CB1 cannabinoid receptor-mediated cell migration

Recent studies have suggested that cell migratory responses are often media ted by G(i) protein-coupled receptors. Because it is known that CB1 cannabi noid receptors are coupled to pertussis toxin-sensitive G proteins, we prop osed that CB1 may mediate cell migration. To test this hypothesis, modified Boyden chamber assays were used to investigate cell migration mediated by CB1 cannabinoid receptors. HU-210, WIN55212-2, and anandamide, three cannab inoid agonists with distinct chemical structures, induced migration of huma n embryonic kidney 293 cells stably transfected with human CB1 gene, but no t 293 cells transfected with an empty expression vector. These migratory re sponses were concentration-dependent. The EC50 values for HU-210, WIN55212- 2, and anandamide were 0.19 +/- 0.04, 12.2 +/- 1.4, and 39.9 +/- 3.7 nM, re spectively. The maximal migration index for HU-210, WIN55212-2, and anandam ide were 8.9 +/- 1.6, 9.5 +/- 1.6, and 8.8 +/- 1.3, respectively. Pretreati ng cells with 100 ng/ml pertussis toxin eliminated the cannabinoid agonist- induced cell migration. SR141716A, a selective antagonist for CB1, inhibite d the cannabinoid agonist-induced migratory responses in a concentration-de pendent manner. Checkerboard analysis demonstrated that anandamide-induced cell migrations are due to chemotaxis as well as chemokinesis. Furthermore, anandamide- induced migratory responses were inhibited, in a concentration - dependent manner, by PD098059, an inhibitor of mitogen- activated protein kinase activation, but not by 8-bromoadenosine-3',5'-cyclic monophosphate, a cell-permeable cAMP analog. These data demonstrate that cannabinoid agon ists are able to induce chemotaxis and chemokinesis, and that these migrato ry responses are mediated by G protein-coupled, CB1 cannabinoid receptors. In addition, these data suggest that activation of mitogen- activated prote in kinase plays an important role, whereas inhibition of adenylate cyclase is probably not involved in the cell migration mediated by CB1.