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

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To clarify the role of mitochondrial Na(+)-Ca(2+) exchange (NCX(mito)) in regulating mitochondrial Ca(2+) (Ca(2+)(mito)) concentration at intact and depolarized mitochondrial membrane potential (DeltaPsi(mito)), we measured Ca(2+)(mito) and DeltaPsi(mito) using fluorescence probes Rhod-2 and TMRE, respectively, in the permeabilized rat ventricular cells. Applying 300 nm cytoplasmic Ca(2+) (Ca(2+)(c)) increased Ca(2+)(mito) and this increase was attenuated by cytoplasmic Na(+) (Na(+)(c)) with an IC(50) of 2.4 mm. To the contrary, when DeltaPsi(mito) was depolarized by FCCP, a mitochondrial uncoupler, Na(+)(c) enhanced the Ca(2+)(c)-induced increase in Ca(2+)(mito) with an EC(50) of about 4 mm. This increase was not significantly affected by ruthenium red or cyclosporin A. The inhibition of NCX(mito) by CGP-37157 further increased Ca(2+)(mito) when DeltaPsi(mito) was intact, while it suppressed the Ca(2+)(mito) increase when DeltaPsi(mito) was depolarized, suggesting that DeltaPsi(mito) depolarization changed the exchange mode from forward to reverse. Furthermore, DeltaPsi(mito) depolarization significantly reduced the Ca(2+)(mito) decrease via forward mode, and augmented the Ca(2+)(mito) increase via reverse mode. When the respiratory chain was attenuated, the induction of the reverse mode of NCX(mito) hyperpolarized DeltaPsi(mito), while DeltaPsi(mito) depolarized upon inducing the forward mode of NCX(mito). Both changes in DeltaPsi(mito) were remarkably inhibited by CGP-37157. The above experimental data indicated that NCX(mito) is voltage dependent and electrogenic. This notion was supported theoretically by computer simulation studies with an NCX(mito) model constructed based on present and previous studies, presuming a consecutive and electrogenic Na(+)-Ca(2+) exchange and a depolarization-induced increase in Na(+) flux. It is concluded that Ca(2+)(mito) concentration is dynamically modulated by Na(+)(c) and DeltaPsi(mito) via electrogenic NCX(mito).