Investigation of nociceptive endogenous opioid dynamics in the periaqueductal gray

项目总结/摘要 慢性疼痛是一种高度普遍和使人衰弱的临床问题，其对健康和精神健康产生负面影响。 数百万人的生活质量。一个共同的和相对有效的战略，以提供急性缓解慢性 疼痛患者是通过阿片类化合物的处方。然而，阿片类镇痛药存在大量滥用 和过量负债，严重助长了持续的阿片类药物流行。这些负面影响 外源性阿片样物质是由它们在内源性μ阿片样物质受体表达（莫尔）脑中的扩散作用产生的 这些区域超出了它们打算调节的疼痛编码神经回路。为了满足紧迫的 由于对有效和安全的止痛药的需求，必须开发新的有针对性的疼痛治疗方法， 集中研究参与疼痛感知的内源性阿片样物质能细胞类型和神经回路（即， 伤害感受）和阿片样物质诱导的镇痛。腹外侧导水管周围灰质（vlPAG）在这方面是至关重要的 因为它可以通过细胞表达的莫尔和脑啡肽产生强有力的抗伤害感受， 它所包含的内容。然而，长期存在的问题仍然是关于内源性阿片信号动力学 在急性和慢性疼痛病症募集的vlPAG中。该提案将开始填补这些空白， 知识与新工具的组合：在确定的神经群体中的遗传重组，莫尔特异性 启动子病毒和荧光脑啡肽传感器。结合起来，这些方法可以实现前所未有的 在vlPAG中体内接近内源性阿片样物质传递的突触前和突触后组分。因此，在本发明中， 我将测试我的中心假设，即vlPAG包含功能性伤害感受性MOR表达集合 其由前脑和局部中间神经元释放的脑啡肽调节以产生抗伤害感受。在Aim中 1，我将使用经典的疼痛测定来鉴定分子身份和钙信号传导活性模式， vlPAG中的伤害感受性MOR表达神经系综（vlPAGNoci/莫尔）。我将进一步确定功能 vlPAGNoci/莫尔神经元在急性和炎症期间通过光遗传学抑制的抗伤害感受中的作用 疼痛状态。在目标2中，我将识别和操纵两个假定的脑啡肽能输入到vlPAG，局部vlPAG 中间神经元和中央杏仁核（CeM）投射的长程内侧核，以确定其 各自对伤害感受的贡献。这些脑啡肽传入如何与表达MOR的 还将确定vlPAG中的神经元。这些拟议实验的结果将推动我们的 了解内源性阿片样物质信号传导过程，参与和改变的急性和慢性 疼痛状况。通过阐明产生VIPAG的内源性阿片样物质回路的组分， 镇痛，缓解慢性疼痛可能通过未来的治疗来实现，这些治疗靶向该系统，同时缺乏 当前阿片类药物的有害成瘾性。完成本奖学金提案将实现我的 培训目标是扩大我的实验专业知识，并使自己成为疼痛和阿片类药物的专家 神经回路

PROJECT SUMMARY/ABSTRACT Chronic pain is a highly prevalent and debilitating clinical problem that negatively impacts the health and quality of life of millions of people. A common and relatively effective strategy to provide acute relief to chronic pain patients is through prescription of opioid compounds. However, opioid analgesics carry substantial abuse and overdose liabilities, contributing heavily to the ongoing opioid epidemic. These negative consequences of exogenous opioids result from their diffuse action at endogenous mu opioid receptor-expressing (MOR) brain regions beyond the pain-encoding neurocircuitry that they are intended to modulate. To meet the pressing demand for effective and safe analgesics, new, targeted pain therapies must be developed that emerge from focused research on the endogenous opioidergic cell types and neural circuits involved in pain perception (i.e., nociception) and opioid-induced analgesia. The ventrolateral periaqueductal gray (vlPAG) is critical in this regard as it can produce robust antinociception through MOR and the enkephalin peptides expressed by the cells and afferents it contains. Yet, long-standing questions remain concerning the endogenous opioid signaling dynamics in the vlPAG that are recruited by acute and chronic pain conditions. This proposal will begin to fill these gaps in knowledge with a combination of novel tools: genetic recombination in defined neural populations, MOR specific promoter viruses, and a fluorescent enkephalin sensor. Combined, these approaches allow for unprecedented in vivo access to the pre- and postsynaptic components of endogenous opioid transmission in the vlPAG. Thus, I will test my central hypothesis that the vlPAG contains a functional nociceptive MOR-expressing ensemble that is modulated by enkephalin release from forebrain and local interneurons to produce antinociception. In Aim 1, I will use classical pain assays to identify the molecular identity and calcium signaling activity patterns of the nociceptive MOR-expressing neural ensemble in the vlPAG (vlPAGNoci/MOR). I will further determine the functional role of vlPAGNoci/MOR neurons in antinociception through optogenetic inhibition during acute and inflammatory pain states. In Aim 2, I will identify and manipulate two putative enkephalinergic inputs to the vlPAG, local vlPAG interneurons and long-range medial nucleus of the central amygdala (CeM) projections, to determine their respective contributions to nociception. How these enkephalingeric afferents interact with the MOR-expressing neurons in the vlPAG will also be determined. The results of these proposed experiments will advance our understanding of endogenous opioid signaling processes that are engaged and altered by acute and chronic pain conditions. By elucidating the components of the endogenous opioid circuitry of the vlPAG that produce analgesia, relief from chronic pain may be realized by future therapies that target this system while lacking the harmful addictive properties of current opioid drugs. Completion of this Fellowship proposal will achieve my training goals to expand my experimental expertise and establish myself as an expert in pain and opioidergic neurocircuitry.