Activating efficiency of Ca2+ and cross-bridges as measured by phosphate analog release

To assess the activating efficiency of Ca2+ and cross-bridges, the release rates of phosphate analogs from skinned fibers were estimated from the reco very of contractility and that of stiffness. Estimations were performed bas ed on the assumptions that contractility was indicative of the population o f analog-free myosin heads and that stiffness reflected the population of f ormed cross-bridges. Aluminofluoride (AIFx) and orthovanadate (Vi) were use d as phosphate analogs with mechanically skinned fibers from rabbit psoas m uscle. The use of the analogs enabled the functional assessment of activati on level in the total absence of ATP. Fibers loaded with the analogs gradua lly recovered contractility and stiffness in normal plain rigor solution. T he addition of Ca2+ to the plain rigor solution significantly accelerated t heir recovery, whereas ADP had no appreciable effect. ATP plus Ca2+ (contra cting condition) accelerated the recovery by several tens of times. These r esults indicate that the cross-bridges formed during contraction have promi nent activating efficiency, which is indispensable to attain full activatio n. A comparison between the activating efficiency evaluated from stiffness and that from contractility suggested that Ca2+ is more potent in accelerat ing the binding of actin to analog-bound myosin heads whereas crossbridges mainly accelerate the subsequent analog-releasing step.