Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Leiden, the Netherlands.
Department of Molecular Physiology, Leiden Institute of Chemistry, Leiden University, Leiden, the Netherlands.
Methods Mol Biol. 2023;2576:189-199. doi: 10.1007/978-1-0716-2728-0_15.
Cannabinoid receptor 1 (CBR) and cannabinoid receptor 2 (CBR) are G protein-coupled receptors (GPCRs) that activate a variety of pathways upon activation by (partial) agonists including the G protein pathway and the recruitment of β-arrestins. Differences in the activation level of these pathways lead to biased signaling. Here, we describe a detailed protocol to characterize the potency and efficacy of ligands to induce or inhibit β-arrestin recruitment to the human CBR and CBR using the PathHunter assay. This is a cellular assay that uses a β-galactosidase complementation system which has a chemiluminescent read-out and can be performed in 384-well plates. We have successfully used this assay to characterize a set of reference ligands (both agonists, antagonists, and an inverse agonist) on human CBR and CBR, of which some examples will be presented here.
大麻素受体 1（CBR）和大麻素受体 2（CBR）是 G 蛋白偶联受体（GPCR），它们在被（部分）激动剂激活后会激活各种途径，包括 G 蛋白途径和β-arrestin 的募集。这些途径的激活水平的差异导致了偏向信号传导。在这里，我们描述了一个详细的方案，以使用 PathHunter 测定法来表征配体诱导或抑制人 CBR 和 CBR 中β-arrestin 募集的效力和效力。这是一种细胞测定法，使用β-半乳糖苷酶互补系统，具有化学发光读数，并且可以在 384 孔板中进行。我们已经成功地使用该测定法对一组参考配体（激动剂、拮抗剂和反向激动剂）在人 CBR 和 CBR 上进行了表征，其中一些示例将在此处呈现。
