the level of single light pulses acting as clock-resetting stimulations, RG108 significantly attenuates period after-effects following acute phase shifts in behavioral rhythms in vivo, and blocks period after-effects on clock gene rhythms following phase resetting by the vasoactive intestinal peptide in the isolated ex vivo SCN. In addition, the DNA methyltransferase inhibitor SGI-1027 blocked period after-effects of optogenetic neuronal stimulation on ex vivo SCN rhythms. Acute clock resetting shifts themselves, however, do not appear to require DNA methylation at the SCN and behavioral levels, in contrast to subsequent period plasticity. Our results demonstrate that DNA methylation inhibitors block light-induced period after-effects in response to photoperiods and single light pulses. Together with previous studies showing that DNA methylation in the SCN is essential for period after-effects of non-24hr light cycles (T-cycles), this suggests that DNA methylation in the SCN may be a widespread mechanism of light-induced circadian period plasticity.
The regulation of sleep, while primarily attributed to the interplay between circadian and homeostatic processes, is significantly influenced by a multitude of additional factors that profoundly impact sleep quantity and quality. These factors encompass both external environmental stimuli, such as ambient temperature and somatosensory inputs, and internal physiological changes. The intricate relationship between metabolism and sleep has been a subject of extensive research, with particular attention given to the role of metabolic signals in sleep regulation. Among these, the brown adipose tissue (BAT) has emerged as a key player, studied from various perspectives including its physiological responses to sleep deprivation, its effects on sleep when activated, the consequences of impaired BAT thermogenesis on sleep patterns, and its metabolic activity across different sleep states. The cumulative evidence from these investigations suggests that BAT plays a crucial role in maintaining an optimal metabolic environment conducive to sleep, a function that becomes particularly significant in contexts of prior sleep loss, inflammatory conditions, and fluctuations in ambient temperature.
Background:Sleep hygiene is integral to health, and sleep regularity may be associated with mental health outcomes in addition to duration. Although sleep and depression relationships are well-studied, the relative impact of different sleep factors remains unclear. As patient-specific factors and health behaviors influence sleep and mental health, we investigated associations between sleep and depression severity considering such factors in a United States sample of adults. Methods:Two cycles (2011-2012, 2013-2014) from the National Health and Nutritional Examination Survey were studied. Objective sleep duration (day and night), and the sleep regularity index (SRI) were calculated from physical activity monitors worn for seven days. Complex survey procedures with four-year weights were used, and backward selection was used to test relevant variables in the fully adjusted regression model. Results:Among participants (n = 7297), we found associations between sleep-associated variables and SRI, with increased daytime sleep being the strongest correlate of decreased SRI. In the fully adjusted model, lower SRI scores and reduced subjective night sleep remained significantly associated with depression. Sex was an additional independent predictor, with females exhibiting higher depression scores, and a significant sex × SRI interaction revealed that the inverse relationship between SRI and depressive symptoms was stronger in females than in males. Health behaviors, including active tobacco and cannabis use, were also associated with increased depression severity in the adjusted model. Conclusions:Daytime sleep may serve as an SRI proxy, although additional cohorts should confirm relationships. Higher depression severity was associated with different sleep components, emphasizing the importance of sleep hygiene in mental health. Behaviors like current smoking and cannabis use were also associated with increased depression. Research exploring the temporality and interactions between these factors may assist in non-pharmacologic depression treatment.
Rett syndrome (RTT) is a severe, progressive neurodevelopmental disorder caused by mutations in the X-linked gene encoding methyl-CpG-binding protein 2 (MECP2). Sleep problems are frequently reported in Rett Syndrome, but the exact nature remains relatively unexplored. Currently there is limited understanding of MECP2's role in sleep architecture and regulation. In this study, we employed longitudinal electroencephalographic (EEG) and electromyographic (EMG) recordings to investigate sleep architecture during baseline conditions as well as the homeostatic response to sleep deprivation (SD) in Mecp2-/y male mice. At baseline, Mecp2-/y mice have more non-rapid-eye-movement (NREM) sleep and less rapid-eye-movement (REM) sleep than their wildtype littermates during the light period. However, Mecp2-/y mice display altered sleep timing during the dark period, spending more time in both NREM and REM during the first half and less time during the second half. Mecp2-/y mice also have lower EEG spectral power during wake and NREM at higher frequencies and higher power at lower frequencies during REM in compared to wildtype mice. In response to SD, Mecp2-/y mice can accumulate and discharge sleep pressure normally and show a sleep rebound. However, baseline differences in sleep architecture are heightened after SD. Overall, our findings show that RTT mice exhibit distinct sleep patterns compared to wildtype mice, with time-of-day-dependent variations in NREM and REM sleep, as well as altered EEG spectral properties, that become more pronounced following SD. Future research should explore the molecular mechanisms through which MECP2 regulates sleep architecture to develop targeted therapeutics for sleep disturbances in RTT patients.
Cardiovascular diseases are paramount cause of morbidity in aging population and aging disrupts normal circadian rhythm cycle. Circadian rhythms, regulated by the suprachiasmatic nucleus in the brain, profoundly influence cardiovascular health through intricate neurobiological mechanisms. These rhythms regulate gene expression in cardiomyocytes, modulate autonomic nervous system (ANS) activity, and synchronize cardiovascular functions with environmental cues, ultimately impacting heart rate, blood pressure, and susceptibility to cardiac events. The intricate relationship between circadian rhythms and cardiovascular health emphasizes the critical role of brain-heart communication in physiological processes.This review explores the neurobiology of circadian clock in cardiovascular disease, exploring how peripheral clocks in cardiovascular tissues influence organ physiology and how their disruption contributes to pathogenesis. The examination of neurobiological pathways linking circadian clock to cardiovascular disease, including ANS function, neuroendocrine signaling, and inflammatory responses, highlights the interplay between brain and heart. By probing environmental and lifestyle factors that modulate the circadian clock, as well as sex-specific variations in circadian rhythms, the review provides a comprehensive understanding of how these factors impact cardiovascular health. The discussion of emerging concepts, such as exosome-mediated intracellular communication in circadian physiology, offers new insights into the molecular mechanisms underlying brain-heart interactions. Furthermore, the exploration of diagnostic potential and therapeutic strategies, particularly chronotherapy, emphasizes the importance of targeting the circadian clock for disease prevention and treatment in cardiovascular medicine. This comprehensive assessment not only advances our understanding about circadian clock's role in cardiovascular health but also paves the way for innovative approaches in theranostic, ultimately improving patient outcomes.
In this article for Dr Krueger's Festschrift, I trace how his early career influenced many aspects in the fields of sleep, neuroimmunology, and the microbiome. Mostly, however, I trace how his career and interests intertwined with those of Abba J. Kastin and mine and how he exerted a profound influence on the direction of our studies. Dr. Krueger, while developing his career as a sleep researcher, encountered resistance to his work that required two major paradigm shifts: 1) that bacterial products could affect sleep and 2) that small peptides can cross the blood-brain barrier (BBB) in sufficient amounts to affect brain functioning. Dr Kastin had also shown that small peptides administered peripherally could affect brain function and postulated that this was because they could cross the BBB. Our efforts to determine whether peptides could or could not cross the BBB were bolstered by Dr Krueger's exemplary struggles.
