Zona incerta GABA neurons modulate energy homeostasis

标题：未确定带GABA神经元调节能量平衡 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、 在美国，中风、癌症和早逝已经成为一个主要的健康问题。在这里我们测试一下将军 旋转内侧不确定带神经元，特别是抑制性GABA神经元发挥作用的假说 在增加食物摄入量和体重方面具有意想不到的深远的促食作用。已完成的大部分工作 关于神经元对能量的调节，动态平衡以前主要集中在其他脑区的神经元上。 第一个目的是研究Zi GABA轴突和它们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重免疫标记电子显微镜 结合依赖cre重组酶的AAV和狂犬病病毒示踪剂，我们检验了ZI的假设 轴突投射到许多部位，包括室旁丘脑(PVT)和下丘脑 腹内侧核(VMH)，与兴奋性神经元建立直接突触联系。我们用 不同神经元选择性调控下表达Cre重组酶的多个转基因小鼠系 启动子包括在vGAT启动子驱动的GABA神经元中表达Cre的小鼠。这些将是 再加上脑内微量注射含有牙线状GFP或td番茄报告基因的AAV病毒载体 研究Zi GABA神经元传出和传入轴突投射的基因。我们还将使用CRE注入 重组酶依赖的脑弓型伪狂犬病病毒进入ZI；在逆行轴突运输后 PrV在野生型细胞中表达红色报告基因，而在表达Cre的GABA细胞中表达红色报告基因 更改为黄色或青色，有助于定义感兴趣的单元格。Aim 2验证了Zi GABA细胞 对包括Ghrelin和Leptin在内的能量动态平衡的长距离信号以及轴突做出反应 释放食物摄入量的神经肽调节剂。全细胞记录允许我们测试不同的 神经递质信号对Zi-GABA细胞的作用机制 能量平衡。将在食物剥夺后检测c-fos的表达，以检验Zi 在食物供应减少时，GABA神经元更加活跃。光遗传学被用在脑片上来测试 Zi GABA轴突释放递质对PVT和PVT有相似的抑制作用 VMH神经元；邻近的Zi多巴胺细胞也进行了测试。在目标3中，我们考察了 旋转内侧Zi GABA神经元通过细胞沉默(使用GI-DREADD)在持续的能量稳态中发挥作用 和caspase)来检验体重和食物摄入量减少的假设。光遗传激活 ChR2变异体负责人将检验刺激不同终末区Zi GABA轴突将 每一种都能增加食物的摄入量和体重。我们还检验了Zi GABA神经元激活的假设 提供一种积极的情感价值。总之，这些实验检验了融合的结构， 电生理和行为分析，重点是GABA Zi神经元在能量中的作用 动态平衡。随着这个国家日益增长的肥胖率接近成年人口的30%， 相关的健康并发症，识别和理解控制和感觉的脑细胞 能量平衡将有助于确定减少肥胖趋势的新方法。

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.