The Vollum Institute, Oregon Health & Science University, Portland, United States.
Medicinal Chemistry Core, Oregon Health & Science University, Portland, United States.
Elife. 2019 Oct 7;8:e49319. doi: 10.7554/eLife.49319.
Identifying neurons that have functional opioid receptors is fundamental for the understanding of the cellular, synaptic and systems actions of opioids. Current techniques are limited to post hoc analyses of fixed tissues. Here we developed a fluorescent probe, naltrexamine-acylimidazole (NAI), to label opioid receptors based on a chemical approach termed 'traceless affinity labeling'. In this approach, a high affinity antagonist naltrexamine is used as the guide molecule for a transferring reaction of acylimidazole at the receptor. This reaction generates a fluorescent dye covalently linked to the receptor while naltrexamine is liberated and leaves the binding site. The labeling induced by this reagent allowed visualization of opioid-sensitive neurons in rat and mouse brains without loss of function of the fluorescently labeled receptors. The ability to locate endogenous receptors in living tissues will aid considerably in establishing the distribution and physiological role of opioid receptors in the CNS of wild type animals.
鉴定具有功能性阿片受体的神经元对于理解阿片类药物的细胞、突触和系统作用至关重要。目前的技术仅限于对固定组织进行事后分析。在这里，我们开发了一种荧光探针，纳曲胺酰基咪唑（NAI），基于一种称为“无痕亲和力标记”的化学方法来标记阿片受体。在这种方法中，高亲和力拮抗剂纳曲胺被用作受体转移反应的导向分子，该反应在受体上将酰基咪唑转移到受体上，生成与受体共价结合的荧光染料，同时释放出纳曲胺并离开结合位点。该试剂诱导的标记允许在不影响荧光标记受体功能的情况下，可视化大鼠和小鼠大脑中的阿片敏感神经元。该能力将极大地帮助确定内源性受体在野生型动物中枢神经系统中的分布和生理作用。
