Time-resolved synchrotron X-ray diffraction studies of a single frog skeletal muscle fiber. Time courses of intensity changes of the equatorial reflections and intracellular Ca2+ transients.

Time-resolved X-ray equatorial diffraction studies on a single frog skeletal muscle fiber were performed with a 10 ms time resolution using synchrotron radiation in order to compare the time courses of the molecular changes of contractile proteins and the intracellular Ca2+ transient during an isometric twitch contraction at 2.7 degrees C. Measurements of the Ca2+ transient using aequorin as an intracellular Ca2+ indicator were conducted separately just before and after the X-ray experiments under very similar experimental conditions. The results, which showed a similar time course of tension to that observed in the X-ray experiment, were compared with the aequorin light signal, tension and the intensity changes of the 1,0 and 1,1 equatorial reflections. No appreciable change in both reflection spacings indicated that the effect of internal shortening of the muscle was minimized during contraction. The intensity change of the equatorial reflections generally occurred after the aequorin light signal. In the rising phase, the time course of increase in the 1,1 intensity paralleled that of the rise of the light signal and the intensity peak occurred 20-30 ms after the peak of the light signal. The decrease in the 1,0 intensity showed a time course similar to that of tension and the intensity minimum roughly coincided with the tension peak, coming at 80-90 ms and about 60 ms after the peaks of the light signal and the 1,1 intensity change, respectively. In the relaxation phase, the 1,1 intensity seemed to fall rapidly just before the tension peak and then returned to the original level in parallel with the decay of tension. The 1,0 intensity returned more slowly than the tension relaxation. Thus, the change of the 1,1 intensity was faster than that of the 1,0 intensity in both the rising and relaxation phases. When the measured aequorin light signal was corrected for the kinetic delay of the aequorin reaction with a first-order rate constant of either 50 or 17 s-1, the peak of the corrected light signal preceded that of the measured one by approx. 30 ms. Thus, the peak of the Ca2+ transient appeared earlier than the peaks of the 1,1 and 1,0 intensity changes by 50-60 and 110-120 ms, respectively. The time lag between the extent of structural change and the Ca2+ transient is discussed in relation to the double-headed attachment of a cross-bridge to actin.