Cannabinoid CB1 receptor-mediated inhibition of prolactin release and signaling mechanisms in GH4C1 cells

The GH(4)C(1) cell line was used to study the cellular mechanisms of cannabinoid-mediated inhibition of PRL release. Cannabinoid CB1 receptor activation inhibited vasoactive intestinal polypeptide- and TRH-stimulated PRL release, but not its basal secretion. The cannabinoid-mediated inhibition of TRH-stimulated PRL release was reversed by the CB1 receptor-specific antagonist, SR141,716A, and was abolished by pertussis toxin pretreatment, indicating that G(alpha) subunits belonging to the G(i)alpha and G(o)alpha family were involved in the signaling. Photoaffinity labeling using [alpha-P-32]azidoaniline GTP showed that cannabinoid receptor stimulation in cell membranes produced activation of four G(alpha) subunits (G(i)alpha 2, G(i)alpha 3, G(o)alpha 1, and G(o)alpha 2), which was also reversed by SR141,716A. The CB1 receptor agonists, WIN55,212-2 and CP55,940, inhibited cAMP formation and calcium currents in GH(4)C(1) cells. The subtypes of calcium currents inhibited by WIN55,212-2 were characterized using holding potential sensitivity and calcium channel blockers. WIN55,212-2 inhibited the omega-conotoxin GVIA (Conus geographus)- and omega-agatoxin IVA (Aigelenopsis aperta)-sensitive calcium currents, but not the nisoldipine-sensitive calcium currents, suggesting the inhibition of N- and P-type, but not L-type, calcium currents. Taken together, the present findings indicate that CB1 receptors can couple through pertussis toxin-sensitive G alpha subunits to inhibit adenylyl cyclase and calcium currents and suppress PRL release from GH(4)C(1) cells.