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Astroglial mechanisms in sleep homeostasis

星形胶质细胞睡眠稳态机制

基本信息

批准号：
10620162
负责人：
MARCOS G FRANK
金额：
$ 58.88万
依托单位：
WASHINGTON STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10620162
关键词：
Address Anatomy Arousal Astrocytes Attention Biological Brain Cells Circadian Dysregulation Circadian Rhythms Coupled Cyclic AMP Data Development Drowsiness Drug Targeting Electroencephalography Equilibrium Etiology Excessive Daytime Sleepiness Exocytosis Feedback G alpha q Protein GTP-Binding Proteins Gene Expression Gene Expression Profiling Glial Fibrillary Acidic Protein Glutamate Transporter Heterogeneity Homeostasis Image Impaired cognition In Situ Hybridization Ion Channel Learning Ligands Measurement Measures Mediating Memory Mental disorders Metabolic Metabolism Methods Microscopy Molecular Mood Disorders Morbidity - disease rate Morphology Mus Neuroglia Neurons Neurotransmitters Pathway interactions Phospholipase C Play Polysomnography Process Proteins REM Sleep Role STIM1 gene Second Messenger Systems Signal Pathway Signal Transduction Signaling Protein Sleep Sleep Deprivation Sleep Disorders Sleeplessness Synapses Techniques Technology Testing Time Transcript United States Wakefulness awake basal forebrain brain cell circadian pacemaker designer receptors exclusively activated by designer drugs differential expression digital experimental study frontal lobe in vivo innovation insight mechanical signal miniaturize nervous system disorder neurotransmitter uptake new therapeutic target next generation sequencing novel promoter response single-cell RNA sequencing sleep abnormalities transcriptome two photon microscopy two-photon

项目摘要

Summary Sleep problems such as excessive daytime sleepiness and insomnia are common in the United States. They are found in many psychiatric and neurological disorders and cause deficits in attention, learning and memory. Some sleep problems may be caused by disrupted circadian rhythms, but others may reflect changes in sleep homeostasis; an enigmatic regulatory mechanism that increases sleep drive, sleep amounts and sleep intensity as a function of prior time awake. The cellular mechanisms of sleep homeostasis are incompletely described but have traditionally thought to be neuronal. We, however, have shown that glial astrocytes are part of this mechanism. More specifically, we propose that sleep homeostasis arises from interactions between astrocytes and neurons. We therefore hypothesize that the normal compensatory response to sleep loss involves intracellular and molecular changes in astrocytes. This A1 submission has been extensively revised in accordance with initial review. New experiments and preliminary data are included (indicated by red font). We will test this overall hypothesis with three innovative approaches in vivo. In Aim 1, we combine genetically encoded Ca2+ indicator (GECI) astrocyte imaging with simultaneous polysomnographic recording in unanesthetized mice in vivo. This allows us to measure astrocyte Ca2+ dynamics in natural states of rapid- eye-movement (REM) sleep, non(N)REM sleep and wakefulness using both 2-photon and epiflorescent microscopy. We also more directly test the necessity of intracellular Ca2+ in sleep homeostasis by inducibly reducing this signal in vivo and measuring changes in sleep expression and homeostasis. In Aim 2, we use inducible molecular techniques to alter the major signaling pathways known to exist in astrocytes (i.e. Gq, Gi and Gs proteins) and examine the resulting changes in sleep expression and homeostasis. In Aim 3, we use next generation sequencing technology (single-cell RNA sequencing (scRNA-seq)) to isolate additional (but currently unknown) signaling pathways that are involved in astrocyte-mediated sleep homeostasis. Mammalian astrocytes are highly diverse based on morphology, cell-specific markers (e.g. GFAP+), ion channels, glutamate transporters and metabolic substrates. The relative contribution of these different astrocytes to sleep is unknown. scRNA-seq provides a new and powerful method to address this problem. Impact: Our characterization of a novel glial sleep mechanism will provide new insights into the etiology of abnormal sleep and arousal. Our experiments will also provide new insights into the function of non-neuronal brain cells. This in turn can lead to the development of new therapeutics that target glia, rather than neurons.

概括白天过度嗜睡和失眠等睡眠问题在美国很常见。他们存在于许多精神和神经系统疾病中，并导致注意力、学习和记忆缺陷。一些睡眠问题可能是由于昼夜节律紊乱引起的，但其他问题可能反映了睡眠的变化体内平衡；一种神秘的调节机制，可以增加睡眠动力、睡眠量和睡眠强度作为先前清醒时间的函数。睡眠稳态的细胞机制不完全已被描述，但传统上认为是神经元的。然而，我们已经证明神经胶质星形胶质细胞是的这个机制。更具体地说，我们认为睡眠稳态是由之间的相互作用产生的星形胶质细胞和神经元。因此，我们假设对睡眠不足的正常补偿反应涉及星形胶质细胞的细胞内和分子变化。此 A1 提交已在按照初步审查。包括新的实验和初步数据（以红色字体表示）。我们将用三种创新方法在体内测试这一总体假设。在目标 1 中，我们结合基因编码 Ca2+ 指示剂 (GECI) 星形胶质细胞成像，同时进行多导睡眠图记录未麻醉的小鼠体内。这使我们能够测量星形胶质细胞在快速自然状态下的 Ca2+ 动态。使用 2 光子和落射荧光进行眼动 (REM) 睡眠、非 (N)REM 睡眠和觉醒显微镜。我们还通过诱导更直接地测试细胞内Ca2+在睡眠稳态中的必要性减少体内这种信号并测量睡眠表达和体内平衡的变化。在目标 2 中，我们使用诱导分子技术可改变星形胶质细胞中已知存在的主要信号传导途径（即 Gq、Gi 和 Gs 蛋白）并检查由此产生的睡眠表达和体内平衡的变化。在目标 3 中，我们使用下一代测序技术（单细胞 RNA 测序 (scRNA-seq)）可分离额外的（但目前未知）参与星形胶质细胞介导的睡眠稳态的信号通路。哺乳动物星形胶质细胞在形态、细胞特异性标记物（例如 GFAP+）、离子通道、谷氨酸转运蛋白和代谢底物。这些不同的相对贡献星形胶质细胞的睡眠状态尚不清楚。 scRNA-seq 提供了一种新的、强大的方法来解决这个问题。影响：我们对一种新型神经胶质睡眠机制的表征将为了解神经胶质细胞睡眠的病因学提供新的见解。睡眠和觉醒异常。我们的实验还将为非神经元功能提供新的见解脑细胞。这反过来又可能导致针对神经胶质细胞而不是神经元的新疗法的开发。