Modulation of CB1 cannabinoid receptor functions after a long-term exposure to agonist or inverse agonist in the Chinese hamster ovary cell expression system

We have investigated the adaptive changes of the human central cannabinoid receptor (CB1) stably expressed in Chinese hamster ovary cells (CHO-CB1), a fter agonist (CP 55,940) or selective CB1 inverse agonist (SR 141716) treat ment. CB1 receptor density and affinity constant as measured by binding ass ays with both tritiated ligands remained essentially unchanged after varyin g period exposure of CHO-CB1 cells (from 30 min to 72 hr) to saturating con centrations of CP 55,940 or SR 141716. However, using a C-myc-tagged versio n of the CB1 receptor, FAGS analysis and confocal microscopy studies on CBI expression indicated that the agonist promoted a disappearance of cell sur face receptor although inverse agonist increased its cell surface density. Taken together these results suggest that 1) agonist induces internalizatio n of the receptor into a cellular compartment that would be still accessibl e to both the hydrophobic ligands CP 55,940 or SR 141716; 2) inverse-agonis t promotes externalization of the receptor from an intracellular preexistin g pool to the cell surface. In parallel, we also investigated the associate d effects of CP 55,940 and SR 141716 on CB1 receptor-coupled second messeng ers. We showed that preexposure of cells to CP 55,940 induced a rapid desen sitization of the CB1 to the agonist response. The ability of CP 55,940 to inhibit the forskolin-stimulated adenylyl cyclase and to activate the mitog en-activated protein kinase activity was dramatically reduced. By striking contrast, SR 141716 pretreatment of CHO-CB1 cells not only had no significa nt effect on the potency of CP 55,940 to inhibit the forskolin-stimulated a denylyl cyclase but also induced a significant enhancement of the CP 55,940 ability to stimulate the mitogen-activated protein kinase activity. These results suggest that the modulation of the number of cell surface receptor could lead to functional desensitization or sensitization of the CB1 recept ors.