Characteristics of voltage gated Ca2+ currents in ovine gonadotrophs

Abstract

1. Voltage-clamp recordings were obtained from gonadotrophs of the ovine pars tuberalis in dissociated cell culture, utilizing the whole-cell recording mode of the patch-clamp technique. 2. The amplitudes of Ca2+ and Ba2+ currents were dependent on the extracellular concentration of divalent cation. 3. Ba2+ tail currents were observed on termination of depolarizing voltage steps. The extrapolated amplitudes of 'instantaneous' tail currents increased with membrane depolarization and showed saturation beyond +15 mV. 4. True inactivation of currents occurred in the presence of both external Ca2+ and Ba2+, judged from decrease in tail current amplitudes with progressive increases in duration of the activating voltage pulse. The inactivation process was fitted by a single-exponential function at membrane potentials below -25 mV, while at more depolarized potentials the inactivation was better described by a double-exponential function. The inactivation time constants decreased with positive shifts in membrane potential favouring a voltage-dependent inactivation. 5. The half-value of steady-state inactivation was observed at -40 mV using a two-pulse protocol. 6. Power spectral analysis of Ba2+ current noise from the steady-state portion of inward current showed a double Lorentzian fit of the power spectrum. 7. Two types of voltage-activated Ca2+ currents were identified based on their kinetics, voltage dependence, dependence on activation frequency, differential sensitivity to intracellular ATP and cyclic AMP, and to extracellular application of nifedipine. The channels with faster kinetics had a lower activation threshold (-50 mV) and the amplitude of the current was sensitive to clamping frequency. 8. From ensemble noise analysis of mean maximal inward current, single-channel amplitude of about 1 pA was estimated in 50 mM-Ba2+.