Central Amygdala CART modulates ethanol withdrawal induced anxiety

描述（由申请人提供）：乙醇依赖是一种普遍的痛苦，每年造成数千亿美元的社会成本。乙醇依赖的治疗因许多因素而复杂，最明显的是复发。复发行为被认为是由乙醇戒断性焦虑（EWIA）引起的。参与EWIA的一个关键脑区是中央杏仁核（CeA）。中央杏仁核的病变，而不是基底外侧杏仁核的病变，导致戒断期间乙醇消耗减少。乙醇依赖引起的神经生物学改变是广泛存在的；在CeA中，这些变化包括谷氨酸能活性和NMDA受体表达的增加。在停药期间，这些增加导致CeA的过度活化；这种过度激活被认为介导了导致复发的戒断性焦虑。可卡因和安非他明调节转录本（CART）是一种与乙醇依赖有关的新型神经肽。它在CeA中表达，并在急性停药期间增加表达。此外，据报道CART可增强NMDA受体介导的电流。本提案的首要假设是CART是EWIA的关键中介。为了验证这一点，我们将对依赖和戒断期间的CART表达进行全面检查。乙醇依赖将通过两种模型诱导：慢性乙醇治疗（CET）和慢性间歇乙醇（CIE）。CIE包含戒断成分，可能涉及戒断引起的焦虑。野生型（WT）和CART敲除型（KO）小鼠的依赖性将采用CET和CIE进行比较。对WT和KO小鼠在停药0、24和72小时的EWIA进行检查。皮质酮elisa和免疫组织化学也将进行。还将评估和比较WT和KO小鼠对急性应激源的应激性饮酒和皮质酮水平。本提案的后半部分将首先在我们手中验证CART的NMDA增强作用，特别是在CeA中。然后，在WT和KO小鼠中进行全面的电生理电压钳分析和比较乙醇幼稚、CET和CIE小鼠在0、24和72小时戒断时CeA中NMDA受体介导的电流。这些实验的发现将阐明EWIA的基础和CART在EWIA诱导的复发中的作用。此外，这些结果将为治疗干预和预防依赖乙醇和其他药物滥用的个体复发提供新的靶点。

DESCRIPTION (provided by applicant): Ethanol dependence is a widespread affliction with societal costs in the hundreds of billions of dollars every year. Treatment of ethanol dependence is complicated by many factors, most notably relapse. Relapse behaviors are believed to result from ethanol withdrawal-induced anxiety (EWIA). A brain region critically involved in EWIA is the central amygdala (CeA). Lesions of the central, but not basolateral, amygdala resulted in reduced ethanol consumption during withdrawal. Neurobiological alterations induced by ethanol dependence are widespread; in the CeA, these changes include increased glutamatergic activity and NMDA receptor expression. During withdrawal, these increases lead to hyperactivation of the CeA; this hyperactivation is believed to mediate the withdrawal-induced anxiety that underlies relapse. Cocaine- and amphetamine-regulated transcript (CART) is a novel neuropeptide implicatied in ethanol dependence. It is expressed in the CeA and has been shown to increase expression here during acute withdrawal. Additionally, CART, has been reported to potentiate NMDA receptor-mediated currents. The overarching hypothesis of this proposal is that CART is a critical mediator of EWIA. To test this, a comprehensive examination of CART expression during dependence and withdrawal will be conducted. Ethanol dependence will-be induced using two models: chronic ethanol treatment (CET) and chronic intermittent ethanol (CIE). CIE contains a withdrawal component that presumably involves withdrawal induced anxiety. Dependence liability in wild type (WT) and CART knockout (KO) mice will compared using both CET and CIE. An examination of EWIA in the WT and KO mice at 0, 24, and 72 hours withdrawal will also be conducted. Corticosterone ELISAs and immunohistochemistry will be performed as well. Stress induced drinking and corticosterone levles in response to an acute stressor will also be assessed and compared between the WT and KO mice. The second half of this proposal will first verify in our hands, the NMDA potentiating actions of CART, specifically in the CeA. Then, a comprehensive electrophysiological voltage clamp analysis and comparison of NMDA receptor-mediated currents in the CeA of ethanol naive, CET, and CIE mice at 0, 24, and 72 hours withdrawal in WT and KO mice will be performed. The findings from these experiments will elucidate the underpinnings of EWIA and the role of CART in EWIA-induced relapse. Furthermore, these results will provide a novel target for therapeutic intervention and prevention of relapse in individuals dependent on ethanol and possibly other drugs of abuse.