Differential Regulation of Na/K-ATPaseα-subunit Isoform Gene Expressions in Cardiac Myocytes by Ouabain and Other Hypertrophic Stimuli

We showed before that partial inhibition of Na/K-ATPase by non-toxic concentrations of ouabain caused hypertrophic growth of neonatal rat cardiac myocytes, and induced several early- and late-response genes that are markers of cardiac hypertrophy. The aim of this study was to determine if the genes of theα-subunit isoforms of Na/K-ATPase were among those regulated by ouabain; and if so, to begin the characterization of the pathways regulating these genes. When neonatal myocytes, expressingα1- andα3-isoform messages, were exposed to 5–100μ ouabain,α1mRNA was not affected, butα3mRNA was decreased in a dose- and time-dependent manner. Oubain-induced down-regulation ofα3mRNA was accompanied by a decrease inα3-protein content in these myocytes. There was a significant correlation between ouabain effects onα3-repression and skeletalα-actin induction; also, ouabainʹs transcriptional effects on both genes were antagonised by retinoic acid. These findings suggested the association ofα3repression with ouabain-induced hypertrophy. Phenylephrine and a phorbol ester, two hypertrophic stimuli that do not inhibit Na/K-ATPase, also down-regulatedα3mRNA without affectingα1mRNA, suggesting thatα3-repression is a common feature of the hypertrophic phenotype in these myocytes. Ouabain-induced repression ofα3required the influx of extracellular Ca2+, and was antagonized by inhibitors of protein kinase C, Ca2+-calmodulin kinase, and mitogen-activated protein kinase but not by inhibition of protein kinase A. These data, and prior findings on the mechanisms of hypertrophic effects of phenylephrine and phorbol esters, suggest that transcriptional repression ofα3by ouabain and other hypertrophic stimuli involves a common step regulated by a mitogen-activated protein kinase.