Effects of Hydrogen Peroxide on Substantia Nigra GABAergic Neuron Excitability

描述（由申请人提供）：最近的工作已经确定过氧化氢是基底神经节核（包括纹状体和黑质丘脑部）中的信号分子。目前的建议，旨在研究功能的过氧化氢作为信号分子在黑质网状部（SNr），这还没有被检查过。黑质GABA能神经元的体外可视化全细胞电流钳记录将用于记录其在施加外源性过氧化氢时的活性，更重要的是，同时通过抑制过氧化物酶来增强内源性过氧化氢水平，这将使用荧光过氧化氢敏感指示剂二氯二氢荧光素（DCF）进行验证。过氧化氢的影响下的通道将通过结合过氧化氢水平的操纵与代谢活化的非选择性阳离子通道（即TRPM）和ATP敏感性钾通道（K-ATP）的药理学阻断剂来揭示，这两者都已被Rice小组证明是过氧化氢在其他基底神经节核中的重要靶点。TRPM和K-ATP通道的激活可能分别增加和降低SNr GABA能神经元的兴奋性。在其他实验中，将使用快速扫描循环伏安法来确定过氧化氢诱导的SNr GABA能神经元的兴奋是否有助于先前观察到的由过氧化氢引起的黑质中的体树突多巴胺释放的抑制。还将研究过氧化氢对NMDA诱导的爆发放电的影响，为SNr中的过氧化氢信号传导提供额外的功能性结果。SNr的GABA能输出神经元将整个基底神经节网络的输出传递到丘脑和上级丘中的靶核。除了介导基底神经节在正常运动控制中的生理作用之外，SNr输出神经元的活动改变似乎也是帕金森病和其他运动障碍的许多病理生理学特征的基础。阐明调节黑质GABA能输出神经元兴奋性的关键因素是了解这些神经元在正常和病理条件下的生理特性的重要前提，并最终导致发现运动障碍的新治疗靶点。具体而言，过氧化氢可能有助于增加在帕金森氏病的SNr神经元的兴奋性，其潜在的机制和后果将在这里进行研究。

DESCRIPTION (provided by applicant): Recent work has established hydrogen peroxide as a signaling molecule in basal ganglia nuclei including the striatum and the substantia nigra pars compacta. The current proposal seeks to examine the functions of hydrogen peroxide as a signaling molecule in the substantia nigra pars reticulata (SNr) which has not been examined previously. In vitro visualized whole cell current clamp recordings of nigral GABAergic neurons will be used to record their activity while applying exogenous hydrogen peroxide, and more importantly, while enhancing endogenous hydrogen peroxide levels by inhibiting peroxidase enzymes, which will be verified using the fluorescent hydrogen peroxide-sensitive indicator, dichlorodihydrofluorescein (DCF). The channels underlying hydrogen peroxide's effects will be uncovered by combining manipulations of hydrogen peroxide levels with pharmacological blockers of metabolically activated nonselective cation channels (namely TRPM) and ATP-sensitive potassium channels (K-ATP), both of which have been shown by the Rice group to be important targets for hydrogen peroxide in other basal ganglia nuclei. The activation of TRPM and K- ATP channels would be expected to increase and decrease the excitability of SNr GABAergic neurons respectively. In other experiments, fast-scan cyclic voltammetry will be used to determine whether hydrogen peroxide-induced excitation of SNr GABAergic neurons contributes to the previously observed suppression of somatodendritic dopamine release in the substantia nigra caused by hydrogen peroxide. The effects of hydrogen peroxide on NMDA-induced burst firing will also be studied, providing an additional functional consequence for hydrogen peroxide signaling in the SNr. The GABAergic output neurons of the SNr convey the output of the entire basal ganglia network to target nuclei in the thalamus and superior colliculus. In addition to mediating the physiological role of the basal ganglia in the control of normal movement, it is altered activity of SNr output neurons which also seems to underlie many of the pathophysiological features of Parkinson's disease and other movement disorders. Elucidating key factors that modulate the excitability of nigral GABAergic output neurons is an important prerequisite to understanding the physiological properties of these neurons under both normal and pathological conditions and ultimately lead to discovery of novel therapeutic targets for movement disorders. Specifically, hydrogen peroxide might contribute to increases in the excitability of SNr neurons in Parkinson's disease, the potential mechanisms and consequences of which will be studied here.