Differential effects of voltage-dependent inactivation and local anesthetics on kinetic phases of Ca2+ release in frog skeletal muscle

In voltage-clamped frog skeletal muscle fibers, Ca2+ release rises rapidly to a peak, then decays to a nearly steady state. The voltage dependence of the ratio of amplitudes of these two phases (p/s) shows a maximum at low voltages and declines with further depolarization. The peak phase has been attributed to a component of Ca2+ release induced by Ca2+, which is proportionally greater at low voltages. We compared the effects of two interventions that inhibit Ca2+ release: inactivation of voltage sensors, and local anesthetics reputed to block Ca2+ release induced by Ca2+. Holding the cells partially depolarized strongly reduced the peak and steady levels of Ca2+ release elicited by a test pulse and suppressed the maximum of the p/s ratio at low voltages. The p/s ratio increased monotonically with test voltage, eventually reaching a value similar to the maximum found in noninactivated fibers. This implies that the marked peak of Ca2+ release is a property of a cooperating collection of voltage sensors rather than individual ones. Local anesthetics reduced the peak of release flux at every test voltage, and the steady phase to a lesser degree. At variance with sustained depolarization, they made p/s low at all voltages. These observations were well-reproduced by the couplon model of dual control, which assumes that depolarization and anesthetics respectively, and selectively, disable its Ca2+-dependent or its voltage-operated channels. This duality of effects and their simulation under such hypotheses are consistent with the operation of a dual, two-stage control of Ca2+ release in muscle, whereby Ca2+ released through multiple directly voltage-activated channels builds up at junctions to secondarily open Ca2+-operated channels.