Despite substantial degradation, 2-arachidonoylglycerol is a potent full efficacy agonist mediating CB1 receptor-dependent G-protein activation in rat cerebellar membranes

1 Two endocannabinoids, arachidonoyl ethanolamide (AEA) and 2-arachidonoylg lycerol (2-AG) bind and activate G-protein-coupled cannabinoid receptors, b ut limited data exist on their relative ability to activate G-proteins. 2 Here we assess agonist potency and efficacy of various cannabinoids, incl uding 2-AG, HU-310 (2-arachidonoyl glyceryl ether, a third putative endocan nabinoid), HU-313 (another ether analogue of 2-AG), AEA, R-methanandamide ( an enzymatically stable analogue of AEA), and CP-55,940 at rat brain CB1 re ceptors using agonist-stimulated [S-35]-GTP gammaS binding to cerebellar me mbranes and whole brain sections. Degradation of endocannabinoids under exp erimental conditions was monitored by HPLC. 3 To enhance efficacy differences, agonist dose-response curves were genera ted using increasing GDP concentrations. At 10(-6) M GDP, all compounds, ex cept HU-313, produced full agonists responses similar to2.5 fold over basal . The superior efficacy of 2-AG over all other compounds became evident by increasing GDP (10(-5) and 10(-4) M). 4 In membrane incubations, 2-AG was degraded by 85% whereas AEA and HU-310 were stable. Pretreatment of membranes with phenylmethylsulphonyl fluoride inhibited 2-AG degradation, resulting in 2 fold increase in agonist potency . Such pretreatment had no effect on AEA potency. 5 Responses in brain sections were otherwise consistent with membrane bindi ng data, but 2-AG evoked only a weak signal in brain sections, apparently d ue to more extensive degradation. 6 These data establish that even under conditions of substantial degradatio n, 2-AG is a full efficacy agonist, clearly more potent than AEA, in mediat ing CB1 receptor-dependent G-protein activity in native membranes.