Isoform-specific regulation of myocardial Na,K-ATPase α-subunit in congestive heart failure: Role of norepinephrine

Background: Myocardial ouabain-binding sites and Na,K-ATPase activity are reduced in congestive heart failure (CHF), but the mechanisms by which CHF reduces the Na,K-ATPase remain unknown. We proposed to investigate whether the changes are accompanied by isoform-specific reductions of the Na,K- ATPase α-subunit proteins in CHF and whether similar changes could be produced by exogenous norepinephrine administration. Methods and Results: CHF was induced in dogs by rapid ventricular pacing at a rate of 225 beats per minute for 8 weeks (protocol 1). A second group of dogs were paced at 100 beats per minute and served as controls. In protocol 2, norepinephrine was infused in normal dogs using a subcutaneous osmotic minipump for 8 weeks. The control dogs received normal saline through the pump. Animals were studied after 8 weeks of pacing or norepinephrine infusion. After the baseline hemodynamics and interstitial norepinephrine concentration had been obtained, the hearts were removed for measuring [³H]ouabain-binding sites and Na,K- ATPase α-subunit proteins using isoform-specific monoclonal antibodies. Results: Myocardial [³H]ouabain-binding sites were reduced in dogs with CHF and chronic norepinephrine infusion. The Western blot analysis showed that adult canine hearts possess both α₁ and α₃ isoforms of the Na,K-ATPase α-subunit but not the α₂ isoform protein. CHF and NE infusion had no effect on the Na,K-ATPase α₁-subunit protein but did reduce the α₃ isoform protein significantly. In addition, there was a significant inverse correlation between the amount of myocardial α₃ isoform protein and interstitial norepinephrine content in the dogs. In contrast, the specific activity of the sarcolemmal marker 5'-nucleotidase did not differ among the groups of animals. Conclusions: The reduction of myocardial Na,K-ATPase in CHF is limited to the α₃ isoform. Furthermore, because similar changes in myocardial ouabain-binding sites and Na,K-ATPase α₃ isoform were produced by chronic norepinephrine infusion, the decrease in the Na,K-ATPase in CHF is most likely mediated via excess sympathetic stimulation.