The effects of fatigue and oxidation on contractile function of intact muscle fibers and myofibrils isolated from the mouse diaphragm

The goal of this study was to investigate the effects of repetitive stimulation and the oxidant H 2 O 2 on fatigue of diaphragm intact fibers and in myofibrils measured with different Ca 2+ concentrations. Intact fibers were isolated from mice diaphragm, and twitch and tetanic contractions (500 ms duration) were performed at different frequencies of stimulation ranging from 15 Hz to 150 Hz to establish a force-frequency relation before and after a fatigue and recovery protocol, without or after a treatment with H 2 O 2 . Fatigue was induced with isometric contractions (500 ms, 40 Hz) evoked every 0.8 seconds, with a total of 625 tetani. After the fatigue, the force recovery was followed by invoking tetanic contractions (500 ms, 40 Hz) every 1 min, with a total duration of 30 min. Individual myofibrils were also isolated from the mouse diaphragm and were tested for isometric contractions before and after treatment with H 2 O 2 and NAC. In a second series of experiments, myofibrils were activated at different pCa (pCa = −log 10 [Ca 2+ ]), before and after H 2 O 2 treatment. After 15 minutes of H 2 O 2 treatment, the myofibrillar force was decreased to 54 ± 12% of its control, maximal value, and a result that was reversed by NAC treatment. The force was also decreased after myofibrils were treated with H 2 O 2 and activated in pCa ranging between 4.5 and 5.7. These results suggest that fatigue in diaphragm intact fibers and at the myofibrils level is caused partially by oxidation of the contractile proteins that may be responsible for changing the force in various levels of Ca 2+ activation.