Delta-kappa opioid receptor interactions: Ligand-dependent effects

说明(申请人提供)：作用于5阿片受体(MOR)的激动剂(如吗啡及其类似物)是止痛的主要药物；但有严重的不良反应(如依赖性)以及限制其使用的社会和法律问题。因此，阿片类药物的外周镇痛作用引起了人们的极大兴趣。除了MOR，4阿片受体(DOR)也是一个有吸引力的药物作用靶点，因为它的副作用比MOR激活时要少。但总体而言，4种激动剂促进镇痛的效果为弱至中度。我们的初步数据表明，DOR激动剂的亲和力和/或疗效可以受到k阿片受体(KOR)拮抗剂Nor-BNI的调节。在R21的探索性应用中，我们建议在大鼠三叉神经节原代培养和用于镇痛的行为模型中研究k阿片拮抗剂对DOR系统的调节。我们的具体目标是：1)考察KOR拮抗剂对选定的DOR激动剂的疗效和效力的影响。为此，我们将确定KOR拮抗剂Nor-BNI对原代培养的大鼠三叉神经节感觉神经元对特定DOR激动剂的浓度-反应曲线的影响。我们还将研究不同的KOR拮抗剂对DOR激动剂DPDPE和DADLE的浓度-反应曲线的影响。2)探讨KOR拮抗剂调节DOR激动剂反应性的机制。我们的初步数据表明，与NOR-BNI一起占据KOR会增加或减少DOR激动剂(分别是DPDPE和DADLE)的效力。为此，我们将通过竞争结合实验来验证NOR-BNI的作用是由于激动剂对DOR亲和力的变化所致的假设。我们还将利用KOR的siRNA击倒来确定NOR-BNI的作用是否由KOR介导。3)在热性和机械性痛觉异常的行为学测试中，观察KOR拮抗剂对DOR激动剂反应性的影响。这一目标提供了目标1-2的基础工作的翻译延伸。在外周镇痛的行为学实验中，我们将确定KOR与Nor-BNI拮抗剂一起使用对一系列DOR激动剂的量效曲线的影响。我们还将研究不同的KOR拮抗剂对DOR激动剂DPDPE的浓度-反应曲线的影响。如果我们的初步结果得到证实，可能会开发出一种治疗效果更好、选择性更高的疼痛控制的新药理学方法。 公共卫生相关性：外周的4阿片受体(DOR)是止痛药的一个有吸引力的靶点，但总的来说，4种阿片受体激动剂通过外周作用促进镇痛的效果是弱到中等的，而且是可变的。在R21的探索性应用中，我们建议在大鼠三叉神经节原代培养和用于镇痛的行为模型中研究k阿片拮抗剂对DOR系统的调节。所获得的结果将为更全面的研究(RO1)提供一个框架，以调节伤害性感受器中DOR的疗效，并可能导致开发一种新的药理学方法来控制疼痛，提高治疗效率和选择性。

DESCRIPTION (provided by applicant): Agonists acting at the 5 opioid receptor (MOR) (e.g. morphine and its analogs) are the mainstay of pain management; however there are serious adverse effects (e.g. dependence) and social and legal issues which limit their use. Consequently there has been considerable interest in the peripheral analgesic effects of opioids. In addition to MOR, the 4 opioid receptor (DOR) is an attractive target for drug action since there are fewer adverse effects than with MOR activation. However, in general, the efficacy of 4 agonists to promote analgesia is weak to moderate. Our preliminary data suggest that the affinity and/or efficacy of DOR agonists can be regulated by the k opioid receptor (KOR) antagonist, nor-BNI. In this R21 exploratory application, we propose to study the regulation of DOR systems by k opioid antagonists in primary cultures of rat trigeminal ganglion neurons and in behavioral models for analgesia. Our specific aims are: 1) To examine the effect of KOR antagonists on the efficacy and potency of selected DOR agonists. In this aim, we will determine the effect of the KOR antagonist, nor-BNI, on concentration-response curves to selected DOR agonists in primary cultures of rat sensory neurons of the trigeminal ganglion. We will also examine the effect of different KOR antagonists on the concentration-response curve to the DOR agonists DPDPE and DADLE. 2) To assess the mechanism by which KOR antagonists regulate DOR agonist responsiveness. Our preliminary data indicate that KOR occupancy with nor-BNI increases or decreases the potency of DOR agonists (DPDPE and DADLE, respectively). In this aim we will test the hypothesis that the effect of nor-BNI is due to a change in agonist affinity for DOR using competition binding experiments. We will also determine if the effect of nor-BNI is mediated by KOR using siRNA knock-down of KOR. 3) To investigate the effect of KOR antagonists on responsiveness to DOR agonists in a behavioral assay of thermal and mechanical allodynia. This aim provides a translational extension of the foundation work of Aims 1-2. We will determine the effect of occupancy of KOR with the antagonist nor-BNI on concentration-response curves to a series of DOR agonists in a behavioral assay of peripheral analgesia. We will also examine the effect of different KOR antagonists on the concentration-response curve to the DOR agonist DPDPE. If our preliminary results are substantiated, a new pharmacological approach for pain management with improved therapeutic efficacy and increased selectivity may be developed. PUBLIC HEALTH RELEVANCE: The 4 opioid receptor (DOR) in the periphery is an attractive target for analgesic drugs, however, in general, the efficacy of 4 opioid receptor agonists to promote analgesia by acting in the periphery is weak to moderate and variable. In this R21 exploratory application, we propose to study the regulation of DOR systems by k opioid antagonists in primary cultures of rat trigeminal ganglion neurons and in behavioral models for analgesia. The results obtained will provide a framework for a more comprehensive study (RO1) of the regulation of DOR agonist efficacy in nociceptors and may lead to the development of a new pharmacological approach for pain management with improved therapeutic efficacy and increased selectivity.