Abstract 1100: Vasoconstrictor Coupling Factor 6 activates c-src by Intracellular Acidosis and Inhibits Prostacyclin Synthesis in Resistance Arterioles Vascular Smooth Muscle Cells
We showed that vasoconstrictor coupling factor 6 (CF6), a component of ATP synthase, suppresses the synthesis of prostacyclin by inducing intracellular acidosis in vascular endothelial cells. We recently showed that CF6 suppresses the synthesis of prostacyclin in an autocrine fashion in the resistance arteriole vascular smooth muscle cells (VSMC). However, little is known about the mechanism for the suppression of prostacyclin synthesis in VSMC and the role of CF6 in the genesis of hypertension. We investigated the intracellular signaling pathway of CF6 in VSMC, the role of cytosolic tyrosine kinase c-src, and the involvement of CF6 in the pathogenesis of hypertension. We cultured the resistance arteriole VSMC from the mesenteric artery network of spontaneously hypertensive rats (SHR, n=8) and Wistar Kyoto rats (WKY, n=8). Flow cytometry and Western blot analysis revealed that the β-subunit of ATP synthase, the receptor of CF6, was present on the surface of VSMC and its affinity was higher in SHR than in WKY, not being associated with difference in the receptor number. CF6 at 10−7M decreased intracellular pH in BCECF-loaded VSMC by activating plasma membrane ATP synthase, and inhibited arachidonic acid (AA) release. Intracellular acidosis and the suppression of AA release were greater in SHR than in WKY. CF6 at 10−7M enhanced phosphorylation of c-src at 2 minutes through 10 minutes, and it was greater in SHR than in WKY. Efrapeptin, a blocker of CF6 receptor, at 10−5M and β-subunit antibody blocked not only intracellular acidosis but the suppression of AA release, whereas PP1, a specific inhibitor of c-src, reversed the suppression of AA release without affecting intracellular acidosis. Blockade of intracellular acidosis with efrapeptine at 10−5M suppressed c-src activation by CF6 at 10−7M. The basal release of CF6 was 3-fold higher in SHR than in WKY, whereas the basal release of prostacyclin was suppressed in SHR and no difference was detected after neutralization with anti-CF6 antibody. These suggest that CF6 activates c-src by intracellular acidosis, resulting in the suppression of prostacyclin production in resistance arteriole VSMC. Since the c-src activation in response to CF6 was enhanced in SHR, CF6 may be involved in the genesis of hypertension.