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Hypocretin (Hcrt), also known as orexin, neuropeptide signaling stabilizes sleep and wakefulness in all vertebrates. A lack of Hcrt causes the sleep disorder narcolepsy, and increased Hcrt signaling has been speculated to cause insomnia, but while the signaling pathways of Hcrt are relatively well-described, the intracellular mechanisms that regulate its expression remain unclear. Here, we tested the role of microRNAs (miRNAs) in regulating Hcrt expression. We found that miR-137, miR-637, and miR-654-5p target the human HCRT gene. miR-137 is evolutionarily conserved and also targets mouse Hcrt as does miR-665. Inhibition of miR-137 specifically in Hcrt neurons resulted in Hcrt upregulation, longer episodes of wakefulness, and significantly longer wake bouts in the first 4 h of the active phase. IL-13 stimulation upregulated endogenous miR-137, while Hcrt mRNA decreased both in vitro and in vivo. Furthermore, knockdown of miR-137 in zebrafish substantially increased wakefulness. Finally, we show that in humans, the MIR137 locus is genetically associated with sleep duration. In conclusion, these results show that an evolutionarily conserved miR-137:Hcrt interaction is involved in sleep–wake regulation.

In the mouse, Period-2 (Per2) expression in tissues peripheral to the suprachiasmatic nuclei (SCN) increases during sleep deprivation and at times of the day when animals are predominantly awake spontaneously, suggesting that the circadian sleep-wake distribution directly contributes to the daily rhythms in Per2. We found support for this hypothesis by recording sleep-wake state alongside PER2 bioluminescence in freely behaving mice, demonstrating that PER2 bioluminescence increases during spontaneous waking and decreases during sleep. The temporary reinstatement of PER2-bioluminescence rhythmicity in behaviorally arrhythmic SCN-lesioned mice submitted to daily recurring sleep deprivations substantiates our hypothesis. Mathematical modeling revealed that PER2 dynamics can be described by a damped harmonic oscillator driven by two forces: a sleep-wake-dependent force and an SCN-independent circadian force. Our work underscores the notion that in peripheral tissues the clock gene circuitry integrates sleep-wake information and could thereby contribute to behavioral adaptability to respond to homeostatic requirements., eLife digest Circadian rhythms are daily cycles in behavior and physiology which repeat approximately every 24 hours. The master regulator of these rhythms is located in a small part of the brain called the supra-chiasmatic nucleus. This brain structure regulates the timing of sleep and wakefulness and is also thought to control the daily rhythms of cells throughout the body on a molecular level. It does this by synchronizing the activity of a set of genes called clock genes. Under normal conditions, the levels of proteins coded for by clock genes change throughout the day following a rhythm that matches sleep-wake patterns. However, keeping animals and humans awake at their preferred sleeping times affects the protein levels of clock genes in many tissues of the body. This suggests that, in addition to the supra-chiasmatic nucleus, sleep-wake cycles may also influence clock-gene rhythms throughout the body. To test this theory, Hoekstra, Jan et al. measured the levels of PERIOD-2, a protein coded for by the clock gene Period-2, while tracking sleep-wake states in mice. They did this by imaging a bioluminescent version of the PERIOD-2 protein in the brain and the kidneys, at the same time as they recorded the brain activity, movement and muscle response of animals. Results showed that PERIOD-2 increased on waking and decreased when mice fell asleep. Additionally, in mice lacking a circadian rhythm in sleep-wake behavior – whose changes in PERIOD-2 levels with respect to time were greatly reduced – imposing a regular sleep-wake cycle restored normal PERIOD-2 rhythmicity. Next, Hoekstra, Jan et al. developed a mathematical model to understand how sleep-wake cycles together with circadian rhythms affect clock-gene activity in the brain and kidneys. Computer simulations suggested that sleep-wake cycles and circadian factors act as forces of comparable strength driving clock-gene dynamics. Both need to act in concert to keep clock-genes rhythmic. The model also predicted the large and immediate effects of sleep deprivation on PERIOD-2 levels, giving further credence to the idea that waking accelerated clock-gene rhythms while sleeping slowed them down. Modelling also suggested that having regular clock-gene rhythms protects against sleep disturbances. In summary, this work shows how sleep patterns contribute to the daily rhythms in clock genes in the brain and body. The findings support the idea that well-timed sleep-wake schedules could help people to adjust to new time zones. It might also be useful to inform other strategies to reduce the health impacts of shift work.

Wakefulness is accompanied by experience-dependent synaptic plasticity and an increase in activity-regulated gene transcription. Wake-induced genes are certainly markers of neuronal activity and may also directly regulate the duration of and need for sleep. We stimulated murine cortical cultures with the neuromodulatory signals that are known to control wakefulness in the brain and found that norepinephrine alone or a mixture of these neuromodulators induced activity-regulated gene transcription. Pharmacological inhibition of the various signaling pathways involved in the regulation of gene expression indicated that the extracellular signal-regulated kinase (ERK) pathway is the principal one mediating the effects of waking neuromodulators on gene expression. In mice, ERK phosphorylation in the cortex increased and decreased with wakefulness and sleep. Whole-body or cortical neuron-specific deletion of Erk1 or Erk2 significantly increased the duration of wakefulness in mice, and pharmacological inhibition of ERK phosphorylation decreased sleep duration and increased the duration of wakefulness bouts. Thus, this signaling pathway, which is highly conserved from Drosophila to mammals, is a key pathway that links waking experience-induced neuronal gene expression to sleep duration and quality.

Humans have an individual profile of the electroencephalographic power spectra at the 8 to 16 Hz frequency during non-rapid eye movement sleep that is stable over time and resistant to experimental perturbations. We tested the hypothesis that this electroencephalographic "fingerprint" is genetically determined, by recording 40 monozygotic and dizygotic twins during baseline and recovery sleep after prolonged wakefulness. We show a largely greater similarity within monozygotic than dizygotic pairs, resulting in a heritability estimate of 96%, not influenced by sleep need and intensity. If replicated, these results will establish the electroencephalographic profile during sleep as one of the most heritable traits of humans.

Significance When and how long we sleep is determined by the time of day and how long we have been awake, which are tracked molecularly by a circadian and a sleep–wake-driven process, respectively. We measured the long-term consequences of a short-term sleep deprivation on gene expression and regulation in the mouse brain, and used mathematical models to determine the relative contributions of the circadian and sleep–wake-driven processes. We find that many genes, including most of the genes that constitute the molecular circadian clock, are perturbed by sleep deprivation long after the mice ceased showing behavioral signs of sleep loss. Our results have implications for human health, given the high prevalence of insufficient and poor-quality sleep in our contemporary society., The timing and duration of sleep results from the interaction between a homeostatic sleep–wake-driven process and a periodic circadian process, and involves changes in gene regulation and expression. Unraveling the contributions of both processes and their interaction to transcriptional and epigenomic regulatory dynamics requires sampling over time under conditions of unperturbed and perturbed sleep. We profiled mRNA expression and chromatin accessibility in the cerebral cortex of mice over a 3-d period, including a 6-h sleep deprivation (SD) on day 2. We used mathematical modeling to integrate time series of mRNA expression data with sleep–wake history, which established that a large proportion of rhythmic genes are governed by the homeostatic process with varying degrees of interaction with the circadian process, sometimes working in opposition. Remarkably, SD caused long-term effects on gene-expression dynamics, outlasting phenotypic recovery, most strikingly illustrated by a damped oscillation of most core clock genes, including Arntl/Bmal1, suggesting that enforced wakefulness directly impacts the molecular clock machinery. Chromatin accessibility proved highly plastic and dynamically affected by SD. Dynamics in distal regions, rather than promoters, correlated with mRNA expression, implying that changes in expression result from constitutively accessible promoters under the influence of enhancers or repressors. Serum response factor (SRF) was predicted as a transcriptional regulator driving immediate response, suggesting that SRF activity mirrors the build-up and release of sleep pressure. Our results demonstrate that a single, short SD has long-term aftereffects at the genomic regulatory level and highlights the importance of the sleep–wake distribution to diurnal rhythmicity and circadian processes.