The delayed component of intramembranous charge movement (hump, I gamma) was studied around the contraction threshold in cut skeletal muscle fibers of the frog (Rana esculenta) in a single Vaseline-gap voltage clamp. Charges (Q) were computed as 50-ms integrals of the ON (QON) and OFF (QOFF) of the asymmetric currents after subtracting a baseline. The hump appeared in parallel with an excess of QON over QOFF by approximately 2.5 nC/mu F. Caffeine (0.75 mM) not only shifted the contraction threshold but moved both the hump and the difference between the ON and OFF charges to more negative membrane potentials. When using 10-mV voltage steps on top of different prepulse levels, the delayed component, if present, was more readily observable. The voltage dependences of the ON and OFF charges measured with these pulses were clearly different: QON had a maximum at or slightly above the contraction threshold, while QOFF increased monotonically in the voltage range examined. Caffeine (0.75 mM) shifted this voltage dependence of QON toward more negative membrane potentials, while that of QOFF was hardly influenced. These results show that the delayed component of intramembranous charge movement either is much slower during the OFF than during the ON, or returns to the OFF position during the pulse. Tetracaine (25 microM) had similar effects on the charge movement currents, shifting the voltage dependence on the ON charge in parallel with the contraction threshold, but to more positive membrane potentials, and leaving QOFF essentially unchanged. The direct difference between the charge movement measured in the presence of caffeine and in control solution was either biphasic or resembled the component isolated by tetracaine, suggesting a common site of caffeine and tetracaine action. The results can be understood if the released Ca plays a direct role in the generation of the hump, as proposed in the first paper of this series (Csernoch et al. 1991. J. Gen. Physiol. 97:845-884).

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