Evidence of a K⁺-H⁺-ATPase in vascular smooth muscle cells

In earlier studies Na⁺-K⁺-adenosinetriphosphatase (ATPase) and Na⁺-K⁺- 2Cl^- cotransport partially accounted for vascular smooth muscle cell (VSMC) K⁺ (Rb⁺) uptake. In other cells Rb⁺ is taken up by a K⁺-H⁺-ATPase that is sensitive to NC-1300-B, SCH28080, omeprazole, and N-ethylmaleimide (NEM). This study examines the effects of K⁺-H⁺-ATPase inhibitors on VSMC. Rubidium uptake by primary cultures of canine coronary artery (CCA) VSMC or cultured rat aortic (CRA) VSMC line A₇r₅ was reduced 19-37% by NC-1300-B, SCH28080, or omeprazole. N-ethylmaleimide reduced CCA VSMC K⁺ content from 1.55 ± 0.02 to 1.24 ± 0.06 μeq/mg protein. The NC-1300-B-sensitive portion of CRA VSMC Rb⁺ uptake was not blocked by ouabain (0.1 mM) or bumetanide (0.1 mM), but was reduced by alkalinization with 7.5 mM NH₄Cl, and increased by acidification with 7.5 mM Na-acetate. Intracellular pH (pH(i)) of CRA VSMC was reduced 0.14 ± 0.03 U by NC-1300-B and 0.22 ± 0.03 U by NEM. pH(i) of CCA VSMC was reduced 0.20 ± 0.03 U by omeprazole (1 mM) and 0.20 ± 0.03 U or 0.20 ± 0.05 U by amiloride in the absence or presence of omeprazole, respectively. Fluorescence of 2',7'-bis(carboxyethyl)-5-(6')- carboxyfluorescein due to excitation at 500:441 nm in rat aortic strips was reduced by 0.21 ± 0.02 U by omeprazole and 0.22 ± 0.03 U by K⁺ removal and increased by 0.21 ± 0.06 U by K⁺ repletion. We conclude that VSMC possess a previously unknown Rb⁺ uptake mechanism. This newly discovered mechanism helps to maintain K⁺ gradient and pH(i) by extruding H⁺ in exchange for K⁺, and is presumably a K⁺-H⁺-ATPase similar to those described in other tissues.