Elevated cytosolic Na+ decreases mitochondrial Ca2+ uptake during excitation-contraction coupling and impairs energetic adaptation in cardiac myocytes

Mitochondrial Ca2+([Ca2+]m) regulates oxidative phosphorylation and thus contributes to energy supply and demand matching in cardiac myocytes. Mitochondria take up Ca2+ via the Ca2+ uniporter (MCU) and extrude it through the mitochondrial Na+/Ca2+ exchanger (mNCE). It is controversial whether mitochondria take up Ca2+ rapidly, on a beat-to-beat basis, or slowly, by temporally integrating cytosolic Ca2+([Ca2+](c)) transients. Furthermore, although mitochondrial Ca2+ efflux is governed by mNCE, it is unknown whether elevated intracellular Na+([Na+](i)) affects mitochondrial Ca2+ uptake and bioenergetics. To monitor [Ca2+](m), mitochondria of guinea pig cardiac myocytes were loaded with rhod-2-acetoxymethyl ester (rhod-2 AM), and [Ca2+](c) was monitored with indo-1 after dialyzing rhod-2 out of the cytoplasm. [Ca2+](c) transients, elicited by voltage-clamp depolarizations, were accompanied by fast [Ca2+](m) transients, whose amplitude (Delta) correlated linearly with Delta[Ca2+](c). Under beta-adrenergic stimulation, [Ca2+](m) decay was approximate to 2.5-fold slower than that of [Ca2+](c), leading to diastolic accumulation of [Ca2+](m) when amplitude or frequency of Delta[Ca2+](c) increased. The MCU blocker Ru360 reduced Delta[Ca2+](m) and increased Delta[Ca2+](c), whereas the mNCE inhibitor CGP-37157 potentiated diastolic [Ca2+](m) accumulation. Elevating [Na+](i) from 5 to 15 mmol/L accelerated mitochondrial Ca2+ decay, thus decreasing systolic and diastolic [Ca2+](m). In response to gradual or abrupt changes of workload, reduced nicotinamide-adenine dinucleotide (NADH) levels were maintained at 5 mmol/L [Na+](i), but at 15 mmol/L, the NADH pool was partially oxidized. The results indicate that (1) mitochondria take up Ca2+ rapidly and contribute to fast buffering during a [Ca2+](c) transient; and (2) elevated [Na+](i) impairs mitochondrial Ca2+ uptake, with consequent effects on energy supply and demand matching. The latter effect may have implications for cardiac diseases with elevated [Na+](i).