Gomes Ivone, Filipovska Julija, Jordan Bryen A, Devi Lakshmi A
Department of Pharmacology, New York University School of Medicine, MSB 408, New York 10016, USA.
Methods. 2002 Aug;27(4):358-65. doi: 10.1016/s1046-2023(02)00094-4.
Opioid receptors belong to the family of G-protein-coupled receptors characterized by their seven transmembrane domains. The activation of these receptors by agonists such as morphine and endogenous opioid peptides leads to the activation of inhibitory G-proteins followed by a decrease in the levels of intracellular cAMP. Opioid receptor activation is also associated with the opening of K(+) channels and the inhibition of Ca(2+) channels. A number of investigations, prior to the development of opioid receptor cDNAs, suggested that opioid receptor types interacted with each other. Early pharmacological studies provided evidence for the probable interaction between opioid receptors. More recent studies using receptor selective antagonists, antisense oligonucleotides, or animals lacking opioid receptors further suggested that interactions between opioid receptor types could modulate their activity. We examined opioid receptor interactions using biochemical, biophysical, and pharmacological techniques. We used differential epitope tagging and selective immunoisolation of receptor complexes to demonstrate homotypic and heterotypic interactions between opioid receptor types. We also used the proximity-based bioluminescence resonance energy transfer assay to explore opioid receptor-receptor interactions in living cells. In this article we describe the biochemical and biophysical methods involved in the detection of receptor dimers. We also address some of the concerns and suggest precautions to be taken in studies examining receptor-receptor interactions.
阿片受体属于G蛋白偶联受体家族，其特征为具有七个跨膜结构域。吗啡和内源性阿片肽等激动剂激活这些受体后，会导致抑制性G蛋白的激活，随后细胞内cAMP水平降低。阿片受体激活还与K(+)通道的开放和Ca(2+)通道的抑制有关。在阿片受体cDNA开发之前的一些研究表明，阿片受体类型之间会相互作用。早期的药理学研究为阿片受体之间可能的相互作用提供了证据。最近使用受体选择性拮抗剂、反义寡核苷酸或缺乏阿片受体的动物进行的研究进一步表明，阿片受体类型之间的相互作用可以调节它们的活性。我们使用生化、生物物理和药理学技术研究阿片受体相互作用。我们使用差异表位标签和受体复合物的选择性免疫分离来证明阿片受体类型之间的同型和异型相互作用。我们还使用基于邻近的生物发光共振能量转移测定法来探索活细胞中的阿片受体-受体相互作用。在本文中，我们描述了检测受体二聚体所涉及的生化和生物物理方法。我们还讨论了一些问题，并对研究受体-受体相互作用时应采取的预防措施提出了建议。
