Elevated Cytosolic Na + Decreases Mitochondrial Ca 2+ Uptake During Excitation-Contraction Coupling and Impairs Energetic Adaptation in Cardiac Myocytes

Abstract

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 (Δ) correlated linearly with Δ[Ca²⁺]_c. Under β-adrenergic stimulation, [Ca²⁺]_m decay was ≈2.5-fold slower than that of [Ca²⁺]_c, leading to diastolic accumulation of [Ca²⁺]_m when amplitude or frequency of Δ[Ca²⁺]_c increased. The MCU blocker Ru360 reduced Δ[Ca²⁺]_m and increased Δ[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.