Cytoplasmic Na+‐dependent modulation of mitochondrial Ca2+ via electrogenic mitochondrial Na+–Ca2+ exchange

To clarify the role of mitochondrial Na+–Ca2+ exchange (NCXmito) in regulating mitochondrial Ca2+ (Ca2+mito) concentration at intact and depolarized mitochondrial membrane potential (ΔΨmito), we measured Ca2+mito and ΔΨmito using fluorescence probes Rhod‐2 and TMRE, respectively, in the permeabilized rat ventricular cells. Applying 300 nm cytoplasmic Ca2+ (Ca2+c) increased Ca2+mito and this increase was attenuated by cytoplasmic Na+ (Na+c) with an IC50 of 2.4 mm. To the contrary, when ΔΨmito was depolarized by FCCP, a mitochondrial uncoupler, Na+c enhanced the Ca2+c‐induced increase in Ca2+mito with an EC50 of about 4 mm. This increase was not significantly affected by ruthenium red or cyclosporin A. The inhibition of NCXmito by CGP‐37157 further increased Ca2+mito when ΔΨmito was intact, while it suppressed the Ca2+mito increase when ΔΨmito was depolarized, suggesting that ΔΨmito depolarization changed the exchange mode from forward to reverse. Furthermore, ΔΨmito depolarization significantly reduced the Ca2+mito decrease via forward mode, and augmented the Ca2+mito increase via reverse mode. When the respiratory chain was attenuated, the induction of the reverse mode of NCXmito hyperpolarized ΔΨmito, while ΔΨmito depolarized upon inducing the forward mode of NCXmito. Both changes in ΔΨmito were remarkably inhibited by CGP‐37157. The above experimental data indicated that NCXmito is voltage dependent and electrogenic. This notion was supported theoretically by computer simulation studies with an NCXmito model constructed based on present and previous studies, presuming a consecutive and electrogenic Na+–Ca2+ exchange and a depolarization‐induced increase in Na+ flux. It is concluded that Ca2+mito concentration is dynamically modulated by Na+c and ΔΨmito via electrogenic NCXmito.