The role of calcium stores in fatigue of isolated single muscle fibres from the cane toad.

1. Intracellular calcium ([Ca2+]i) and tension were measured from single muscle fibres dissected from the cane toad (Bufo marinus). The amount of Ca2+ which could be released from the sarcoplasmic reticulum (SR) was estimated by brief (approximately 20 s) exposures to 4-chloro-m-cresol (4-CmC) or caffeine. 2. Muscle fatigue was produced by repeated tetani at 4 s or shorter intervals and continued until tension had fallen to 50% of the control. The intracellular free calcium concentration during a tetanus (tetanic [Ca2+]i) first increased and then steadily declined to 43+/-2% of control by the time tension had fallen to 50%. Over the period of fatigue the rapidly releasable Ca2+ from the SR fell to 46+/-6% of control. Tension and tetanic [Ca2+]i recovered to 93+/-3% and 100+/-4% of the control values after 20 min of rest. Over the same period rapidly releasable SR Ca2+ recovered to 98+/-12%. 3. When a similar number of tetani (200) were repeated at longer intervals (10 s), fibres showed only a small reduction in tension (to 85+/-1%) and tetanic [Ca2+]i did not change significantly. Under these conditions the rapidly releasable SR Ca2+ did not change significantly. 4. The recovery of rapidly releasable SR Ca2+ after fatigue was unaffected by removal of extracellular calcium but did not occur when oxidative phosphorylation was inhibited with cyanide. 5. These results suggest that an important cause of the decline of tetanic [Ca2+]i during fatigue is an equivalent decline in the amount of rapidly releasable SR Ca2+. The results show that the decline of rapidly releasable SR Ca2+ is related to a metabolic consequence of fatigue and are consistent with the hypothesis that Ca2+ precipitates with phosphate in the SR during fatigue.