Na-H antiport in cultured rat aortic smooth muscle: its role in cytoplasmic pH regulation

Abstract

We have investigated the role of the Na-H antiport in the regulation of intracellular pH (pHi) in vascular smooth muscle. Experiments were conducted on contractile-state rat aortic smooth muscle cells grown in primary culture and loaded with the pH-sensitive, fluorescent indicator 2',7',-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF). Cells equilibrated in a normal physiological salt solution (PSS) containing 135 mM Na, pH 7.4 at 37 degrees C, had a pHi of 7.16 +/- 0.04 (means +/- SE; n = 8). 5-(N-ethyl-N-isopropyl)amiloride (EIPA) caused a concentration-dependent fall in pHi. Removal of extracellular Na caused an intracellular acidification that was rapidly reversed on replacement of Na. The rate of recovery from NH4Cl-induced intracellular acidosis was dependent on extracellular Na concentration (Km 14.6 +/- 2.8 mM) and was accelerated by increasing the transmembrane Na gradient and slowed by decreasing it. Recovery from acidosis was completely abolished by either EIPA or the absence of extracellular Na. These results demonstrate that the Na-H antiport is an important mechanism for the maintenance and regulation of pHi in vascular smooth muscle cells. The BCECF fluorescence technique provides an ideal method for further studies on the mechanisms for pHi regulation in these cells.