Phosphatidyl inositol‐4,5‐bisphosphate bound to bovine cardiac Na+/Ca2+ exchanger displays a MgATP regulation similar to that of the exchange fluxes

This work shows the existence of a phosphatidylinositol 4,5‐bisphosphate (PtdIns‐4,5‐P2) bound form of the cardiac sarcolemmal Na+/Ca2+ exchanger. That was demonstrated in Western blots and cross‐immunoprecipitation by using specific antibodies against the NCX1 exchanger (NCX1) and against PtdIns‐4,5‐P2. In addition, PtdIns‐4,5‐P2 bound to the Na+/Ca2+ exchanger and the Na+/Ca2+ exchange fluxes displayed a similar MgATP regulation: (a) both increase by 100–130% when membrane vesicles are incubated (15–20 s at 37 °C) with 1 mm MgATP and 1 µm Ca2+ (b) in the presence of 100 µm Ca2+, MgATP fails to stimulate the exchange fluxes and does not modify the levels of PtdIns‐4,5‐P2 bound to the exchanger. In addition, in the absence of Ca2+, the net synthesis of total membrane PtdIns‐4,5‐P2 is greatly reduced compared with that in the presence of 1 µm Ca2+. Furthermore, in the absence of Ca2+ there is no effect of MgATP on the levels of PtdIns‐4,5‐P2 bound to the exchanger. These results indicate that, in bovine heart, MgATP‐stimulation of Na+/Ca2+ exchange is associated with intracellular Ca2+‐dependent levels of PtdIns‐4,5‐P2 bound to the exchanger molecule.