Regulation of NA/K-atpase /3-subuntt expression by ouabain and other hypertrophic stimuli in neonatal cardiac myocyteS

Partial inhibition of Na/K-ATPase by ouabain causes hypertrophie growth and regulates several early and late response genes including Na/K-ATPase aj subunit in cultured neonatal rat cardiac myocytes. When Na/K-ATPase β subunit mRNA was determined after myocytes were exposed to ouabain, ouabain increased β subunit mRNA in a dose and time dependent manner. Phenylephrine and PMA, two hypertrophie stimuli that do not inhibit Na/KATPase, also increased /3-subunit mRNA, suggesting an association of β subunit up-regulation with cardiac hypertrophy. When myocytes were exposed to ouabain in the presence of actinomycin D, ouabain failed to increase β subunit mRNA, suggesting that ouabain may transcriptionally regulate expression of the β subunit. When myocytes were transfected with luciferase reporter gene directed by the 5'-flanking region of β subunit, ouabain caused a significant increase in luciferase activity in the transfected cells, supporting a transcriptional regulation of β subunit gene by ouabain. The effects of ouabain on β subunit expression are extracellular Ca2+-, calmodulin-, and PKC-dependent. Inhibition of Ras by adenovirus-mediated expression of dominant negative Ras partially prevent ouabain-induced β expression. Inhibition of mitogen-activated protein kinase kinase (MEK) by PD98059 also caused a partial repression of ouabain-induced β expression. These data indicate that both Ras-dependent and Rasindependent pathways are involved in ouabain-induced β subunit expression in cardiac myocytes. Supported by a Grant-in-Aid from the American Heart Association.