Regulation of the Na/K
ATPase by
protein kinases is model-specific. We have observed a profound activation of the sarcolemmal Na/K
ATPase during cardiac
ischemia, which is masked by an inhibitor of the
enzyme in the cytosol. The aim of these studies was to characterize the pathways involved in this activation in the Langendorff-perfused rat heart. Na/K
ATPase activity was determined by measuring
ouabain-sensitive
phosphate generation by cardiac homogenates at 37 degrees C. In isolated sarcolemma,
ischemia (30 min) caused a substantial activation of the Na/K
ATPase compared with aerobic controls, which was abolished by perfusing the heart with
staurosporine or
H89. However, the alpha1 subunit of the Na/K
ATPase was not phosphorylated during
ischemia. The sarcolemmal
protein phospholemman (PLM) was found associated with the Na/K
ATPase alpha1 and beta1 but not alpha2 subunits, and PLM increased its association with the catalytic subunit of PKA following
ischemia. In vitro 14-3-3 binding assays indicated that PLM was phosphorylated following
ischemia. These results indicate that the
ischemia-induced activation of the Na/K
ATPase is indirect, through phosphorylation of PLM, which is an integral part of the Na/K
ATPase enzyme complex in the heart. The role of PLM is analogous to
phospholamban in regulating the
sarcoplasmic reticulum calcium ATPase.