Physiological Implications of Opioid Receptor Regulation

描述(申请人提供)：吗啡，缓解疼痛的黄金标准，在临床上受到几种不良特性的限制，包括耐受性、依赖性和呼吸抑制。阿片类镇痛剂，如吗啡，主要通过激活u阿片受体(MuOR)来调节其生物学效应。MUOR是一种G蛋白偶联受体(GPCRR)，在受体脱敏的过程中受GPCRK(GRK)磷酸化和随后与Betaarrestins结合的调节。在小鼠体内，Betaarrestin2(Betaarr2)的基因消融似乎具有矛盾的效应；它导致增强和延长吗啡的镇痛作用，并显著降低吗啡的耐受性。此外，这些小鼠在身体依赖性方面没有变化；相反，呼吸抑制实际上被消除了。在这个小鼠模型中，吗啡的作用接近于一种假想的、临床使用的最佳阿片类止痛剂。在Betaarr2基因敲除小鼠(betaarr2-KO)中，与吗啡镇痛相关的脑区，muOR与G蛋白的偶联增加，这可能与增强的镇痛反应有关。然而，Betaarr2在吗啡诱导的呼吸抑制中的作用尚不清楚。此外，尽管在Betaarr2-KO小鼠中增强了吗啡的止痛作用，但对其他阿片类药物如芬太尼和美沙酮的反应没有改变。我们假设GRK和Betaarrestins调节MUOR，这种调节的特异性由船上的阿片激动剂决定。因此，在MUOR水平上的这些调节差异可能是多种临床相关阿片类药物产生不同药理作用的基础，如吗啡、芬太尼、美沙酮和丁丙诺啡。我们有独特的机会通过研究缺乏单个GRK2(GRK2、GRK3、GRK4、GRK5和GRK6)或Betarr2的小鼠的阿片中介行为和生理反应来评估GRKs和Betarr2在体内阿片类药物反应中的作用。这些动物的神经化学改变将在行为研究的同时进行评估，重点是受体运输、受体脱敏和下游神经适应性变化。HEK-293细胞将被用作模型系统，以进一步阐明体内观察到的现象背后的分子机制。这项研究的总体目标是在确定药物对生理和病理条件的影响的特异性时，更好地了解MUOR调节。为了达到这一目标，这些研究集中于研究GRK和Parretins在阿片耐受性(AIM I)、依赖(AIM II)和阿片类药物引起的副作用(AIM III)发展过程中对MUOR调节的贡献。阐明MUOR调节的复杂性可能指向微调受体的反应性，以增加镇痛效果，限制滥用的可能性，并在开发阿片类药物治疗时消除不良副作用。

DESCRIPTION (provided by applicant): Morphine, the gold standard for pain relief, is clinically limited by several adverse properties including tolerance, dependence and the onset of respiratory suppression. Opiate analgesics, such as morphine, mediate their biological effects mainly via activation of the mu opioid receptor (muOR). The muOR, a G protein coupled receptor (GPCR), is regulated by GPCR kinase (GRK) phosphorylation and subsequent binding of betaarrestins in a process known as receptor desensitization. The genetic ablation of betaarrestin2 (betaarr2)in mice has seemingly paradoxical effects; it leads to enhanced and prolonged morphine analgesia and dramatically reduces morphine tolerance. Moreover, these mice display no change in physical dependence; in contrast, respiratory suppression is practically eliminated. The effects of morphine in this mouse model approach a hypothetical, optimal opiate analgesic for clinical use. The coupling of the muOR to G proteins is elevated in brain regions associated with morphine analgesia in the betaarr2knockout (betaarr2-KO) mice which may contribute to the enhanced analgesic responses. However, the role of betaarr2 in morphine-induced respiratory suppression is unclear. Furthermore, while morphine analgesia is enhanced in the betaarr2-KO mice, lresponses to other opiates such as fentany and methadone, are unaltered. We have hypothesized that GRKs and betaarrestins regulate muORs and the specificity of this regulation is determined by the opiate agonist onboard. These regulatory differences at the level of the muOR may thereby underlie the diverse pharmacological effects produced by a wide-range of clinically relevant opiates such as morphine, fentanyl, methadone and buprenorphine. We have the unique opportunity to evaluate the contributions of GRKs and betarr2 to opiate responses in vivo by studying opiatemediated behaviors and physiological responses in strains of mice lacking individual GRKs (GRK2, GRK3, GRK4, GRK5, and GRK6) or betarr2. Neurochemical alterations will be assessed in these same animals in parallel to the behavioral studies with a focus on receptor trafficking, receptor desensitization, and downstream neuroadaptive changes. HEK-293 cells wil be used as a model system to further elucidate the molecular mechanisms underlying the observed in vivo phenomena. The overall objective of this study is to gain a greater understanding of muOR regulation in determining the specificity of drug effects on physiological and pathological conditions. Toward this goal, these studies focus on examining the contribution of GRKs and parrestins to muOR regulation in the development of opiate tolerance (AIM I), dependence (AIM II), and the side effects induced by opiates (AIM III). Illuminating the intricacies of muOR regulation may point to fine-tuning receptor responsiveness to increase analgesic efficacy, limit abuse liabilityand eliminate adverse side effects in developing opiate pharmaceutical therapies.