Amygdala Modulation of Adolescent Alcohol Effects on Pain

摘要 青少年时期的酒精会在大脑中产生永久性的适应不良变化。青少年饮酒者 报告称，成年人的疼痛障碍患病率更高，功能性磁共振成像显示， 与不喝酒的人相比，杏仁核的激活程度更高。杏仁核，更确切地说是中央核 杏仁核（CeA）是痛觉相关脑区谷氨酸传递的关键脑区 地区CeA对酒精敏感，但根据暴露时间的不同，结果也不同。 在成年啮齿动物模型中，长期饮酒通过上调N-甲基-D-天冬氨酸（NMDAR）增加CeA活性 亚基和AMPAR介导的兴奋性传递导致疼痛处理增加。这些影响是 短暂的，因为NMDAR亚基表达和谷氨酸传递的变化在1-2周内恢复正常 暴露后。在青少年酒精暴露模型中， 信号持续到成年期的扩展杏仁核，这表明青少年酒精的影响更多 坚持不懈黑皮质素系统和神经特异性黑皮质素4受体（MC 4 R）在小鼠黑素细胞中高度表达， 并调节神经递质的传递MC 4 R在突触后增加AMPAR，调节 NMDAR功能，并增加其他疼痛相关区域的谷氨酸信号。黑素皮质素信号在 杏仁核调节疼痛相关行为，因为CeA内MC 4 R拮抗作用逆转了酒精诱导的 成年大鼠的痛觉过敏。成人酒精暴露降低MC 4 R表达，但青少年酒精暴露 增加成年期MC 4 R的表达。黑皮质素系统如何调节谷氨酸传输 青少年酒精对CeA谷氨酸和黑皮质素系统的纵向影响，以及如何 黑皮质素系统纵向调节疼痛处理还有待解决。使用大鼠模型， 我们将测试青少年酒精暴露对黑皮质素和CeA中的多巴胺能信号的影响， 慢性MC 4 R拮抗作用在此期间对细胞效应的影响，我们将测试MC 4 R拮抗作用 在成年期对青少年间歇性乙醇（AIE）蒸汽模型引起的痛觉过敏。男性和 雌性Wistar大鼠从出生后第28天（PND）至PND 56天接受酒精蒸汽，其中一组接受 在此期间，通过脑室内微型泵进行慢性MC 4 R拮抗。我们将使用Western blot 检测AIE（PND）后即刻AMPAR和NMDAR亚基及黑素皮质素系统蛋白表达 56)成年期（PND 86）。我们将使用切片电生理学来测量脑电传递 在青春期和成年期的CeA中，以及MC 4 R药物对其的调节。最后，我们将测试效果 CeA和鼻内MC 4 R拮抗剂对成年期伤害感受的影响。我们的假设是AIE 在介导AIE的CeA中产生黑皮质素和多巴胺能信号的持久增加， 诱导的痛觉过敏，在青春期慢性MC 4 R阻滞逆转AIE诱导的细胞 结果表明，MC 4 R拮抗作用在成年期将挽救AIE诱导的痛觉过敏。

Abstract Alcohol during the adolescent period produces permanent maladaptive changes in the brain. Adolescent drinkers report greater prevalence of pain disorders in adulthood, and functional magnetic resonance imaging has shown greater amygdala activation compared to non-drinkers. The amygdala, and more specifically the central nucleus of the amygdala (CeA) is a key brain region in pain processing by glutamate transmission from pain-related brain regions. The CeA is sensitive to alcohol, however there are divergent results depending on timing of exposure. In adult rodent models, chronic alcohol increases CeA activity by upregulating N-methyl-D-Aspartate (NMDAR) subunits and AMPAR mediated excitatory transmission leading to increased pain processing. These effects are transient, as changes in NMDAR subunit expression and glutamate transmission normalize within 1-2 weeks following exposure. In an adolescent alcohol exposure model, increases in NMDAR subunit and glutamate signaling lasts into adulthood in the extended amygdala, suggesting adolescent alcohol effects are more persistent. The melanocortin system and neural specific melanocortin 4 receptor (MC4R) are highly expressed in the CeA and regulate glutamatergic transmission. MC4R increases AMPAR post-synaptically, modulates NMDAR function, and increases glutamate signaling in other pain-related regions. Melanocortin signaling in the amygdala modulates pain-related behaviors as intra-CeA MC4R antagonism reverses alcohol-induced hyperalgesia in adult rats. Adult alcohol exposure decreases MC4R expression, but adolescent alcohol exposure increases MC4R expression in adulthood. How the melanocortin system modulates glutamate transmission in the CeA, the longitudinal effects of adolescent alcohol on CeA glutamate and melanocortin systems, and how the melanocortin system modulates pain processing longitudinally have yet to be addressed. Using a rat model, we will test the effect of adolescent alcohol exposure on melanocortin and glutamatergic signaling in CeA, the effect of chronic MC4R antagonism on cellular effects during this period, and we will test MC4R antagonism during adulthood on hyperalgesia induced by an adolescent intermittent ethanol (AIE) vapor model. Male and female Wistar rats will receive alcohol vapor from postnatal day (PND) 28 to PND 56, with one cohort receiving chronic MC4R antagonism via intracerebroventricular minipumps during this period. We will use Western blots to measure AMPAR and NMDAR subunits, and melanocortin system protein expression acutely after AIE (PND 56) and during adulthood (PND 86). We will use slice electrophysiology to measure glutamatergic transmission in the CeA during adolescence and adulthood, and its modulation by MC4R drugs. Finally, we will test the effects of intra-CeA and intranasal MC4R antagonism on nociception in adulthood. Our hypothesis is that AIE produces lasting increases in melanocortin and glutamatergic signaling in the CeA that mediates AIE- induced hyperalgesia, that chronic MC4R blockade during adolescence reverses AIE-induced cellular effects, and that MC4R antagonism in adulthood will rescue AIE-induced hyperalgesia.