Biochemical separation of delayed rectifier currents in frog short skeletal muscle fibres

Frog short (1-1.5 mm) skeletal muscle fibres in sucrose hypertonic Ringer at 5 degrees C were voltage clamped employing a two-electrode technique. Decreasing pH from 7.0 to 5.0 dramatically increased the rate of turn on of the slow delayed rectifier (GK,s) current, so that it became similar to the rapidly activated form. The accelerating effects of decreased pH upon slow current kinetics had a pKa of 5.8. Decreased pH also appeared to shift the voltage dependence of fast and GK,s gating to more positive potentials. In addition, a decrease in pH from 7.0 to 5.0 shifted the reversal potential of GK,s by over 11 mV in the positive direction. GK,s was selectively and irreversibly abolished by applying 1 mM-diethylpyrocarbonate (DEP), a histidine reagent. This effect occurred even after GK,s had been accelerated by low pH. DEP had no effect upon fast delayed rectifier current except for a small positive shift in voltage dependence. Application of 1 or 2 mM-N-ethylmaleimide, a sulphydryl reagent, depressed the fast delayed rectifier while sparing the GK,s currents. However, the muscle fibres also developed markedly increased leakage currents.