Zona incerta GABA neurons modulate energy homeostasis

未定带 GABA 神经元调节能量稳态

• 批准号: 10242745
• 负责人: MURAT GUNEL
• 金额: $ 39.95万
• 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 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10242745
• 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; Injections; 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：未定带GABA神经元调节能量稳态 抽象。肥胖，这往往会导致继发性健康并发症，包括心脏病，糖尿病， 中风、癌症和早逝，已经成为美国的主要健康问题。在这里我们测试将军 这一假说认为，在吻内侧三叉神经（ZI）的神经元，特别是抑制性GABA神经元，发挥作用， 在增加食物摄入量和体重方面具有意想不到的深刻的食欲作用。所做的大部分工作 关于能量稳态的神经元调节，以前集中在其他脑区的神经元上。 第一个目的是检查ZI GABA轴突和它们的神经元之间相互作用的结构底物。 突触后靶点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重组酶依赖的AAV和狂犬病病毒示踪剂，我们测试了ZI 轴突投射到许多部位，包括室旁丘脑（PVT）和下丘脑（HHT）。 腹内侧核（VMH），其中与兴奋性神经元进行直接突触连接。我们使用 在各种神经元选择性控制下表达Cre重组酶的多个转基因小鼠系 启动子，包括在由vGAT启动子驱动的GABA神经元中表达Cre的小鼠。这些将是 结合脑内显微注射含有floxed GFP或tdTomato报告基因的AAV病毒载体 基因来研究ZI GABA神经元传出和传入轴突投射。我们还将采用注射Cre 重组酶依赖的Brainbow型伪狂犬病病毒进入ZI;逆行轴突运输后， PRV在野生型细胞中表达红色报告基因，但在表达Cre的GABA细胞中， 更改为黄色或青色，帮助定义感兴趣的单元格。目的2：验证ZI型GABA细胞 对长距离的能量稳态信号（包括ghrelin和leptin）作出反应， 释放的神经肽调节剂的食物摄入。全细胞记录使我们能够测试不同的 其他神经元参与的神经调质信号对ZI GABA细胞的作用机制 能量平衡将在食物剥夺后检测C-fos表达，以检验ZI GABA神经元在食物供应减少时更活跃。光遗传学被用于大脑切片， 假设从ZI GABA轴突释放递质将对PVT产生类似的抑制作用， VMH神经元;相邻的ZI多巴胺细胞也进行了测试。在目标3中，我们研究了 吻内侧ZI GABA神经元通过细胞沉默（使用Gi-DREADDs）在持续的能量稳态中发挥作用 和胱天蛋白酶）来检验体重和食物摄入减少的假设。光遗传激活， ChR 2变体ChIEF将检验以下假设，即刺激不同终末区中的ZI GABA轴突， 每一种都能增加食物摄入量和体重。我们还检验了ZI GABA神经元激活的假设 提供了一个积极的情绪效价。总之，这些实验研究收敛结构， 电生理和行为分析，重点是GABA ZI神经元在能量中的作用， 体内平衡随着这个国家肥胖率的不断上升，接近成年人口的30%， 相关的健康并发症，识别和理解控制和感知的脑细胞， 能量平衡将有助于确定减少肥胖趋势的新方法。