Zona incerta GABA neurons modulate energy homeostasis

未定带 GABA 神经元调节能量稳态

• 批准号: 9426268
• 负责人: ANTHONY N VAN DEN POL
• 金额: $ 41.1万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-15 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9426268
• 关键词: Address; Adult; Animals; Appetite Stimulants; Area; Axon; Axonal Transport; Behavioral; Body Weight; Brain; Brain region; CASP3 gene; Caspase; Cell Nucleus; Cells; Cessation of life; Country; Coupled; Data; Desire for food; Diabetes Mellitus; Dopamine; Eating; Efferent Neurons; Electron Microscopy; Electrophysiology (science); Emotional; Enterobacteria phage P1 Cre recombinase; FOS gene; Food; Food deprivation (experimental); Foundations; Genes; Glutamates; Health; Heart Diseases; Homeostasis; Hypothalamic structure; Injection of therapeutic agent; Investigation; Label; Laser Scanning Confocal Microscopy; Lasers; Lateral; Leptin; Light; LoxP-flanked allele; Malignant Neoplasms; Measures; Methods; Microinjections; Mus; Neuromodulator; Neurons; Neuropeptides; Obesity; Play; Population; Presynaptic Terminals; Pseudorabies; Rabies; Rabies virus; Regulation; Reporter; Reporter Genes; Reporting; Rewards; Role; Secondary to; Signal Transduction; Site; Slice; Stroke; Structure; Structure of paraventricular nucleus of thalamus; Substrate Interaction; Synapses; Testing; Thalamic structure; Tracer; Transgenic Mice; Variant; Viral Vector; Weight; Weight Gain; Whole-Cell Recordings; Work; base; behavior test; brain cell; cell type; designer receptors exclusively activated by designer drugs; dopaminergic neuron; excitatory neuron; experimental study; feeding; gamma-Aminobutyric Acid; ghrelin; immunocytochemistry; in vivo; inhibitory neuron; interest; mad itch virus; novel; novel strategies; optogenetics; patch clamp; postsynaptic; postsynaptic neurons; promoter; reduced food intake; response; trend; ventromedial hypothalamic nucleus; voltage clamp; zona incerta

Title: Zona incerta GABA neurons modulate energy homeostasis Abstract. Obesity, which often leads to secondary health complications including heart disease, diabetes, stroke, cancer, and early death, has become a major health concern in the US. Here we test the general hypothesis that neurons in the rostromedial zona incerta (ZI), and particularly inhibitory GABA neurons, play an unexpectedly profound orexigenic role in increasing food intake and body weight. Most of the work done on the neuronal regulation of energy homeostasis has previously focused on neurons in other brain regions. The first Aim examines the structural substrates for interaction between ZI GABA axons and their postsynaptic targets. Using confocal scanning laser microscopy and dual immunolabel electron microscopy coupled with cre recombinase-dependent AAV and rabies virus tracers, we test the hypothesis that ZI axons project to a number of sites, including the paraventricular thalamus (PVT) and hypothalamic ventromedial nucleus (VMH) where direct synaptic connections are made with excitatory neurons. We use multiple transgenic mouse lines expressing Cre recombinase under control of various neuron-selective promoters including mice that express Cre in GABA neurons driven by a vGAT promoter. These will be coupled with intracerebral microinjections of AAV viral vectors containing floxed GFP or tdTomato reporter genes to study ZI GABA neuron efferent and afferent axon projections. We will also employ injection of a Cre recombinase-dependent Brainbow-type pseudorabies virus into the ZI; after retrograde axonal transport this PRV expresses a red reporter in wild-type cells, but in Cre-expressing GABA cells, reporter expression changes to yellow or cyan, helping define the cell of interest. Aim 2 tests the hypothesis that ZI GABA cells respond to long distance signals of energy homeostasis including ghrelin and leptin, and also to axonally released neuropeptide modulators of food intake. Whole cell recording allows us to test different mechanisms of action on ZI GABA cells produced by neuromodulator signals from other neurons involved in energy homeostasis. C-fos expression will be examined after food deprivation to test the hypothesis that ZI GABA neurons are more active during reduced food availability. Optogenetics is used in brain slices to test the hypothesis that release of transmitter from ZI GABA axons will exert similar inhibitory effects on PVT and VMH neurons; neighboring ZI dopamine cells are also tested. In Aim 3, we examine the role that rostromedial ZI GABA neurons play in ongoing energy homeostasis by cell silencing (using Gi-DREADDs and caspase) to test the hypothesis that body weight and food intake is reduced. Optogenetic activation with ChR2 variant ChIEF will test the hypothesis that stimulation of ZI GABA axons in different terminal zones will each enhance food intake and body weight. We also test the hypothesis that ZI GABA neuron activation provides a positive emotional valence. Together, these experiments examine converging structural, electrophysiological, and behavioral analyses, focusing on the role of the GABA ZI neurons in energy homeostasis. With the growing levels of obesity in this country approaching 30% of the adult population, and the associated health complications, identifying and understanding the brain cells that control and sense energy homeostasis will help to identify novel approaches to reducing the trend toward obesity.

标题：Zona Incerta Gaba神经元调节能量稳态 抽象的。肥胖通常会导致继发性健康并发症，包括心脏病，糖尿病， 中风，癌症和早期死亡已成为美国的主要健康问题。在这里我们测试一般 假设神经元在zona zona incerta（Zi），尤其是抑制性GABA神经元中的假设，播放 出乎意料的深刻的甲状酸在增加食物摄入量和体重中的作用。大多数完成的工作 关于能量稳态的神经元调节，以前侧重于其他大脑区域的神经元。 第一个目的检查了Zi Gaba轴突及其相互作用的结构基板及其之间的相互作用 突触后靶标。使用共聚焦扫描激光显微镜和双免疫标记电子显微镜 再加上CRE重组酶依赖性AAV和狂犬病病毒示踪剂，我们检验了Zi的假设 轴突项目针对多个地点，包括旁侧丘脑（PVT）和下丘脑 腹侧核（VMH），其中直接突触连接是用兴奋性神经元进行的。我们使用 在各种神经元选择性的控制下表达CRE重组酶的多个转基因小鼠系 包括VGAT启动子驱动的GABA神经元中表达CRE的小鼠在内的启动子。这些将是 结合含有Floxed GFP或TDTOMATO报告的AAV病毒载体的脑内显微注射 研究Zi GABA神经元的传出和传入轴突投影的基因。我们还将使用CRE注射 重组酶依赖性的脑弓型假型病毒进入Zi；逆行轴突运输后 PRV在野生型细胞中表达红色记者，但是在表达CRE的GABA细胞中，记者表达 变为黄色或青色，有助于定义感兴趣的细胞。 AIM 2检验了Zi Gaba细胞的假设 响应包括生长素和瘦素在内的能量体内平衡的长距离信号，也响应轴突 释放食物摄入的神经肽调节剂。全细胞记录使我们能够测试不同的 来自其他参与的神经调节剂信号产生的对Zi Gaba细胞的作用机制 能量稳态。食物剥夺后将检查C-FOS表达，以检验Zi的假设 GABA神经元在降低食物的可用性过程中更为活跃。光遗传学用于大脑切片测试 从Zi Gaba轴突中释放发射机的假设将对PVT和PVT产生相似的抑制作用 VMH神经元；还测试了相邻的Zi多巴胺细胞。在AIM 3中，我们研究了 细胞沉默的延伸性Zi Gaba神经元在持续的能量稳态中发挥作用（使用Gi-Dreadds 和caspase），以测试降低体重和食物摄入的假设。使用 CHR2变体负责人将检验以下假设：不同终端区域中Zi GABA轴突的刺激将会 每种都可以增强食物摄入量和体重。我们还测试了Zi GABA神经元激活的假设 提供积极的情感价。这些实验一起检查了融合结构， 电生理和行为分析的重点是GABA ZI神经元在能量中的作用 稳态。这个国家的肥胖水平不断增长，接近成年人口的30％，并且 相关的健康并发症，识别和理解控制和感官的脑细胞 能量稳态将有助于确定减少肥胖趋势的新颖方法。