Neural control of NREM sleep in the medulla

Project Summary/Abstract In this proposal, we make use of optogenetic and chemogenetic tools, in vivo calcium imaging, RNA-seq, viral-based circuit tracing, and genetic targeting techniques to dissect the neural circuits that control sleep behavior in the mammalian brain. At a fundamental level, the work presented in this research proposal may provide valuable information for developing new treatments for various human sleep disorders, such as insomnia. Decades of studies have revealed neuromodulatory circuits as key regulators of sleep and wakefulness. However, most of these studies focus on the arousal system. Sleep-promoting neurons are theoretically required for the initiation and maintenance of sleep states based on the flip-flop model of wake-sleep switching, in which sleep-promoting neurons are mutually inhibitory with wake-promoting neurons. Previous studies have identified several brain structures that promote non-rapid eye movement (or NREM) sleep, but whether these structures are involved in the initiation or the maintenance remains largely unknown. In this research project, we will primarily focus on neural control of sleep initiation, i.e. the transition from wakefulness to sleep – under physiological conditions, always to NREM sleep. We aim to decipher the circuit mechanisms underlying the wake-sleep transition. In pilot studies, we have identified a novel population of glutamatergic neurons in the ventrolateral medulla (VLM) that project to the preoptic area (POA), a prominent sleep center. We present preliminary data showing that these VLM neurons are activated during wake-sleep transitions and optogenetic activation of these neurons induces long-lasting NREM sleep in awake mice. These results lead to our working hypothesis: VLM glutamatergic neurons induce the transition from wakefulness to NREM sleep, which subsequently activate their downstream targets to maintain NREM sleep. In Aim 1, we will activate and inactivate VLM glutamatergic neurons and examine the sufficiency and necessity of their activity in the wake-sleep transition. In Aim2, we will perform microendoscopic calcium imaging in VLM glutamatergic neurons and examine their activity during sleep. Then, we will use single-cell RNA-seq to identify genetic markers of sleep-active VLM glutamatergic neurons for further in vivo experiments. In Aim 3, we will use viral-based circuit tracing to characterize the inputs and outputs of VLM neurons and examine their function in sleep regulation. Together, the results obtained from this proposal will expand our understanding of the neural basis of sleep behavior.