Role of BK Channel Interactome in Excessive Ethanol Drinking

描述(由申请人提供)：酒精滥用和依赖影响了大约8.5%的美国人口，并造成了巨大的健康和社会成本。大电导钙激活钾(BK)通道，以及超过20%的已知与BKα亚基相互作用的蛋白质，已被INIA-West联盟确定为潜在的乙醇偏好的遗传决定因素。在本方案中，我们假设酒精对相关脑区BK通道相互作用网络的干扰是导致过度饮酒的原因之一。我们将重点研究抑制酒精兴奋效应的神经回路(伏隔核、杏仁核和前额叶皮质)。我们进一步假设，乙醇诱导的BK通道相互作用组的重组介导了在酒精依赖动物中观察到的神经传递和突触可塑性的变化。为了验证这些假设，我们将量化和操纵BKα亚单位的关键相互作用伙伴的表达水平。第一个特定的目的是描述缺乏BK通道的两个神经元辅助亚基之一的基因敲除小鼠的表型。我们将使用酒精中毒、耐受和戒断的分析，以及自愿饮酒导致适度或过量酒精摄入的范例。第二个具体目标是绘制已知的BK相互作用伙伴在伏隔核、杏仁核和前额叶皮质的表达图，并评估过量乙醇暴露如何改变它们的蛋白质水平。一种创新的蛋白质分析将被用于同时定量大脑样本中19个BK通道亚基和相互作用伙伴。然后，我们将评估病毒介导的最有希望的基因的局部沉默如何影响乙醇自我给药。在第三个特定目标中，将与INIA-West的研究人员合作，使用类似的功能方法来探索这些基因对杏仁核的GABA能神经传递和伏隔核的突触可塑性的贡献。这些拟议的实验有望揭示BK通道相互作用组对乙醇自我给药的贡献，并潜在地确定治疗酒精中毒的新分子靶点。 与公共健康相关：这项研究项目调查了大脑的一个主要钾通道对过度饮酒的贡献。我们期待这项工作为调节酒精的行为和细胞效应的分子机制提供完整的见解。翻译含义包括为开发更有效的酒精中毒治疗方法确定新的靶点。

DESCRIPTION (provided by applicant): Alcohol abuse and dependence affect an estimated 8.5% of the U.S. population and are responsible for substantial health and societal costs. The large conductance calcium-activated potassium (BK) channel, together with more than 20% of the proteins known to interact with BK a subunit, have been identified by the INIA-West consortium as potential genetic determinants of ethanol preference. In the present proposal, we hypothesize that perturbation of BK channel interaction network by alcohol in relevant brain regions contributes to excessive drinking. We will focus our investigation on the neurocircuitry subtending the motivational effects of ethanol (nucleus accumbens, amygdala and prefrontal cortex). We further hypothesize that ethanol-induced reorganization of the BK channel interactome mediates changes in neurotransmission and synaptic plasticity observed in ethanol-dependent animals. To test these hypotheses, we will quantify and manipulate expression levels of key interaction partners of the BK a subunit. The first Specific Aim is to characterize the phenotype of knockout mice deficient for either of the two neuronal auxiliary subunits of the BK channel. We will use assays of ethanol intoxication, tolerance and withdrawal, as well as paradigms of voluntary drinking leading to moderate or excessive ethanol intake. The second Specific Aim is to map the expression of known BK interaction partners in the nucleus accumbens, amygdala and prefrontal cortex, and assess how excessive ethanol exposure alters their protein levels. An innovative protein assay will be exploited for the simultaneous quantification of 19 BK channel subunits and interaction partners in brain samples. We will then assess how virally-mediated local silencing of the most promising genes affects ethanol self-administration. In a third Specific Aim, a similar functional approach will be used to probe the contribution of these genes to GABAergic neurotransmission in the amygdala and synaptic plasticity in the nucleus accumbens, in collaboration with INIA-West investigators. The proposed experiments are expected to uncover the contribution of BK channel interactome to ethanol self-administration and potentially pinpoint novel molecular targets for the treatment of alcoholism. PUBLIC HEALTH RELEVANCE: This research project investigates the contribution of a major potassium channel of the brain to excessive alcohol drinking. We anticipate this work to provide integrated insights into the molecular mechanisms mediating the behavioral and cellular effects of alcohol. Translational implications include the identification of novel targets for the development of a more efficient treatment of alcoholism.