Regulation of binge-like ethanol intake by arcuate POMC projection neurons

摘要 内源性阿片系统强烈地牵涉到奖赏、强化和动机效应， 乙醇，如阿片受体拮抗剂纳洛酮和 纳美芬在减少乙醇摄入量、复发倾向和渴望方面的作用。动物研究表明， 乙醇激活脑内各种含有阿片肽的回路，包括脑内的 中皮质边缘奖赏回路和杏仁核。最近我们发现了多条证据 表明乙醇激活了下丘脑弓状核（ArcN）内的一部分神经元 表达阿黑皮素原（POMC），其产生许多生物活性神经肽，包括β- 内啡肽使用膜片钳电生理学，我们观察到浴中应用乙醇（5 - 40 mM） 增加记录的ArcN POMC神经元的约35%的放电频率。同样，使用FosB 免疫组织化学，我们证明了酗酒样乙醇摄入激活了大约一个亚组的ArcN POMC神经元，其中大部分合成β-内啡肽与β-MSH。逆行追踪显示有酗酒- 像乙醇摄入主要激活投射到杏仁核的ArcN POMC神经元， 投射到腹侧被盖区（VTA）或延髓核（NAc）的神经元。令人惊讶的是， 没有亚群划分的ArcN POMC神经元的调制对乙醇摄入没有影响。然而，在这方面， 我们推测，这些影响的缺乏是由于一些ArcN激活的非特异性， POMC神经元包含子电路。观察到酗酒样乙醇摄入的基线性别差异，其中 雌性小鼠比雄性小鼠消耗更多的乙醇，我们观察到ER β是 与ER β或孕酮相比，位于ArcN POMC神经元上的初级雌性激素受体 受体。总之，这些观察结果导致了我们的总体假设，即ArcN POMC神经元 对中脑边缘奖励系统区域的投射调节了酒精摄入量的狂欢， 主要通过ER依赖性机制。为了检验这一假设，我们制定了以下公式 既相互关联又相互独立的具体目标。在目标1中，我们将确定乙醇对 ArcN POMC投射神经元的神经生理学特性。在目标2中，我们将确定 特定ArcN POMC传出投射神经元对酒精摄入的化学遗传学调节。 最后，在目标3中，我们将确定POMC神经元上的ER β在酒精狂欢样调节中的作用 摄入和与中脑多巴胺神经元的潜在相互作用。总之，这些研究将阐明具体的 阿片类药物回路和机制调节酗酒样乙醇摄入，这将指导改善 用于治疗酒精使用障碍的神经调节和/或药理学方法。

ABSTRACT The endogenous opioid system is strongly implicated in the rewarding, reinforcing, and motivational effects of ethanol, as evidenced by the established clinical efficacy of the opioid receptor antagonists naltrexone and nalmefene in reducing ethanol intake, relapse propensity, and craving. Animal studies have demonstrated that ethanol activates various opioid peptide-containing circuits within the brain, including regions of the mesocorticolimbic reward circuitry and amygdala. Recently we have generated multiple lines of evidence indicating that ethanol activates a subset of neurons within the arcuate nucleus (ArcN) of the hypothalamus expressing pro-opiomelanocortin (POMC), which gives rise to numerous bioactive neuropeptides including - endorphin. Using patch clamp electrophysiology, we observed that bath application of ethanol (5-40 mM) increases the firing frequency of ~35% of recorded ArcN POMC neurons. Similarly, using FosB immunohistochemistry, we demonstrated that binge-like ethanol intake activates approximately a subset of ArcN POMC neurons, the majority of which synthesize -endorphin vs. -MSH. Retrograde tracing revealed binge- like ethanol intake primarily activates ArcN POMC neurons projecting to the amygdala, with fewer activated neurons projecting to the ventral tegmental area (VTA) or nucleus accumbens (NAc). Surprisingly, chemogenetic modulation of ArcN POMC neurons without subpopulation delineation had no effect on ethanol intake. However, we speculate that these lack of effects were due to the non-specific nature of activation of a number of ArcN POMC neuron containing subcircuits. Baseline sex differences in binge-like ethanol intake were observed, where female mice consumed significantly more ethanol than their male counterparts, and we observed that ER is the primary female sex hormone receptor located on ArcN POMC neurons as compared to ER or progesterone receptors. Together, these observations have led to our overarching hypothesis that ArcN POMC neuron projections to regions of the mesolimbic reward system regulate binge-like ethanol intake in a sex-dependent manner primarily via ER dependent mechanisms. To test this hypothesis, we have formulated the following inter-related yet independent Specific Aims. In Aim 1, we will determine the effects of ethanol on the neurophysiological properties of ArcN POMC projection neurons. In Aim 2, we will determine the effects of chemogenetic modulation of specific ArcN POMC efferent projection neurons on binge-like ethanol intake. Finally, in Aim 3, we will determine the role of ER on POMC neurons in the regulation of binge-like ethanol intake and potential interactions with midbrain dopamine neurons. Together, these studies will elucidate specific opioid circuits and mechanisms regulating binge-like ethanol intake, which will guide the improvement of neuromodulatory and/or pharmacological approaches for the treatment of alcohol use disorders.