CHARACTERIZATION OF THE CALCIUM-SENSITIVE VOLTAGE-GATED DELAYED RECTIFIER POTASSIUM CHANNEL IN ISOLATED GUINEA-PIG HEPATOCYTES

The voltage-dependent K+ channel was examined in enzymatically isolate d guinea pig hepatocytes using whole-cell, excised outside-out and ins ide-out configurations of the patch-clamp technique. The resting membr ane potential in isolated hepatocytes was -25.3 +/- 4.9 mV (n = 40). U nder the whole-cell voltage-clamp, the time-dependent delayed rectifie r outward current was observed at membrane potentials positive to -20 mV at physiological temperature (37 degrees C). The reversal potential of the current, as determined from tail current measurements, shifted by similar to 57 mV per 10-fold change in the external K+ concentrati on. In addition, the current did not appear when K+ was replaced with Cs+ in the internal and external solutions, indicating that the curren t was carried by K+ ions. The envelope test of the tails demonstrated that the growth of the tail current followed that of the current activ ation. The ratio between the activated current and the tail amplitude was constant during the depolarizing step. The time course of growth a nd deactivation of the tail current were best described by a double ex ponential function. The current was suppressed in Ca2+-free, 5 mM EGTA internal or external solution (pCa > 9). The activation curve (P-infi nity curve) was not shifted by changing the internal Ca2+ concentratio n ([Ca2+](i)). The current was inhibited by bath application of 4-amin opyridine or apamin. alpha(1)-Adrenergic stimulation with noradrenalin e enhanced the current but beta-adrenergic stimulation with isoprotere nol had no effect on the current. In single-channel recordings from ou tside-out patches, unitary current activity was observed by depolarizi ng voltage-clamp steps whose slope conductance was 9.5 +/- 2.2 pS (n = 10). The open time distribution was best described by a single expone ntial function with the mean open lifetime of 18.5 +/- 2.6 ms (n = 14) , while at least two exponentials were required to fit the closed time distributions with a time constant. for the fast component of 2.0 +/- 0.3 ms (n = 14) and that for the slow component of 47.7 +/-5.9 ms (n = 14). Ensemble averaged current exhibited delayed rectifier nature wh ich was consistent with whole-cell measurements. In excised inside-out patch recordings, channel open probability was sensitive to [Ca2+](i) . The concentration of Ca2+ at the half-maximal activation was 0.031 m u M These results suggest that guinea pig hepatocytes possess voltage- gated delayed rectifier K+ channels which are modified by intracellula r Ca2+.