VOLTAGE-GATED TRANSIENT CURRENTS IN BOVINE ADRENAL FASCICULATA CELLS .1. T-TYPE CA2+ CURRENT

The whole cell version of the patch clamp technique was used to identi fy and characterize voltage-gated Ca2+ channels in enzymatically disso ciated bovine adrenal zona fasciculata (AZF) cells. The great majority of cells (84 of 86) expressed only low voltage-activated, rapidly ina ctivating Ca2+ current with properties of T-type Ca2+ current describe d in other cells. Voltage-dependent activation of this current was fit by a Boltzmann function raised to an integer power of 4 with a midpoi nt at -17 mV. Independent estimates of the single channel gating charg e obtained from the activation curve and using the ''limiting logarith mic potential sensitivity'' were 8.1 and 6.8 elementary charges, respe ctively. Inactivation was a steep function of voltage with a v1/2 of - 49.9 mV and a slope factor K of 3.73 mV. The expression of a single Ca 2+ channel subtype by AZF cells allowed the voltage-dependent gating a nd kinetic properties of T current to be studied over a wide range of potentials. Analysis of the gating kinetics of this Ca2+ current indic ate that T channel activation, inactivation, deactivation (closing), a nd reactivation (recovery from inactivation) each include voltage-inde pendent transitions that become rate limiting at extreme voltages. Ca2 + current activated with voltage-dependent sigmoidal kinetics that wer e described by an m4 model. The activation time constant varied expone ntially at test potentials between -30 and +10 mV, approaching a volta ge-independent minimum of 1.6 ms. The inactivation time constant (tau( i)) also decreased exponentially to a minimum of 18.3 ms at potentials positive to 0 mV. T channel closing (deactivation) was faster at more negative voltages; the deactivation time constant (tau(d)) decreased from 8.14 +/- 0.7 to 0.48 +/- 0.1 ms at potentials between -40 and -15 0 mV. T channels inactivated by depolarization returned to the closed state along pathways that included two voltage-dependent time constant s. tau(rec-s), ranged from 8.11 to 4.80 s when the recovery potential was varied from -50 to -90 mV, while T(rec-f) decreased from 1.01 to 0 .372 s. At potentials negative to -70 mV, both time constants approach ed minimum values. The low voltage-activated Ca2+ current in AZF cells was blocked by the T channel selective antagonist Ni2+ with an IC50 o f 20 muM. At similar concentrations, Ni2+ also blocked cortisol secret ion stimulated by adrenocorticotropic hormone. Our results indicate th at bovine AZF cells are distinctive among secretory cells in expressin g primarily or exclusively T-type Ca2+ channels. These channels serve as the major pathway for voltage-gated Ca2+ entry and may mediate pept ide hormone-stimulated cortisol secretion in these cells.