Kir 3 Channel Subunits in Drug Abuse with GABAB Agonists

描述(由申请人提供)：我们建议研究日益流行的设计药物伽马-羟丁酸(GHB)对腹侧被盖区(中脑边缘多巴胺系统)神经元的急性影响。GHB的作用是通过GABAB类型的G蛋白偶联受体(GPCRs)介导的，激活下游效应分子，如G蛋白门控的内向整流钾(Kir3或GIRK)通道。我们假设GIRK通道的特定组合在介导GABAg受体激动剂对中脑边缘多巴胺系统的急性奖赏效应中起主要作用。我们将(1)阐明G蛋白信号转导调节蛋白(RGS)在调节GABAB受体偶联效率中的作用，(2)评估GIRK通道亚单位(GIRK1、GIRK2和GIRK3)在调节GABAB受体在VTA的多巴胺和GABA神经元偶联效率中的作用，以及(3)确定VTA中异构体GIRK通道的单通道特性和组装规则。我们将使用双管齐下的方法-系统水平的方法，包括来自野生型、GIRK缺陷和转基因小鼠的急性脑片的膜片钳记录和双光子成像；以及细胞方法，涉及在异源细胞和VTA的培养神经元中表达的GIRK通道的膜片钳记录和先进的成像技术。总之，这些研究将揭示大脑中GIRK通道组装和偶联到Gabas受体的细胞和分子事件，并阐明GIRK通道在决定GHB和相关药物在大脑中的疗效中的作用。与公共卫生相关。毒品滥用是美国和世界范围内的一个问题，影响着数百万人。滥用毒品会带来强烈的奖赏感，这可能会导致重复服药、依赖和上瘾。上瘾的特征是对无法控制的渴望做出反应而复发。最初的奖励效应和渴望与中脑边缘多巴胺系统的激活有关。在过去的几年里，羟基丁酸的滥用急剧增加。通过揭示GIRK通道在调节GHB作用中的作用，我们可能揭示直接激活这些通道或改变GPCRs偶联效率的新药的靶点。这种方法可能会使GIRK通道成为治疗成瘾的强大药物靶点。

DESCRIPTION (provided by applicant): We propose to study the acute effects of the increasingly popular designer drug gamma-hydroxy butyric acid (GHB) on neurons in the ventral tegmental area (mesolimbic dopamine system). The actions of GHB are mediated through G protein-coupled receptors (GPCRs) of the GABAB type, activating downstream effectors, such as G protein-gated inwardly rectifying potassium (Kir3 or GIRK) channels. We hypothesize that specific combinations of GIRK channels play a primary role in mediating acute rewarding effects of GABAg receptor agonists on mesolimbic dopamine system. We will (1) elucidate the role of regulator of G-protein signaling (RGS) proteins in modulating the coupling efficiency of GABAB receptors, (2) asses the role of GIRK channel subunits (GIRK1, GIRK2, and GIRK3) on setting the coupling efficiency of GABAB receptors in dopamine and GABA neurons of the VTA, and (3) determine single-channel properties and rules of assembly of heteromeric GIRK channels in the VTA. We will use a two-pronged approach - a systems level approach involving patch-clamp recordings and two-photon imaging in acute brain slices from wild-type, GIRK- deficient, and transgenic mice; and a cellular approach, involving patch-clamp recordings and advanced imaging techniques from GIRK channels expressed in heterologous cells and in cultured neurons of the VTA. Together, these studies will reveal the cellular and molecular events underlying GIRK channel assembly and coupling to GABAs receptors in the brain, as well as elucidate the role of GIRK channels in determining the efficacy of GHB and related drugs in the brain. Public health relevance. Drug abuse is a US and world-wide problem affecting millions of people. Drugs of abuse impart strong sensations of reward, which may lead to repetitive drug administration, dependence and addiction. Addiction is characterized by relapse in response to uncontrollable cravings. The initial rewarding effects as well as the cravings are associated with the activation of the mesolimbic dopamine system. The abuse of GHB has increased dramatically over the last few years. By unraveling the role of GIRK channels in mediating GHB actions, we may reveal targets for new drugs that either directly activate these channels or alter the coupling efficiency to GPCRs. Such an approach may establish GIRK channels as formidable drug targets for treating addiction.