The Na,K-ATPase-Dependent Src Kinase Signaling Changes with Mesenteric Artery Diameter

Abstract

Inhibition of the Na,K-ATPase by ouabain potentiates vascular tone and agonist-induced contraction. These effects of ouabain varies between different reports. In this study, we assessed whether the pro-contractile effect of ouabain changes with arterial diameter and the molecular mechanism behind it. Rat mesenteric small arteries of different diameters (150–350 µm) were studied for noradrenaline-induced changes of isometric force and intracellular Ca2+ in smooth muscle cells. These functional changes were correlated to total Src kinase and Src phosphorylation assessed immunohistochemically. High-affinity ouabain-binding sites were semi-quantified with fluorescent ouabain. We found that potentiation of noradrenaline-sensitivity by ouabain correlates positively with an increase in arterial diameter. This was not due to differences in intracellular Ca2+ responses but due to sensitization of smooth muscle cell contractile machinery to Ca2+. This was associated with ouabain-induced Src activation, which increases with increasing arterial diameter. Total Src expression was similar in arteries of different diameters but the density of high-affinity ouabain binding sites increased with increasing arterial diameters. We suggested that ouabain binding induces more Src kinase activity in mesenteric small arteries with larger diameter leading to enhanced sensitization of the contractile machinery to Ca2+.