Regulated trafficking and compartmentalized signaling of opioid receptors

这一建议解决了三角洲阿片受体(DOR)生物学的一个令人兴奋的方面，DOR是一种原型 和临床相关的G蛋白偶联受体(GPCR)，长期以来一直被认为是一个有希望的靶点 治疗疼痛和阿片成瘾--这两种高度流行和并存的疾病。疼痛，最常见的一种 在医院出现的症状，目前主要通过阿片类止痛药来管理，这些止痛药针对的是u-阿片类药物。 受体。虽然有效，但由于广泛的副作用和很高的成瘾可能性，它们的使用受到高度限制。 DOR激活是一种很有前途的替代策略，可以在不导致成瘾的情况下减轻疼痛，因为受体不是 在奖励途径中表达很多。而表达DOR的神经元对不同通道的贡献 这一策略的关键问题是在体内有效地靶向DOR 一直都很难。DOR激动剂在孤立的系统中有效地激活受体和信号，但它们表明 体内止痛反应差。中枢作用的DOR激动剂可以在高剂量的动物模型中抑制疼痛， 但它们也会导致抽搐，从而排除了使用它们的可能性。这种在体内的低止痛效果，是一种 针对DOR的止痛药开发的关键限制因素正在不同层面上进行研究，包括在 药理和神经回路水平。然而，我们仍然没有完全了解其背后的机制 这种低效率。有趣的是，与许多其他GPCR相比，DOR的独特之处在于它的本地化 大部分在细胞内表达，在细胞表面表达很少。我们对机制的证明 实验表明，将DOR重新定位到神经元表面会增加DOR激动剂的效果， 这暗示了一个令人兴奋的想法，即这种细胞内的位置是导致低止痛效力的原因。 因此，了解DOR是如何以及为什么在神经元中存在于细胞内，对于理解阿片类药物生理学至关重要 并将DOR发展为疼痛管理的目标。该提案涉及机制和 DOR定位对神经元细胞内隔间的影响。我们假设这个序列- DOR的特异性相互作用介导了新合成的DOR在细胞内隔间的滞留 神经元，并允许隔室特定的信号。我们将通过确定机制来检验我们的假设 DOR在细胞内的滞留，并测试这些机制在隔室特异性DOR信号中的作用。 具体地说，我们将遵循两个目标：1)确定调控DOR在细胞内定位的机制 隔室，以及2)研究来自细胞内膜的隔室特异性DOR信号。我们 将使用有重点的、以候选人为基础的和公正的全基因组方法来追求这些目标。完成 该项目将验证DOR如何定位到细胞内的一个新的和生理相关的模型 神经元中的隔间，确定DOR运输的关键原则，这些原则将在整个家族中广泛相关 临床相关蛋白，并潜在地识别靶向DOR的新的和未尝试的策略的靶点 疼痛管理。

This proposal addresses an exciting aspect of the biology of the delta opioid receptor (DOR), a prototypical and clinically relevant G protein-coupled receptor (GPCR) that has long been considered a promising target for treating pain and opioid addiction - two highly prevalent and comorbid diseases. Pain, one of the most common symptoms presented at hospitals, is currently managed primarily by opioid analgesics that target the mu-opioid receptor. While effective, their use is highly limited due to extensive side effects and a high potential for addiction. DOR activation is a promising alternate strategy to reduce pain without causing addiction as the receptor is not expressed much in the reward pathway. While the contribution of DOR-expressing neurons to distinct modalities of pain is being heavily explored, the critical problem with this strategy is that effectively targeting DOR in vivo has been difficult. DOR agonists activate the receptor and signal efficiently in isolated systems, but they show poor analgesic responses in vivo. Centrally acting DOR agonists can inhibit pain at high doses in animal models, but they also induce convulsions which preclude their use. This low analgesic effectiveness in vivo, which is a critical limiting factor in developing analgesics targeting DOR, is being studied at various levels including at the pharmacological and neural circuit level. However, we still do not fully understand the mechanisms underlying this low effectiveness. Interestingly, DOR is unique when compared to many other GPCRs in that it localizes mostly to intracellular compartments, with very little expressed on the cell surface. Our proof of mechanism experiments showing that relocating DOR to the neuronal surface increases the effectiveness of DOR agonists, suggesting the exciting idea that this intracellular location is what contributes to the low analgesic potency. Understanding how and why DOR is intracellular in neurons is therefore critical to understand opioid physiology and to develop DOR as a target for pain management. This proposal addresses the mechanisms and consequences of DOR localization to intracellular compartments in neurons. We hypothesize that sequence- specific interactions of DOR mediate retention of newly synthesized DOR in intracellular compartments in neurons and allow compartment-specific signaling. We will test our hypothesis by determining the mechanisms of DOR intracellular retention and testing the role of these mechanisms in compartment-specific DOR signaling. Specifically, we will follow two aims, 1) To determine the mechanisms regulating DOR localization in intracellular compartments, and 2) To investigate compartment-specific DOR signaling from intracellular membranes. We will use focused candidate-based and unbiased genome-wide approaches to pursue these aims. Completion of this project will validate a novel and physiologically relevant model for how DOR is localized to intracellular compartments in neurons, identify key principles of DOR trafficking that will be broadly relevant across this family of clinically relevant proteins, and potentially identify targets for new and untried strategies for targeting DOR in pain management.