Inhibitory effects of SR141716A on G-protein activation in rat brain

N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR141716A), a cannabinoid CB1 receptor antagonist, has inverse agonist effects in cannabinoid CB1 receptor-expressing cell lines, brain and peripheral organs. These studies characterized SR141716A-inhibited G-protein activity by measuring [S-35]GTP gammaS binding. Maximal inhibition of basal [S-35]GTP gammaS binding in cerebellar membranes was 50%. The EC50 value for inhibition of [S-35]GTP gammaS binding was 4.4 muM, whereas the K-c for inhibition of R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate (WIN 55,212-2)-stimulated [S-35]GTP gammaS binding was 0.6 nM. [S-35]GTP gammaS autoradiography was used to examine the regional specificity of SR141716A inhibition. SR141716A inhibited basal [S-35]GTP gammaS binding in all regions examined, with inhibition ranging from approximately 20% in caudate-putamen to 40% in hippocampus. These studies demonstrate that SR141716A is a competitive antagonist at nanomolar concentrations, whereas it inhibits basal receptor-mediated G-protein activity at micromolar concentrations. These data suggest that the apparent inverse agonist effect is either not cannabinoid CB1 receptor-specific or that SR141716A is binding to different sites on the cannabinoid CB1 receptor to produce inverse agonist versus competitive antagonist effects. (C) 2001 Elsevier Science B.V. All rights reserved.