Characterization of human cannabinoid CB2 receptor coupled to chimeric Gαqi5 and Gαqo5 proteins

Cannabinoid CB2 receptors may couple to a variety of G proteins and intracellular effector systems to regulate physiological and pathophysiological processes involved in inflammatory and neuropathic pain. In this study, the coupling of cannabinoid hCB2 receptors to Gαqo5 and Gαqi5 proteins was studied and compared by investigating the pharmacological properties of HEK-293 cells co-expressing cannabinoid hCB2 with chimeric Gαqo5 (HEK-hCB2-Gqo5) or Gαqi5 (HEK-hCB2-Gqi5). Both cell lines were found to be amendable for measuring cannabinoid CB2 receptor agonist evoked Ca2+ mobilization in a high-throughput manner. Comparison of binding affinities of ligands in homogenates prepared from both cell lines revealed similar affinities for [3H]CP55,940 displacement with the following rank order: CP55,940 ~ WIN55,212-2 > SR144528 > JWH015 ~ AM1241 ~ AM630 > SR141617A ~ AM251. In comparison at cannabinoid hCB1 receptors: the rank order was: SR141617A ~ CP55,940>AM251 > WIN55,212-2 > AM1241 ~ SR144528 > JWH015 ~ AM630. No significant differences in cannabinoid receptor agonist (CP55,940 ~ WIN55,212-2 > JWH015) or antagonist (SR144528 ~ AM1241 > AM630 > AM251 ~ SR141617A) profiles were observed in HEK-hCB2-Gqo5 and HEK-hCB2-Gqi5 cells as determined using intracellular Ca2+ measurements. Experiments with HEK-hCB2-Gqi5 cells carried out by investigating interactions among CP55,940, carbachol, thapsigargin, and U73122 revealed that the mechanism of cannabinoid hCB2 receptor coupling via chimeric G proteins to Ca2+ mobilization involves phospholipase C-inositol trisphosphate (PLC-IP3) and that it is less efficient in comparison to the endogenous muscarinic mediated PLC-IP3-Ca2+ pathway. This study demonstrates that expressed cannabinoid CB2 receptors couple equally well to Gαqo5 and Gαqi5 proteins and that receptor agonist or antagonist pharmacology is not influenced by the nature of these coupled G proteins when heterologously expressed.