Electrochemical Analysis of Dendritic Dopamine Release

描述（由申请人提供）：多巴胺（DA）是大脑运动和情绪通路中的关键递质。多巴胺能神经传递功能障碍是多种脑部疾病的基础，对社会产生重大影响，包括帕金森氏症和亨廷顿氏病，精神分裂症和成瘾。中脑DA细胞群是中枢神经系统DA的主要来源。黑质致密部（SNc）的细胞向侧投射支配背纹状体（黑纹状体DA系统），而邻近的腹侧被盖区（VTA）的细胞向伏隔核和其他边缘结构（中边缘DA系统）投射支配。DA神经元的一个潜在的独特特征是它们从中脑的树突以及远端轴突终端释放DA。树突和末端DA的释放对DA介导的行为至关重要，但对中脑中调节DA释放的因素知之甚少。使用碳纤维微电极和快速扫描循环伏安法实时检测脑切片中诱发的DA释放，我们发现树突和末端释放所需的Ca2+依赖性和Ca2+通道存在显著差异。此外，我们还发现了谷氨酸和GABA对SNc、VTA和纹状体中嗜离子受体的不同调节模式。值得注意的是，我们还发现了一种新的内源性黑质纹状体DA释放调节剂：活性氧（ROS） H202。在这篇续文中，我们将研究树突DA释放的Ca2+依赖性，重点是突触输入对DA细胞的贡献和细胞内储存Ca2+的释放（Aim I）。我们还将阐明ROS作为树突和末端DA溢出调节剂的作用（Aim 1 - 4）。Aim II将研究H202和其他活性氧在黑质纹状体和中边缘DA系统中的作用。目的研究活性氧的来源，包括末端和邻近DA释放位点的细胞。目的IV检测H202是否通过引起DA细胞超极化抑制树突状DA溢出；对膜性能的影响将通过全细胞记录来显示。

DESCRIPTION (provided by applicant): Dopamine (DA) is a key transmitter in motor and emotive pathways of the brain. Dysfunction of dopaminergic neurotransmission underlies a variety of brain disorders that have a critical impact on society, including Parkinson's and Huntington's diseases, schizophrenia, and addiction. DA cell groups in the midbrain provide the primary source of DA to the CNS. Cells of the substantia nigra pars compacta (SNc) project rostrally to innervate the dorsal striatum (nigrostriatal DA system) whereas those of the adjacent ventral tegmental area (VTA) project to the nucleus accumbens and other limbic structures (mesolimbic DA system). A potentially unique characteristic of DA neurons is that they release DA from their dendrites in midbrain, as well as from distant axon terminals. Dendritic, as well as terminal release of DA is critical for DA-mediated behaviors, yet little is known about factors that regulate DA release In midbrain. Using carbon-fiber microelectrodes and fast-scan cyclic voltammetry to detect evoked DA release in real-time in brain slices, we have found significant differences in the Ca2+-dependence and Ca2+-channels required for dendritic vs. terminal release. Further, we have found distinct patterns of regulation by glutamate and GABA acting at ionotropic receptors in SNc, VTA and striatum. Significantly, we also discovered a novel, endogenous regulator of the nigrostriatal DA release: the reactive oxygen species (ROS), H202. In this continuation, we will investigate the distinct Ca2+ dependence of dendritic DA release, with emphasis on contributions from synaptic input to DA cells and release of Ca2+ from intracellular stores (Aim I). We will also elucidate the role of ROS as modulators of dendritic and terminal DA overflow (Aim ll-IV). Aim II will examine the involvement of H202 and other ROS in nigrostriatal vs. mesolimbic DA systems. Aim Ill will investigate sources of ROS, including terminals and cells adjacent to DA release sites. Aim IV will test whether H202 inhibits dendritic DA overflow by causing hyperpolarization of DA cells; effects on membrane properties will be indicated by whole cell recording.