Optogenetic Analysis of Different Forms of Aversion-Resistant Ethanol Intake

描述（由申请人提供）：人类酗酒者经历复发和强迫性寻求酒精的主要时期，尽管有不利的经济、社会和身体后果，但酒精摄入仍然存在。虽然强迫性是酒精中毒治疗的一个主要临床障碍，因为它会促进复发和持续饮酒，但实际上我们对驱动强迫性酒精摄入的分子和神经机制一无所知。人类研究表明，强迫行为可能是由前额皮质区域调节的，该区域向伏隔核（NAcore）发送谷氨酸能输入，而NAcore谷氨酸受体信号对于许多动机性、成瘾性和习惯性行为的表达至关重要。尤其是岛叶皮层与产生强烈的上瘾欲望有关，至少对尼古丁是如此。在这里，本研究的主要目标是一组探索性的大鼠实验，以确定一个共同的分子回路（脑岛输入到NAcore和NAcore NMDA受体的激活）是否驱动不同形式的厌恶性乙醇摄入，这可能模拟人类强迫性酒精摄入的某些方面。部分基于广泛的初步数据，我们假设，无论摄入模式（从瓶子里喝乙醇，操作杠杆按压乙醇）或与乙醇相结合的厌恶刺激的感觉方式（奎宁的味道，脚休克的体感），这种共同的分子电路都促进了厌恶性乙醇的摄入。因此，Specific Aim 1使用最先进的体内光遗传学技术来研究在不同形式的抗厌恶乙醇摄入过程中，抑制胰岛素到nacore的输入是否同样减少了乙醇摄入。特别是，我们使用光视紫红质，一种光激活的抑制蛋白，可以在特定的大脑区域内病毒表达，以选择性地调节轴突终端。此外，我们的体外电生理实验发现，NMDAR功能在胰岛素-NAcore输入下增强，因此，Specific Aim 2研究了NMDAR在NAcore内的药理阻断是否也抑制了不同摄入模型和不同的厌恶性刺激对乙醇的抗性摄入。我们还研究了当乙醇没有明显与厌恶刺激配对时，脑岛-脑核输入和脑核NMDARs是否介导摄入。谷氨酸能输入到NAcore可能在厌恶性酒精摄入的表达中发挥核心作用，我们将光遗传学与颅内药理学相结合，可能为强迫性酒精摄入的分子机制提供关键的治疗相关信息。

DESCRIPTION (provided by applicant): Human alcoholics undergo major periods of relapse and compulsive alcohol seeking, where alcohol intake persists despite adverse economic, social, and physical consequences. Although compulsion represents a major clinical hurdle to treatment of alcoholism by promoting relapse and continued alcohol intake, virtually nothing is known about molecular and neuronal mechanisms that drive compulsive aspects of alcohol intake. Human studies suggest that compulsive behavior may be regulated by frontal cortical areas that send glutamatergic inputs to the nucleus accumbens core (NAcore), and NAcore glutamate receptor signaling is critical for the expression of many motivated, addictive, and perhaps habitual behaviors. The Insular cortex in particular is implicated in generating strong drives that promote addiction, at least for nicotine. Here, the major goal of this proposal is an exploratory set of rat experiments addressing whether a common molecular circuit (Insula inputs to the NAcore and activation of NAcore NMDA receptors) drives different forms of aversion-resistant ethanol intake, which may model some aspects of human compulsive alcohol intake. We hypothesize, based in part on extensive preliminary data, that this common molecular circuit promotes aversion-resistant ethanol intake regardless of intake model (drinking ethanol from a bottle, operant lever pressing for ethanol) or the sensory modality of the aversive stimulus paired with ethanol (taste for quinine, somatosensory for footshock). Thus, Specific Aim 1 uses state-of-the- art optogenetic techniques in vivo to examine whether inhibiting Insula-to-NAcore inputs similarly reduces ethanol intake during the different forms of aversion-resistant ethanol intake. In particular, we use halorhodopsin, a light-activated inhibitory protein than can be viraly expressed within a particular brain region to selectively modulate axon terminals. Also, our in vitro electrophysiology experiments find enhanced NMDAR function under Insula-to-NAcore inputs, and thus Specific Aim 2 examines whether pharmacological block of NMDARs within the NAcore also inhibits aversion-resistant ethanol intake with different intake models and different aversive stimuli paired with ethanol. We also examine whether Insula-NAcore inputs and NAcore NMDARs mediate intake when ethanol is not overtly paired with an aversive stimuli. Glutamatergic inputs to the NAcore likely play a central role in the expression of aversion-resistant alcohol intake, and our use of optogenetics in combination with intracranial pharmacology is likely to provide critical and therapeutically relevant information about molecular mechanisms underlying compulsive alcohol intake.