Na (+) -K (+) -ATPase that redistributes to apical membrane during ATP depletion remains functional
Molitoris, B.A.
American Journal of Physiology 265(5 Pt 2): F693-F697
1993
ISSN/ISBN: 0002-9513 PMID: 8238549 Document Number: 422191
We have previously demonstrated using immunocytochemical, histochemical, and biochemical techniques that ischemia in vivo and ATP depletion in vitro result in dissociation of Na+-K+-adenosinetriphosphatase (ATPase) from the actin cytoskeleton and redistribution to the apical domain in renal proximal tubule cells. To directly evaluate whether apical Na+-K+-ATPase retained Na+ pumping activity, a rapidly reversible model of cellular ATP depletion in confluent LLC-PK-1 cells grown on semipermeable membranes was utilized. Tight-junction integrity, monitored by electrical resistance, was lost during ATP depletion and reestablished during 2 h of ATP repletion. Total cellular Na+-K+-ATPase activity and total surface membrane (3H)ouabain binding remained constant, but specific apical (3H)ouabain binding increased (7 vs. 26 fmol/filter, P lt 0.01). Apical (3H)ouabain binding returned to base-line during 5 h of ATP repletion. Apically applied ouabain was then used to selectively inhibit apical Na+-K+-ATPase. It had no effect on apical-to-basolateral Na+ flux under physiological conditions (1.3 +- 0.61 vs. 1.27 +- 0.46 meq cntdot filter-1 cntdot 30 min-1), but it increased the apical-to-basolateral flux in ATP-depleted and then repleted monolayers (0.39 +- 0.12 vs. 0.83 +- 0.27 meq cntdot filter-1 cntdot 30 min-1, P lt 0.01), implying that apical Na+-K+-ATPase retained Na+ pumping activity. Together, these data imply that ATP depletion induce loss of surface membrane polarity resulting in redistribution of functional proteins to the alternate domain.