Institute of Translational Medicine and New Drug Development, China Medical University, Taichung 40402, Taiwan.
Center for Drug Abuse and Addiction, China Medical University Hospital, Taichung 40447, Taiwan.
Int J Mol Sci. 2023 Jan 5;24(2):1048. doi: 10.3390/ijms24021048.
We attempted to examine the alterations elicited by opioids via coexpressed μ-opioid (MOP) and nociceptin/orphanin FQ (NOP) receptors for receptor localization and Erk1/2 (p44/42 MAPK) in human embryonic kidney (HEK) 293 cells. Through two-photon microscopy, the proximity of MOP and NOP receptors was verified by fluorescence resonance energy transfer (FRET), and morphine but not buprenorphine facilitated the process of MOP-NOP heterodimerization. Single-particle tracking (SPT) further revealed that morphine or buprenorphine hindered the movement of the MOP-NOP heterodimers. After exposure to morphine or buprenorphine, receptor localization on lipid rafts was detected by immunocytochemistry, and phosphorylation of Erk1/2 was determined by immunoblotting in HEK 293 cells expressing MOP, NOP, or MOP+NOP receptors. Colocalization of MOP and NOP on lipid rafts was enhanced by morphine but not buprenorphine. Morphine stimulated the phosphorylation of Erk1/2 with a similar potency in HEK 293 cells expressing MOP and MOP+NOP receptors, but buprenorphine appeared to activate Erk1/2 solely through NOP receptors. Our results suggest that opioids can fine-tune the cellular localization of opioid receptors and phosphorylation of Erk1/2 in MOP+NOP-expressing cells.
我们试图通过在人胚肾（HEK）293 细胞中共表达μ-阿片受体（MOP）和孤啡肽/孤啡肽受体（NOP）来检测阿片类药物引起的改变，以研究受体定位和 Erk1/2（p44/42 MAPK）。通过双光子显微镜，通过荧光共振能量转移（FRET）验证了 MOP 和 NOP 受体的接近程度，并且吗啡而不是丁丙诺啡促进了 MOP-NOP 异二聚体化的过程。单颗粒跟踪（SPT）进一步表明，吗啡或丁丙诺啡阻碍了 MOP-NOP 异二聚体的运动。在暴露于吗啡或丁丙诺啡后，通过免疫细胞化学检测到表达 MOP、NOP 或 MOP+NOP 受体的 HEK 293 细胞中脂质筏上的受体定位，并通过免疫印迹法测定 Erk1/2 的磷酸化。吗啡增强了 MOP 和 NOP 在脂质筏上的共定位，但丁丙诺啡没有。吗啡以相似的效力刺激表达 MOP 和 MOP+NOP 受体的 HEK 293 细胞中 Erk1/2 的磷酸化，但丁丙诺啡似乎仅通过 NOP 受体激活 Erk1/2。我们的结果表明，阿片类药物可以精细调节表达 MOP+NOP 的细胞中阿片受体的细胞定位和 Erk1/2 的磷酸化。
