In vivo cytosolic HO changes and Ca homeostasis in mouse skeletal muscle.

Hydrogen peroxide (HO) and calcium ions (Ca) are functional regulators of skeletal muscle contraction and metabolism. Although HO is one of the activators of the type-1 ryanodine receptor (RyR1) in the Ca release channel, the interdependence between HO and Ca dynamics remains unclear. This study tested the following hypotheses using an in vivo model of mouse tibialis anterior (TA) skeletal muscle. ) Under resting conditions, elevated cytosolic HO concentration ([HO]) leads to a concentration-dependent increase in cytosolic Ca concentration ([Ca]) through its effect on RyR1; and ) in hypoxia (cardiac arrest) and muscle contractions (electrical stimulation), increased [HO] induces Ca accumulation. Cytosolic HO (HyPer7) and Ca (Fura-2) dynamics were resolved by TA bioimaging in young C57BL/6J male mice under four conditions: ) elevated exogenous HO; ) cardiac arrest; ) twitch (1 Hz, 60 s) contractions; and ) tetanic (30 s) contractions. Exogenous HO (0.1-100 mM) induced a concentration-dependent increase in [HO] (+55% at 0.1 mM; +280% at 100 mM) and an increase in [Ca] (+3% at 1.0 mM; +8% at 10 mM). This increase in [Ca] was inhibited by pharmacological inhibition of RyR1 by dantrolene. Cardiac arrest-induced hypoxia increased [HO] (+33%) and [Ca] (+20%) 50 min postcardiac arrest. Compared with the exogenous 1.0 mM HO condition, [HO] after tetanic muscle contractions rose less than one-tenth as much, whereas [Ca] was 4.7-fold higher. In conclusion, substantial increases in [HO] levels evoke only modest Ca accumulation via their effect on the sarcoplasmic reticulum RyR1. On the other hand, contrary to hypoxia secondary to cardiac arrest, increases in [HO] from muscle contractions are small, indicating that HO generation is unlikely to be a primary factor driving the significant Ca accumulation after, especially tetanic, muscle contractions. We developed an in vivo mouse myocyte HO imaging model during exogenous HO loading, ischemic hypoxia induced by cardiac arrest, and muscle contractions. In this study, the interrelationship between cytosolic HO levels and Ca homeostasis during muscle contraction and hypoxic conditions was revealed. These results contribute to the elucidation of the mechanisms of muscle fatigue and exercise adaptation.