A DAMPED OSCILLATION IN THE INTRAMEMBRANOUS CHARGE MOVEMENT AND CALCIUM-RELEASE FLUX OF FROG SKELETAL-MUSCLE FIBERS

Asymmetric membrane currents and calcium transients were recorded simu ltaneously from cut segments of frog skeletal muscle fibers voltage cl amped in a double Vaseline-gap chamber in the presence of high concent ration of EGTA intracellularly. An inward phase of asymmetric currents following the hump component was observed in all fibers during the de polarization pulse to selected voltages (similar or equal to -45 mV). The average value of the peak inward current was 0.1 A/F (SEM = 0.01, n = 18), and the time at which it occurred was 34 ms (SEM = 1.8, n = 1 8). A second delayed outward phase of asymmetric current was observed after the inward phase, in those experiments in which hump component a nd inward phase were large. It peaked at more variable time (between 6 0 and 130 ms) with amplitude 0.02 A/F (SEM = 0.003, n = 11). The trans membrane voltage during a pulse, measured with a glass microelectrode, reached its steady value in less than 10 ms and showed no oscillation s. The potential was steady at the time when the delayed component of asymmetric current occurred. ON and OFF charge transfers were equal fo r all pulse durations. The inward phase moved 1.4 nc/mu F charge (SEM = 0.8, n = 6), or about one third of the final value of charge mobiliz ed by these small pulses, and the second outward phase moved 0.7 nC/mu F (SEM = 0.8, n = 6), bringing back about half of the charge moved du ring the inward phase. When repolarization intersected the peak of the inward phase, the OFF charge transfer was independent of the repolari zation voltage in the range -60 to -90 mV. When bath pre- and post-pul se voltages were changed between -120 mV and -60 mV, the equality of O N and OFF transfers of charge persisted, although they changed from 11 3 to 81% of their value at -90 mV. The three delayed phases in asymmet ric current were also observed in experiments in which the extracellul ar solution contained Cd2+, La3+ and no Ca2+. Large increases in intra cellular [Cl-] were imposed, and had no major effect on the delayed co mponents of the asymmetric current. The Ca2+ transients measured optic ally and the calculated Ca2+ release fluxes had three phases whenever a visible outward phase followed the inward phase in the asymmetric cu rrent. Several interventions intended to interfere with Ca release, re duced or eliminated the three delayed phases of the asymmetric current . We conclude that these phases are capacitive, with no significant io nic component. All results could be well described by a model of EC co upling in which the oscillations of the asymmetric current are intrame mbranous charge movements, driven by changes in local voltage. In the model, the oscillations are the consequence of two feedback processes. One is positive: Ca2+, released from the SR binds to a hypothetical s ite near the voltage sensors, increasing the local voltage. The other is the spontaneous inactivation of release, presumably also induced by Ca2+, which reduces the local Ca2+ concentration and the transmembran e voltage.