TY - JOUR
T1 - Cellular Assay to Study β-Arrestin Recruitment by the Cannabinoid Receptors 1 and 2
AU - Bouma, Jara
AU - Soethoudt, Marjolein
AU - van Gils, Noortje
AU - Xia, Lizi
AU - van der Stelt, Mario
AU - Heitman, Laura H.
N1 - Publisher Copyright: © 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
PY - 2023
Y1 - 2023
N2 - Cannabinoid receptor 1 (CB1R) and cannabinoid receptor 2 (CB2R) 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 CB1R and CB2R 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 CB1R and CB2R, of which some examples will be presented here.
AB - Cannabinoid receptor 1 (CB1R) and cannabinoid receptor 2 (CB2R) 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 CB1R and CB2R 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 CB1R and CB2R, of which some examples will be presented here.
KW - Biased signaling
KW - Cannabinoid receptors
KW - Desensitization
KW - DiscoverX PathHunter®
KW - Functional selectivity
KW - GPCRs
KW - Internalization
KW - Receptor signaling
KW - β-arrestin
UR - https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=85138458617&origin=inward
UR - https://www.ncbi.nlm.nih.gov/pubmed/36152187
U2 - https://doi.org/10.1007/978-1-0716-2728-0_15
DO - https://doi.org/10.1007/978-1-0716-2728-0_15
M3 - Article
C2 - 36152187
SN - 1064-3745
VL - 2576
SP - 189
EP - 199
JO - Methods in molecular biology (Clifton, N.J.)
JF - Methods in molecular biology (Clifton, N.J.)
ER -