Effect of the calcium buffer EGTA on the "hump" component of charge movement in skeletal muscle.

Abstract

Three manifestations of excitation-contraction (E-C) coupling were measured in cut skeletal muscle fibers of the frog, voltage clamped in a double Vaseline gap: intramembrane charge movements, myoplasmic Ca2+ transients, and changes in optical transparency. Pulsing patterns in the presence of high [EGTA] intracellularly, shown by García et al. (1989. J. Gen. Physiol. 94:973-986) to deplete Ca2+ in the sarcoplasmic reticulum, were found to change the above manifestations. With an intracellular solution containing 15 mM EGTA and 0 Ca, 10-15 pulses (100 ms) to -20 mV at a frequency of 2 min-1 reduced the "hump" component of charge movement current. This effect was reversible by 5 min of rest. The same effect was obtained in 62.5 mM EGTA and 0 Ca by pulsing at 0.2 min-1. This effect was reversible by adding calcium to the EGTA solution, for a nominal [Ca2+]i of 200 nM, and was prevented by adding calcium to the EGTA solution before pulsing. The suppression of the hump was accompanied by elimination of the optical manifestations of E-C coupling. The current suppressed was found by subtraction and had the following properties: delayed onset, a peak at a variable interval (10-20 ms) into the pulse, a negative phase (inward current) after the peak, and a variable OFF transient that could be multi-phasic and carried less charge than the ON transient. In the previous paper (Csernoch et al., 1991. J. Gen. Physiol. 97:845-884) it was shown that several interventions suppress a similar component of charge movement current, identified with the "hump" or Q gamma current (I gamma). Based on the similarity to that component, the charge movement suppressed by the depletion protocols can also be identified with I gamma. The fact that I gamma is suppressed by Ca2+ depletion and the kinetic properties of the charge suppressed is inconsistent with the existence of separate sets of voltage sensors underlying the two components of charge movement, Q beta and Q gamma. This is explicable if Q gamma is a consequence of calcium release from the sarcoplasmic reticulum.