Evidence for a new G protein-coupled cannabinoid receptor in mouse brain

The purpose of these studies was to support the hypothesis that an undiscov ered cannabinoid receptor exists in brain. [S-35]GTP gammaS binding was sti mulated by anandamide and WIN55212-2 in brain membranes from both CB1+/+ an d CB1-/- mice. In contrast, a wide variety of other compounds that are know n to activate CB1 receptors, including CP55940, HU-210, and Delta (9)-tetra hydrocannabinol, failed to stimulate [S-35]GTP gammaS binding in CB1-/- mem branes. In CB1-/- membranes, SR141716A affected both basal and anandamide- or WIN55212-2-induced stimulation of [S-35]GTP gammaS binding only at conce ntrations greater than 1 muM. In CB1+/+ membranes, SR141716A inhibited only 84% of anandamide and 67% of WIN55212-2 stimulated [S-35]GTP gammaS bindin g with an affinity appropriate for mediation by CB1 receptors (K-B approxim ate to 0.5 nM). The remaining stimulation seemed to be inhibited with lower potency (IC50 approximate to 5 muM) similar to that seen in CB1-/- membran es or in the absence of agonist. Further experiments determined that the ef fects of anandamide and WIN55212-2 were not additive, but that the effect o f mu opioid, adenosine A1, and cannabinoid ligands were additive. Finally, assays of different central nervous system (CNS) regions demonstrated signi ficant activity of cannabinoids in CB1-/- membranes from brain stem, cortex , hippocampus, diencephalon, midbrain, and spinal cord, but not basal gangl ia or cerebellum. Moreover, some of these same CNS regions also showed sign ificant binding of [H-3]WIN55212-2, but not [H-3]CP55940. Thus anandamide a nd WIN55212-2 seemed to be active in CB1-/- mouse brain membranes via a com mon G protein-coupled receptor with a distinct CNS distribution, implying t he existence of an unknown cannabinoid receptor subtype in brain.