Reciprocal Polarization of the Mitochondrial Ca2+ Uniporter and NA+-CA2+ Exchanger in Cardiac Muscle

Abstract

Control of myocardial energetics by Ca2+ signal propagation to the mitochondrial matrix includes local Ca2+ delivery from sarcoplasmic reticulum (SR) ryanodine receptors (RyR2) to the inner mitochondrial membrane (IMM) through the Ca2+ uniporter (mtCU). Ca2+ is extruded by the Na+-Ca2+ exchanger (NCLX), predominantly active in excitable tissues such as heart, and/or the H+-Ca2+ exchanger preferentially active in non-excitable cells. We have recently reported that the mtCU is strategically positioned in cardiac muscle at the mitochondria-junctional SR association in order to support more efficiently the local Ca2+ signals. In those studies we established a comparative abundance profile for a range of mitochondrial proteins between the mitochondrial and SR fractions of mouse and rat heart homogenates. While mtCU constituents were equally distributed (MCU, MICU1) or enriched in the SR fraction (EMRE), the NCLX was strongly (multiple fold) enriched in the mitochondrial fraction. We thus hypothesized, that NCLX was excluded from the mtCU hot spots at the dyad junctions and that it would be localized at areas where mitochondria associate with the network SR. Indeed, percoll purification of the crude mitochondrial fraction that decreased the abundance of MCU and EMRE lead to an enrichment of NCLX. Mitochondrial 45Ca2+ uptake assays using isolated mitochondria or SR fraction with vs. without NCLX inhibition (pharmacological or Na+ removal) revealed a robust Na+-dependent efflux working against the uptake in the mitochondrial but not the SR fraction. Direct measurement of the fractional Ca2+ release from the two membrane fractions also pointed toward much more efficient NCLX-mediated Ca2+ extrusion in the mitochondrial fraction than in the SR. Thus, mitochondrial Ca2+ uptake and Na+ dependent extrusion seem to be reciprocally polarized to optimize both the efficacy of receiving local Ca2+ signals at the dyadic interface and of feeding back the uptake Ca2+ to the SERCA pumps of the network SR.