We used single electrode voltage-clamp methods to investigate the inactivation of Ca2+ currents in melanotrophs of the intermediate lobe of the pituitary. The low threshold transient current was inactivated by brief prepulses to potentials above -30 mV and inhibition remained complete as prepulse potential was increased from 0 to +70 mV. Both the high threshold transient and sustained currents, however, were inhibited to the greatest extent (60%) by prepulses to 0 mV. Prepulses to more positive potentials close to the Ca2+ reversal potential produced much less (15%) inactivation. Buffering intracellular Ca2+ by including BAPTA in the recording electrode or replacing extracellular Ca2+ with Ba2+ reduced the effect of prepulses. Slowing Ca2+ extrusion by reducing the Na+ gradient across the cell increased the duration of the effect of prepulses. We conclude that the low threshold, transient current is inactivated primarily by membrane voltage while both the high threshold currents are inhibited by elevation of intracellular Ca2+ although the two currents display different sensitivities to Ca2+ concentration. Inhibition of the high threshold transient current by the neurotransmitter dopamine, however, acts by a different mechanism not mediated by Ca(2+)-dependent inactivation.