Voltage-dependent Ca2+ currents appear to be involved in the actions of hormones that regulate pituitary secretion. In order to investigate modulation of Ca2+ currents by release-inducing and release-inhibiting hormones, we performed whole-cell clamp experiments in the pituitary cell line GH3. The resting potential was approximately -40 mV; spontaneous action potentials were observed in the majority of cells. Superfusion of cells with the stimulatory hormone, LHRH, depolarized the plasma membrane to approximately -10 mV, whereas the inhibitory hormone, somatostatin, caused hyperpolarization to approximately -60 mV; both hormones suppressed spontaneous action potentials. Under voltage clamp conditions, GH3 cells exhibited slowly and fast inactivating Ca2+ currents. LHRH increased whereas somatostatin decreased the slowly inactivating currents; fast inactivating currents were not affected by these hormones. The stimulatory effect of LHRH was not mimicked by intracellularly applied cAMP. In contrast to vasoactive intestinal peptide and forskolin, LHRH did not activate adenylate cyclase in membranes of GH3 cells, but rather appeared to cause inhibition of the enzyme. Hormonal stimulation and inhibition of inward currents were abolished by pretreatment of the cells with pertussis toxin. In membranes of GH3 cells, we identified a pertussis toxin-sensitive G-protein of the Gi-type and Go. We conclude that LHRH and somatostatin modulate voltage-dependent Ca2+ currents via cAMP-independent mechanisms involving pertussis toxin-sensitive G-proteins. The occurrence of both pertussis toxin-sensitive hormonal stimulation and inhibition of voltage-dependent Ca2+ currents in one cell type suggest that these opposite regulations are mediated by distinct G-proteins.