Function and Regulation of the Na + -Ca2+ Exchanger NCX3 Splice Variants in Brain and Skeletal Muscle

Background: The Na+-Ca2+ exchanger isoform 3 (NCX3) participates in maintaining calcium homeostasis in brain and skeletal muscle. Results: The two splice variants of NCX3 have distinct regulation and capacity of exchange. Conclusion: Distinct calcium uptake capacity of muscle NCX3 suggests a novel function in muscle exercise physiology. Significance: This study brings a new understanding of the role of NCX3 variants in excitable tissues. Isoform 3 of the Na+-Ca2+ exchanger (NCX3) is crucial for maintaining intracellular calcium ([Ca2+](i)) homeostasis in excitable tissues. In this sense NCX3 plays a key role in neuronal excitotoxicity and Ca2+ extrusion during skeletal muscle relaxation. Alternative splicing generates two variants (NCX3-AC and NCX3-B). Here, we demonstrated that NCX3 variants display a tissue-specific distribution in mice, with NCX3-B as mostly expressed in brain and NCX-AC as predominant in skeletal muscle. Using Fura-2-based Ca2+ imaging, we measured the capacity and regulation of the two variants during Ca2+ extrusion and uptake in different conditions. Functional studies revealed that, although both variants are activated by intracellular sodium ([Na+](i)), NCX3-AC has a higher [Na+](i) sensitivity, as Ca2+ influx is observed in the presence of extracellular Na+. This effect could be partially mimicked for NCX3-B by mutating several glutamate residues in its cytoplasmic loop. In addition, NCX3-AC displayed a higher capacity of both Ca2+ extrusion and uptake compared with NCX3-B, together with an increased sensitivity to intracellular Ca2+. Strikingly, substitution of Glu(580) in NCX3-B with its NCX3-AC equivalent Lys(580) recapitulated the functional properties of NCX3-AC regarding Ca2+ sensitivity, Lys(580) presumably acting through a structure stabilization of the Ca2+ binding site. The higher Ca2+ uptake capacity of NCX3-AC compared with NCX3-B is in line with the necessity to restore Ca2+ levels in the sarcoplasmic reticulum during prolonged exercise. The latter result, consistent with the high expression in the slow-twitch muscle, suggests that this variant may contribute to the Ca2+ handling beyond that of extruding Ca2+.