Mechanisms Through Which Agents Of Muscle Fatigue, Acidosis And Phosphate, Inhibit Muscle Myosin Function.

Abstract

PURPOSE: During muscle fatigue from intense activity, elevated concentrations of hydrogen ions (acidosis) and inorganic phosphate (Pi) inhibit muscle’s ability to generate force and motion. However, it is not clear exactly how these metabolic by-products reduce the force and enzymatic function of muscle’s molecular motor, myosin. METHODS: To determine these mechanisms we directly measured the effect of these fatigue agents on the force generating capacity of isolated myosin in a laser trap assay and on its ability to hydrolyze ATP in an ATPase assay. RESULTS: Acidosis (pH 7.4 vs. 6.5) in a mini-ensemble laser trap assay reduced myosin’s average force production by 20% (p < 0.05) due to a slowed rate of actomyosin binding. This conclusion was supported by the observation that acidosis slowed myosin’s ability to hydrolyze ATP by roughly 90% (p < 0.05) in a solution assay. By contrast elevated levels of Pi (0 vs. 10-15mM), in the presence of low pH (6.5), caused a similar reduction in force. However, this was likely due to an accelerated rate of myosin’s detachment from actin, because myosin’s ATPase rate also recovered back toward the control value (pH 7.4, no Pi) when Pi was added. CONCLUSION: Thus, these data provide unique insight into the molecular mechanisms that underlie the loss of muscle function during fatigue. In our current work we are using these findings to explore methods to mitigate these effects in vitro in a first step toward attenuating fatigue in diseases such as chronic heart failure.