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In this study we investigated the roles of cytoplasmic ATP as both an energy source and a regulatory molecule in various steps of the excitation-contraction (E-C) coupling process in fast-twitch skeletal muscle fibres of the rat. Using mechanically skinned fibres with functional E-C coupling, it was possible to independently alter cytoplasmic ATP and free Mg2+. Electrical field stimulation was used to elicit action potentials (APs) within the sealed transverse tubular (T-) system, producing either twitch or tetanic (50 Hz) force responses. Measurements were also made of the amount of Ca2+ released by an AP in different cytoplasmic conditions. The rate of force development and relaxation of the contractile apparatus was measured using rapid step changes in Ca2+. Twitch force decreased substantially (approximately 30%) at 2 mm ATP compared to the level at 8 mm ATP, whereas peak tetanic force only declined by approximately 10% at 0.5 mm ATP. The rate of force development of the twitch and tetanus was slowed only slightly at ATP > or = 0.5 mm, but was slowed greatly (> 6-fold) at 0.1 mm ATP, the latter being due primarily to slowing of force development by the contractile apparatus. AP-induced Ca2+ release was decreased by approximately 10 and 20% at 1 and 0.5 mm ATP, respectively, and by approximately 40% by raising the Mg2+ to 3 mm. Adenosine inhibited Ca2+ release and twitch responses in a manner consistent with its action as a competitive weak agonist for the ATP regulatory site on the ryanodine receptor (RyR). These findings show that (a) ATP is a limiting factor for normal voltage-sensor activation of the RyRs, and (b) large reductions in cytoplasmic ATP, and concomitant elevation of Mg2+, substantially inhibit E-C coupling and possibly contribute to muscle fatigue in fast-twitch fibres in some circumstances.
Dutka et al. (Fri,) studied this question.