Pharmacological Comparison of a Recombinant CB1 Cannabinoid Receptor with Its Ga16 Fusion Product

Abstract

The pharmacology of G protein-coupled receptors is widely accepted to depend on the G protein subunit to which the agonist-stimulated receptor couples. In order to investigate whether CB₁ agonist-mediated signal transduction via an engineered Ga_α16 system is different than that of the G_i/o coupling normally preferred by the CB₁ receptor, we transfected the human recombinant CB₁ receptor (hCB₁) or a fusion protein comprising the hCB₁ receptor and G_α16 (hCB₁-G_α16) into HEK293 cells. From competition binding studies, the rank order of ligand affinities at the hCB₁-Ga_α16 fusion protein was found to be similar to that for hCB₁: HU 210 > CP 55,940 ≥ SR 141716A > WIN 55212-2 > anandamide > JWH 015. Agonists increased [³⁵S]GTPγS binding or inhibited forskolin-stimulated cAMP, presumably by coupling to G_i/o, in cells expressing hCB₁ but not hCB₁-G_α16. However, an analogous rank order of potencies was observed for these agonists in their ability to evoke increases in intracellular calcium concentration in cells expressing hCB_q-Ga_α16 but not hCB₁. These data demonstrate that ligand affinities for the hCB₁, receptor are not affected by fusion to the G_α16 subunit. Furthermore, there is essentially no difference in the function of the hCB₁, receptor when coupled to G_i/o, or G_α16 .