Abstract 17464: Regulation of Intracellular Ca2+ Waves and Triggered Activities by Mitochondrial Ca2+ Flux in Mouse Cardiomyocytes
Background: Recent studies have shown that mitochondria play important roles in Ca2+ homeostasis of cardiac myocytes. However, it is still unclear if mitochondrial Ca2+ flux can regulate the generation of Ca2+ waves and triggered activities in cardiomyocytes.
Methods and Results: Cytosolic Ca2+ (Cai2+) was imaged in fluo-4-AM loaded ventricular myocytes isolated from mice. Spontaneous sarcoplasmic reticulum (SR) Ca2+ release and Ca2+ waves were induced in the presence of high external Ca2+ (Cao2+, 4 mM). The protonophore carbonyl cyanide p —(trifluoromethoxy)phenylhydrazone (FCCP) reversibly raised basal Cai2+ levels in the presence, as well as absence of Cao2+, suggesting Ca2+ release from intracellular stores. Mitochondrial membrane potential (ΔΨ m) was monitored by TMRM fluorescence. FCCP at 0.01–0.1 μM, which partially depolarized ΔΨ m, increased the frequency and amplitude of Ca2+ waves in a dose-dependent manner. Simultaneous recording of cell membrane potentials showed the augmentation of delayed afterdepolarization amplitudes and frequencies, and induction of triggered action potentials. On the contrary, FCCP at higher concentrations (>0.5 μM), which completely dissipated ΔΨ m, eliminated Ca2+ waves while the basal Cai2+ remained high. The cease of Ca2+ waves was most likely due to the reduction of SR Ca2+ content as evaluated by rapid exposure to10 mM caffeine. Blocking sarcolemmal Na+-Ca2+ exchanger by substituting Na+ with Li+ in the perfusant further elevated basal Cai2+ and restored Ca waves. The effect of FCCP on Ca2+ waves was mimicked by antimycin A (an electron transport chain inhibitor disrupting ΔΨ m) or Ru360 (a mitochondrial Ca2+ uniporter inhibitor), but not by oligomycin (an ATP synthase inhibitor) or iodoacetic acid (a glycolytic inhibitor), excluding the contribution of intracellular ATP levels. The effects of FCCP on Ca2+ waves were counteracted by the mitochondrial permeability transition pore blocker cyclosporine A, or the mitochondrial Ca2+ uniporter activator kaempferol.
Conclusions: Mitochondrial Ca2+ release and uptake exquisitely control the local Ca2+ level in the micro-domain near SR ryanodine receptors and plays an important role in regulation of intracellular Ca2+ waves and arrhythmogenesis.
- © 2010 by American Heart Association, Inc.