Distinct Mechanisms of Sodium–Calcium Exchanger NCX1.4 Regulation by Ca2+, ATP, and PIP2

Mammalian Na+/Ca2+ exchangers (NCXs) mediate inward (Ca2+-exit) or outward (Ca2+-entry) currents (INCX), with the peak current (Ipeak) declining to the steady-state current (Iss). The Ca2+-, ATP-, and PIP2-dependent activation of cardiac (NCX1.1) inward INCX has been previously documented. To investigate possible interactions among regulatory modes, we examined the effects of ATP and PIP2 on outward INCX when the CBD1 and CBD2 domains of NCX1.4 (brain variant) are Ca2+-occupied and -unoccupied. At nominally zero cytosolic Ca2+, ATP and PIP2 significantly reduced the outward Ipeak amplitude. While increasing cytosolic Ca2+ increased Ipeak and Iss/Ipeak, consistent with CBD1- and CBD2-dependent regulation, no significant effects of ATP or PIP2 were detected at 0.1–10 μM cytosolic Ca2+. These observations indicate distinct responses under the Ca2+ conditions examined and warrant direct mechanistic investigation. Although we observed opposing effects of ATP and PIP2 on inward (in NCX1.1) and outward (in NCX1.4) currents using different patch-clamp configurations and protocols, the data collectively suggest that Ca2+, ATP, and PIP2 regulatory modes may operate differently on the forward and reverse modes in NCX1.1 and NCX1.4 variants. Given the important physiological implications across distinct cell types (e.g., cardiomyocytes, neurons, glia, and kidney), careful investigation is needed to resolve how distinct regulatory modes affect the forward and reverse modes of mammalian NCX variants.