The purpose of this study was to investigate the mechanism underlying sarcoplasmic reticulum (SR) Ca leakage after in vivo contractions. Rat gastrocnemius muscles were electrically stimulated in vivo, and then mechanically skinned fibers and SR microsomes were prepared from the muscles excised 30 min after repeated high-intensity contractions. The mechanically skinned fibers maintained the interaction between dihydropyridine receptors (DHPRs) and ryanodine receptors (RyRs), whereas the SR microsomes did not. Interestingly, skinned fibers from the stimulated muscles showed increased SR Ca leakage, whereas Ca leakage decreased in SR microsomes from the stimulated muscles. To enhance the orthograde signal of DHPRs, SR Ca leakage in the skinned fiber was measured ) under a continuously depolarized condition and ) in the presence of nifedipine. As a result, in either of the two conditions, SR Ca leakage in the rested fibers reached a level similar to that in the stimulated fibers. Furthermore, the increased SR Ca leakage from the stimulated fibers was alleviated by treatment with 1 mM tetracaine (Tet) but not by treatment with 3 mM free Mg (3 Mg). Tet exerted a greater inhibitory effect on the DHPR signal to RyR than 3 Mg, although their inhibitory effects on RyR were almost similar. These results suggest that the increased Ca leakage after muscle contractions is mainly caused by the orthograde signal of DHPRs to RyRs.
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