The activity‐induced reduction of myofibrillar Ca 2+ sensitivity in mouse skeletal muscle is reversed by dithiothreitol

The aim of this study was to further characterize the reduction of myofibrillar Ca 2+ sensitivity in mouse muscle which has been observed after fatigue at 37°C. Muscle bundles and single fibres were isolated from mouse flexor digitorum brevis muscle and studied at 37°C. The single fibres were injected with the Ca 2+ indicator indo‐1. Muscle fatigue was produced by 0.4 s tetani repeated at 4 s intervals until force had fallen to less than 50% of initial. Excitation–contraction coupling was assessed by measuring the cytosolic calcium concentration ([Ca 2+ ] i ) during tetani, and the maximum Ca 2+ ‐activated force and the myofibrillar Ca 2+ sensitivity were estimated from a series of tetani at different stimulation frequencies. Two main results were found. (i) The reduction of Ca 2+ sensitivity only occurred when the muscle was intensely stimulated leading to fatigue. When the muscle was rested for 10 min at 37°C there was no significant change in Ca 2+ sensitivity. (ii) If the membrane‐permeant thiol‐specific reducing agent dithiothreitol (0.5 m m ) was applied to the muscle for 2 min following the fatigue protocol, the reduction in Ca 2+ sensitivity was reversed. Dithiothreitol had no effect on Ca 2+ sensitivity in unfatigued preparations. There was no effect of fatigue or dithiothreitol on tetanic [Ca 2+ ] i or on the maximum Ca 2+ ‐activated force. These results suggest that intense activity of skeletal muscle at 37°C causes the production of reactive oxygen species which oxidize a target protein. We propose that critical sulphydryl groups on the target protein(s) are converted to disulphide bonds and this reaction reduces Ca 2+ sensitivity.