Effect of Na+ on Na+,K+-ATPase alpha-subunit expression and Na+-pump activity in aortic smooth muscle cells.
In earlier studies we demonstrated that cyclical mechanical strain on vascular smooth muscle cells increases intracellular Na+ and upregulates the alpha-1 and alpha-2 isoform expression of Na+,K+-ATPase, and that the increase of intracellular Na+ and upregulation of the alpha-2 isoform expression are blocked by Gd3+, which blocks entry of ions (including 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 treatment. In parallel experiments, we measured Na+,K+-ATPase alpha isoform expression, Na+-pump activity and intracellular Na+ in cultured aortic smooth muscle cells after treatment with two concentrations of ouabain for various time periods. Treatment with 100 nM ouabain resulted in a significant elevation in intracellular Na+ after 1 (21%) and 2 h (12%), but the value returned to baseline after 12 h. Both alpha-1 and alpha-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 microM of ouabain, the effects on intracellular Na+, isoform expression and Na+-pump activity at earlier time points (1 h through 1 day) were similar to those with 100 nM treatment, but prolonged treatment (2 and 4 days) resulted in an accumulation of intracellular Na+ and inhibition of the isoform expression and Na+-pump activity, possibly due to general dysfunction of the cells as a result of chronic exposure to high concentrations of ouabain. We conclude that elevated intracellular Na+ can serve as a signal to mediate the alpha isoform upregulation and the regulatory process requires less than one day.
Liu
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Department of Pharmacology and Experimental Therapeutics, Louisiana State University Medical Center, New Orleans 70112, USA.
Songu-Mize
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D000818:Animals; D001011:Aorta / Q000187:drug effects*; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005682:Gadolinium / Q000494:pharmacology; D006706:Homeostasis / Q000187:drug effects; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects*; D010042:Ouabain / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012964:Sodium / Q000032:analysis / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D013997:Time Factors; D015854:Up-Regulation
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010042:Ouabain; D012964:Sodium; D005682:Gadolinium; D000254:Sodium-Potassium-Exchanging ATPase