EFFECT OF NA-ATPASE ALPHA-SUBUNIT EXPRESSION AND NA+-PUMP ACTIVITY INAORTIC SMOOTH-MUSCLE CELLS( ON NA+,K+)

In earlier studies we demonstrated that cyclical mechanical strain on vascular smooth muscle cells increases intracellular Na+ and upregulat es the alpha-l and alpha-2 isoform expression of Na+,K+-ATPase, and th at the increase of intracellular Na+ and upregulation of the alpha-2 i soform expression are blocked by Gd3+, which blocks entry of ions (inc luding Na+) through stretch-activated channels. The present study was designed to investigate the role of intracellular Na+ in Na+,K+-ATPase regulation by increasing intracellular Na+ with chronic ouabain treat ment. In parallel experiments, we measured Na+,K+-ATPase alpha isoform expression, Na+-pump activity and intracellular Na+ in cultured aorti c smooth muscle cells after treatment with two concentrations of ouaba in for various time periods. Treatment with 100 nM ouabain resulted in a significant elevation in intracellular Na+ after 1 (21%) and 2 h (1 2%), but the value returned to baseline after 12 h. Both alpha-1 and a lpha-2 subunits of Na+,K+-ATPase were significantly upregulated after 1 through 4 days. Na+-pump activity was also stimulated, and the time course of this effect closely followed protein expression. At 200 mu M of ouabain, the effects on intracellular Na+, isoform expression and Na+-pump activity at earlier time points (1 h through 1 day) were simi lar to those with 100 nM treatment, but prolonged treatment (2 and 4 d ays) resulted in an accumulation of intracellular Na+ and inhibition o f the isoform expression and Na+-pump activity, possibly due to genera l dysfunction of the cells as a result of chronic exposure to high con centrations of ouabain. We conclude that elevated intracellular Na+ ca n serve as a signal to mediate the a isoform upregulation and the regu latory process requires less than one day. (C) 1998 Elsevier Science B .V. All rights reserved.