Enhancement of the endogenous opioid system by ketamine

项目摘要： 内源性阿片系统参与许多不同的生物学过程，并有助于一系列 具有巨大的健康和社会后果的疾病，如阿片类药物使用障碍，疼痛，和重大 抑郁症内源性阿片系统由数十种阿片肽组成， 不同的前体蛋白，和三种细胞表面受体，这些受体被这些肽激活， 鸦片和合成毒品。最近，氯胺酮已成为一种有效的治疗方法， 抑郁症和慢性疼痛，并且还可以有效治疗阿片类药物使用障碍。而克他命 麻醉作用是由于其抑制谷氨酸受体的亚型，其其他机制， 生物效应仍然难以捉摸。我们实验室和其他研究小组的研究发现， 与内源性阿片系统相互作用，但这些相互作用的全部范围尚不清楚。 此外，单剂量氯胺酮在数分钟/小时内产生快速效应，以及持续效应 持续数天/数周。长期效果显著，不像氯胺酮的麻醉作用， 与血浆水平相关。我们的中心假设是氯胺酮与内源性 阿片类药物系统产生快速和长期的影响。这一假设得到了我们初步的支持。 数据显示氯胺酮作为阿片受体的正变构调节剂， 内源性阿片肽 我们的中心假设将在三个高度相关的具体目标中得到检验。目标1A将 使用培养的细胞来表征氯胺酮作为内源性阿片系统的变构调节剂的活性， 细胞和膜制品。这些研究将调查氯胺酮的短期和长期影响， （包括外消旋氯胺酮以及立体异构体S-和R-氯胺酮）和主要生物活性代谢物 （去甲氯胺酮和羟基去甲氯胺酮）。使用脑切片中的电生理学，Aim 1B将测量急性 氯胺酮对阿片受体介导的信号传导的影响及氯胺酮引起的持续性变化 局目的2A将测试氯胺酮与高度纯化的阿片受体制剂的相互作用。 目的2B将确定μ阿片受体与氯胺酮和肽复合物的冷冻电镜结构 配体。目的3研究氯胺酮对小鼠脑内内源性阿片肽水平的影响。 总的来说，这些研究将提供对氯胺酮与 内源性阿片样物质系统，提供机制来解释这种药物的快速和长期作用。 从历史上看，了解药物的分子靶点有助于深入了解疾病的病理生理学 和新疗法。因为关于氯胺酮作为抗抑郁药的机制知之甚少 以及这些疾病与阿片类药物使用障碍的共病，我们的研究旨在确定 氯胺酮与内源性阿片系统的相互作用与人体健康密切相关。

Project Summary: The endogenous opioid system is involved in many diverse biological processes and contributes to a range of disorders that have enormous health and societal consequences such as opioid use disorder, pain, and major depressive disorder. The endogenous opioid system consists of dozens of opioid peptides derived from three distinct precursor proteins, and three cell surface receptors that are activated by these peptides as well as by opiates and synthetic drugs. Recently, ketamine has emerged as an effective treatment for both major depressive disorder and chronic pain, and may also be effective in treating opioid use disorder. While ketamine’s anesthetic actions are due to its inhibition of a subtype of glutamate receptors, the mechanisms of its other biological effects remain elusive. Studies from our laboratory and other groups have found that ketamine interacts with the endogenous opioid system, however the full scope of these interactions is not known. Moreover, a single dose of ketamine produces rapid effects within minutes/hours, as well as sustained effects that last days/weeks. The long-term effects are remarkable and unlike the anesthetic action of ketamine which correlates with plasma levels. Our central hypothesis is that ketamine interacts with the endogenous opioid system to produce both rapid and long-term effects. This hypothesis is supported by our preliminary data showing that ketamine acts as a positive allosteric modulator of opioid receptors to synergize with endogenous opioid peptides. Our central hypothesis will be tested in three Specific Aims that are highly interrelated. Aim 1A will characterize the activity of ketamine as an allosteric modulator of the endogenous opioid system using cultured cells and membrane preparations. These studies will investigate short- and long-term effects of ketamine (including racemic ketamine and the stereoisomers S- and R-ketamine) and major bioactive metabolites (norketamine and hydroxynorketamine). Using electrophysiology in brain slices, Aim 1B will measure acute effects of ketamine on opioid receptor-mediated signaling and persistent changes induced by in vivo ketamine administration. Aim 2A will test the interaction of ketamine with highly purified preparations of opioid receptors. Aim 2B will determine the cryoEM structure of the mu opioid receptor in complex with ketamine and a peptide ligand. Aim 3 will examine the effect of ketamine on the levels of endogenous opioid peptides in mouse brain. Collectively, these studies will provide a solid understanding of the interactions of ketamine with the endogenous opioid system, providing mechanisms to account for the rapid and long-term effects of this drug. Historically, understanding the molecular targets of drugs have led to insights into the pathophysiology of disease and novel therapeutics. Because little is known regarding the mechanism(s) of ketamine as an antidepressant and analgesic, and the comorbidity of these diseases with opioid use disorder, our studies to determine the interaction of ketamine and the endogenous opioid system are highly relevant to human health.