Using whole-cell and perforated-patch recordings, we have examined the part played by endogenous G-protein βγ subunits in neurotransmitter-mediated inhibition of N-type Ca2+ channel current ICa) in dissociated rat superior cervical sympathetic neurones.Expression of the C-terminus domain of β-adrenergic receptor kinase 1 (βARK1), which contains the consensus motif (QXXER) for binding Gβγ, reduced the fast (pertussis toxin (PTX)-sensitive) and voltage-dependent inhibition of ICa by noradrenaline and somatostatin, but not the slow (PTX-insensitive) and voltage-independent inhibition induced by angiotensin II. βARK1 peptide reduced GTP-γ-S-induced voltage-dependent and PTX-sensitive inhibition of ICa but not GTP-γ-S-mediated voltage-independent inhibition.Overexpression of Gβ1γ2, which mimicked the voltage-dependent inhibition by reducing ICa density and enhancing basal facilitation, occluded the voltage-dependent noradrenaline- and somatostatin-mediated inhibitions but not the inhibition mediated by angiotensin II.Co-expression of the C-terminus of βARK1 with β1 and γ2 subunits prevented the effects of Gβγ dimers on basal Ca2+ channel behaviour in a manner consistent with the sequestering of Gβγ.The expression of the C-terminus of βARK1 slowed down reinhibition kinetics of ICa following conditioning depolarizations and induced long-lasting facilitation by cumulatively sequestering βγ subunits.Our findings identify endogenous Gβγ as the mediator of the voltage-dependent, PTX-sensitive inhibition of ICa induced by both noradrenaline and somatostatin but not the voltage-independent, PTX-insensitive inhibition by angiotensin II. They also support the view that voltage-dependent inhibition results from a direct Gβγ-Ca2+ channel interaction.