Na/K-ATPase α1 regulation of cardiac myocyte metabolism: role of HIF-1α

Two novel cellular pathways of Na/K-ATPase (NKA)-mediated regulation of the cardiac myocyte metabolism have emerged. The first one is through inhibition of NKA’s ion-transporting function. The resulting elevation of intracellular Na + (Na i ) decreases mitochondrial calcium through a modulation of mitochondrial sodium calcium exchange, which decreases availability to calcium-dependent mitochondrial enzymes of the Tricarboxylic Acid Cycle and the Electron Transport Chain. When NKA expression is intact, this also induces a reversible metabolic shift to glycolysis via the hypoxia-inducible factor 1-alpha (HIF-1α). A second regulation occurs through NKAα1/Src signaling without NKA inhibition, but its role in HIF-1α activation and the glycolytic switch during Na i elevation have not been explored. To address this, we generated a CRISPR-Cas9-mediated knockout (KO) of NKAα1 in human ventricular cardiomyocytes (AC16 cells). In the NKAα1 KO, NKAα1 expression was reduced by 65% (p<0.001, n=10), resulting in an 80% decrease of cellular Na/K-ATPase activity (p<0.05, n=5) and a significant increase in Na i by 20% (p<0.05, n=3). Acute NKA inhibition with ouabain (100nM, 1 hr) to increase Na i activated HIF-1α in WT AC16 cells by 19% (p<0.05, n=3). However, despite elevated Na i HIF-1α activation was not observed in the NKAα1 KO. Functionally, Seahorse analyses revealed a significant reduction in basal respiration, maximal respiration, spare respiratory capacity, and ATP production (n=5-7, p<0.05), consistent with impaired mitochondrial function in NKAα1 KO cells. Notably, a glycolytic switch was not observed. In fact, glycolytic capacity and reserve were significantly reduced by 23% and 31%, respectively (n=5, p<0.05). These findings are consistent with a role of NKAα1 signaling in HIF-1α activation and the metabolic switch in response to increased Na i . Therefore, targeting NKAα1 signaling could be a new therapeutic strategy to address metabolic dysregulation in heart failure with altered cardiac NKAα1 content. NIH Grant R15 HL145666 (PI, Pierre) American Heart Association Grant 22POST917776 (PI, Pessoa) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.