Ethanol Dependence Induced Changes in Dopamine Signaling in Basolateral Amygdala

 描述(由申请人提供)：目前申请的总体目标是了解有助于赋予病理行为的神经生物学机制，如酒精身体依赖后增强的负面影响。最近的研究表明，杏仁内侧/基底外侧(BLA)GABA能神经元严密控制负面情绪的表达，包括慢性酒精暴露后戒断时表达的情绪。来自腹侧被盖/黑质致密部的多巴胺能输入调节这一GABA能系统，并抑制驱动焦虑样行为表达的BLA主神经元。基于强有力的初步证据，我们提出的实验将检验这一中心假设，即酒精依赖通过失调这些GABA能神经元的多巴胺能调制来抑制BLA的输出。我们将测试我们的中心假设，并通过利用一个成熟的慢性酒精暴露的大鼠模型，并通过整合光遗传学、突触前和突触后的多巴胺神经生理学以及行为实验方法来实现我们的总体目标。BLA被广泛认为是神经回路的重要调节成分，控制着慢性酒精暴露戒断时的焦虑样行为，以及药物幼稚和暴露动物的奖赏寻求。具体目标1将通过检查酒精依赖戒断过程中突触前多巴胺的功能来检验我们的中心假设。我们将通过结合体外快速扫描循环伏安法和光遗传控制DA释放和慢性乙醇暴露来直接测量DA的释放和再摄取。我们假设，根据我们以前的文献，慢性酒精暴露将以不同的方式调节基础或强直的DA水平和DA释放的时相，最终增强DA信号。具体目标2将使用体外全细胞膜片钳电生理学和创新的光遗传学方法来检测突触后DA受体信号，以研究突触后DA受体功能是如何在BLA改变的。我们的工作假设是，酒精的身体依赖将增加突触后D1和D2样DA受体信号，从而使DA介导的对GABA能功能的抑制被上调。具体目标3将通过整合对DA释放的光遗传控制和体内对BLA依赖行为的测量，将依赖DA信号的详细细胞效应置于整个动物的背景下。我们的工作假设是，多巴胺神经传递和信号传递中与依赖相关的变化最终控制着依赖戒断的焦虑样行为。这项拟议的工作采用了一种独特的、高度集成的实验方法，为控制慢性酒精暴露的负面强化效应的神经生物学机制提供了无与伦比的见解。最终，这些研究将为人类酗酒者控制滥用和复发的潜在细胞机制提供洞察力。

 DESCRIPTION (provided by applicant): The overall goal of the current application is to understand the neurobiological mechanisms that help confer pathological behaviors like enhanced negative affect following ethanol physical dependence. Recent studies suggest that intrinsic lateral/basolateral amygdala (BLA) GABAergic neurons tightly control the expression of negative emotions including those expressed during withdrawal following chronic ethanol exposure. Dopaminergic inputs from the ventral tegmentum/substantia nigra pars compacta have been shown to regulate this GABAergic system and disinhibit BLA principal neurons which drive the expression of anxiety-like behaviors. Based on strong preliminary evidence, our proposed experiments will test the central hypothesis that ethanol dependence dis-inhibits BLA output by dysregulating dopaminergic modulation of these GABAergic neurons. We will test our central hypothesis and accomplish our overall goal by utilizing a well- established rat model of chronic ethanol exposure and by integrating optogenetic, pre- and post-synaptic dopamine neurophysiology, and behavioral experimental approaches. The BLA has been extensively implicated as an important regulatory component of the neural circuitry controlling both anxiety-like behavior during withdrawal from chronic ethanol exposure as well as reward-seeking in drug-naïve and -exposed animals. Specific Aim 1 will test our central hypothesis hypothesis by examining presynaptic dopamine function during withdrawal from ethanol dependence. We will directly measure DA release and reuptake in vitro by integrating in vitro fast-scan cyclic voltammetry with optogenetic control of DA release and chronic ethanol exposure. We hypothesize that, based on our previous publications, chronic ethanol exposure will differentially modulate basal or `tonic' DA levels and phasic DA release to ultimately enhance DA signaling. Specific Aim 2 will examine how postsynaptic DA receptor function is altered in the BLA using in vitro whole cell patch clamp electrophysiology with innovative optogenetic approaches to measure postsynaptic DA receptor signaling. Our working hypothesis is that ethanol physical dependence will increase postsynaptic D1- and D2-like DA receptor signaling such that DA-mediated inhibition of GABAergic function is up-regulated. Specific Aim 3 will place the detailed cellular effects of dependence on DA signaling within a whole-animal context by integrating optogenetic control of DA release with in vivo measures of BLA-dependent behaviors. Our working hypothesis is that dependence-related changes in dopamine neurotransmission and signaling ultimately control withdrawal-dependent anxiety-like behavior. The proposed work employs a unique and highly integrated experimental approach to provide unparalleled insight into the neurobiological mechanisms governing the negative reinforcing effects of chronic ethanol exposure. Ultimately, these studies will provide insight into potential cellular mechanisms governing abuse and relapse in human alcoholics.