Characterization and cyclic AMP-dependence of a hyperpolarization-activated chloride conductance in Leydig cells from mature rat testis.
We recently described a cyclic AMP-activated current in the membrane of Leydig cells from mature rat testis by using the whole-cell configuration of the patch-clamp technique (Noulin & Joffre, 1992a). In the present study, further experiments were performed in symmetrical CsCl solutions. We show that this current corresponds to a hyperpolarization-activated chloride conductance. Voltage jumps to negative potentials, applied from a holding potential of +60 mV, activated a time-dependent inward current. In control cells, the curve of steady-state current activation typically ranged from +60 mV (0) to -120 mV (1) and had its midpoint at -40 mV. Deactivation at positive potential was characterized by an instantaneous outwardly rectifying current which decayed with time. The kinetics of activation and deactivation were described by a double and a single exponential, respectively. Cyclic AMP, added to the pipette solution, increased both the inward rectification and the amplitude of the steady-state current in the range of 0 to -60 mV. The activation threshold was unchanged, while the V0.5 of the activation curve was shifted by 24 mV to more positive potentials. Consequently, the activation curve was steeper. The two rate constants of activation were increased and were strongly voltage dependent. In parallel, the amplitude of the instantaneous outward current and the rate constant of deactivation were increased. The reversal potential of this current was close to ECl. It did not change with equimolar replacement of cesium by TEA, and shifted with the chloride concentration gradient. This current was inhibited by chloride channel blockers. These results indicate a hyperpolarization-activated chloride conductance in the membrane of Leydig cells which is modulated by cyclic AMP. This nucleotide acts by modifying the kinetics of inward current and both the kinetics and the amplitude of deactivating outward current.