Astroglial mechanisms in sleep homeostasis

星形胶质细胞睡眠稳态机制

基本信息

批准号：
10163932
负责人：
MARCOS G FRANK
金额：
$ 61.65万
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/10163932
关键词：
Address Anatomy Arousal Astrocytes Attention Biological Brain Cells 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 Glial Fibrillary Acidic Protein Glutamate Transporter Heterogeneity Homeostasis Image Impaired cognition In Situ Hybridization Ion Channel Lead Learning Ligands Measurement Measures Mechanics Mediating Memory Mental disorders Metabolic Metabolism Methods Microscopy Molecular Mood Disorders Morbidity - disease rate Morphology Mus Neuroglia Neurons Neurotransmitters Pathway interactions Phospholipase C Plant Roots 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 base brain cell circadian pacemaker designer receptors exclusively activated by designer drugs differential expression digital experimental study frontal lobe in vivo innovation insight miniaturize nervous system disorder neurotransmitter uptake new therapeutic target next generation sequencing non rapid eye movement 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.

概括在美国，诸如白天过度嗜睡和失眠之类的睡眠问题在美国很常见。他们在许多精神病和神经系统疾病中发现，并引起注意力，学习和记忆的缺陷。某些睡眠问题可能是由于昼夜节律中断引起的，但另一些可能反映了睡眠的变化稳态；一种神秘的调节机制，可增加睡眠驱动，睡眠量和睡眠强度是先前时间的函数。睡眠体内平衡的细胞机制不完全描述但传统上认为是神经元。但是，我们已经表明胶质星形胶质细胞是一部分这种机制。更具体地说，我们建议睡眠体内平衡来自星形胶质细胞和神经元。因此，我们假设对睡眠损失的正常补偿性反应涉及星形胶质细胞的细胞内和分子变化。此A1提交已被广泛修订根据初步审查。包括新的实验和初步数据（由红色字体表示）。我们将通过体内的三种创新方法检验这一总体假设。在AIM 1中，我们从遗传上结合编码的Ca2+指示器（GECI）星形胶质细胞成像，同时在体内的无动鼠。这使我们能够在快速的自然状态下测量星形胶质细胞CA2+动力学眼动（REM）睡眠，非（N）REM睡眠和使用2光子和表面的清醒显微镜。我们还可以直接测试通过诱导的睡眠体内稳态中细胞内Ca2+的必要性减少体内信号并测量睡眠表达和稳态的变化。在AIM 2中，我们使用可诱导的分子技术改变星形胶质细胞中已知存在的主要信号通路（即GQ，GI 和GS蛋白）并检查睡眠表达和稳态的结果变化。在AIM 3中，我们使用下一代测序技术（单细胞RNA测序（SCRNA-SEQ））以隔离额外（但是目前未知的）信号通路与星形胶质细胞介导的睡眠体内平衡有关。哺乳动物星形胶质细胞基于形态，细胞特异性标记（例如GFAP+），离子高度多样通道，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入睡是未知的。 Scrna-Seq提供了一种解决此问题的新方法。影响：我们对新型神经胶质睡眠机制的表征将为对病因的病因提供新的见解异常睡眠和唤醒。我们的实验还将提供有关非神经元功能的新见解脑细胞。反过来，这可能导致靶向神经胶质而不是神经元的新疗法的发展。