Studies are proposed to examine the physiological regulation of myocardial Na, K-ATPase in the course of early neonatal development and by thyroid hormone (T3). Plasma membrane- bound Na, K-ATPase consists of two dissimilar subunits, alpha and beta, present in an equimolar ratio. Three distinct mRNA isoforms encoding the alpha-subunit of the enzyme (alpha1, alpha2, and alpha3) have been characterized and the corresponding genes recently mapped to different chromosomes. Expression of the alpha1 and alpha2 polypeptide subunits moreover has been correlated respectively with the presence of low- and high-ouabain-affinity Na, K- pump sites. The beta- subunit of the enzyme, in contrast, appears to be represented by a single gene. Alpha1 and alpha3 mRNA isoforms are expressed in neonatal rat myocardium, whereas the alpha1 and alpha2 mRNA isoforms are present in adult cardiac tissue. As a part of this developmental process, Na, K-Atpase genes additionally become responsive to various epigenetic stimuli such as T3, a known inducer of myocardial Na, K-ATPase in adult mammals. In the present project mechanisms underlying both the T3 and developmental regulation of Na, K-ATPase activity in rat ventricular myocardium will be investigated. Special attention will be devoted to determining the precise time of onset of T3- inducibility of Na, K-ATPase in the course of development. As an initial goal, changes in Na, K-ATPase activity will be correlated with concomitant changes in enzyme molecules characterized by differing ouabain sensitivities. To explore the potential role of Na, K-ATPase biosynthesis, the abundance and translational activity of messenger RNA's encoding the alpha-subunit isoforms and the beta-subunit of the enzyme will be determined. Finally, the role of transcriptional regulation in mediating the positive and negative modulation of myocardial Na, K-ATPase gene expression will be investigated in developing hypothyroid, euthyroid, and hyperthyroid animals. The studies described in this application are designed to explore the molecular basis for the physiological regulation of myocardial Na, K-ATPase expression. Elucidation of basic mechanisms underlying the ontogeny and genomic regulation of cardiac Na, K- ATPases should be of value in future analyses of a variety of physiological and pathological states such as cardiac hypertrophy, congestive heart failure, and various hypermetabolic states.
