Circulation Research, Vol 67, 814-825, Copyright © 1990 by American Heart Association

ARTICLES

Intracellular pH in human arterial smooth muscle. Regulation by Na+/H+ exchange and a novel 5-(N-ethyl-N-isopropyl)amiloride-sensitive Na(+)- and HCO3(-)-dependent mechanism

CB Neylon, PJ Little, EJ Cragoe Jr and A Bobik
Alfred Baker Medical Unit, Alfred Hospital, Prahran, Australia.

We investigated in a physiological salt solution (PSS) containing HCO3- the intracellular pH (pHi) regulating mechanisms in smooth muscle cells cultured from human internal mammary arteries, using the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) and 22Na+ influx rates. The recovery of pHi from an equivalent intracellular acidosis was more rapid when the cells were incubated in CO2/HCO3(-)- buffered PSS than in HEPES-buffered PSS. Recovery of pHi was dependent on extracellular Na+ (Km, 13.1 mM); however, it was not attenuated by 4- acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS), indicating the absence of SITS-sensitive HCO3(-)-dependent mechanisms. Recovery instead appeared mostly dependent on processes sensitive to 5- (N-ethyl-N-isopropyl)amiloride (EIPA), indicating the involvement of Na+/H+ exchange and a previously undescribed EIPA-sensitive Na(+)- and HCO3(-)-dependent mechanism. Differentiation between this HCO3(-)- dependent mechanism and Na+/H+ exchange was achieved after depletion of cellular ATP. Under these conditions, the NH4Cl-induced 22Na+ influx rate stimulated by intracellular acidosis was markedly attenuated in HEPES-buffered PSS but not in CO2/HCO3(-)-buffered PSS. EIPA also appeared to inhibit the two mechanisms differentially. In HEPES- buffered PSS containing 20 mM Na+, the EIPA inhibition curve for the intracellular acidosis-induced 22Na+ influx was monophasic (IC50, 39 nM), whereas in an identical CO2/HCO3(-)-buffered PSS, the inhibition curve exhibited biphasic characteristics (IC50, 37.3 nM and 312 microM). Taken together, the results indicate that Na+/H+ exchange and a previously undescribed EIPA-sensitive Na(+)- and HCO3(-)-dependent mechanism play an important role in regulating the pHi of human vascular smooth muscle. The involvement of the latter mechanism depends on the severity of the intracellular acidosis, varying from approximately 25% in severe intracellular acidosis up to 50% at lesser, more physiological, levels of induced acidosis.

This article has been cited by other articles:

				E. Boedtkjer, J. Praetorius, and C. Aalkjaer NBCn1 (slc4a7) Mediates the Na+-Dependent Bicarbonate Transport Important for Regulation of Intracellular pH in Mouse Vascular Smooth Muscle Cells Circ. Res., March 3, 2006; 98(4): 515 - 523. [Abstract] [Full Text] [PDF]

				E. J. Rios, M. Fallon, J. Wang, and L. A. Shimoda Chronic hypoxia elevates intracellular pH and activates Na+/H+ exchange in pulmonary arterial smooth muscle cells Am J Physiol Lung Cell Mol Physiol, November 1, 2005; 289(5): L867 - L874. [Abstract] [Full Text] [PDF]

				M. Ten Hove, M. G. J. Nederhoff, and C. J. A. Van Echteld Relative contributions of Na+/H+ exchange and Na+/HCO3- cotransport to ischemic Nai+ overload in isolated rat hearts Am J Physiol Heart Circ Physiol, January 1, 2005; 288(1): H287 - H292. [Abstract] [Full Text] [PDF]

				P.A. Underwood and S. M. Mitchell Low density lipoproteins in human plasma make vascular smooth muscle cells resistant to growth inhibition by heparin Cardiovasc Res, September 1, 2000; 47(4): 749 - 758. [Abstract] [Full Text] [PDF]

				H. Portner, C Bock, and A Reipschlager Modulation of the cost of pHi regulation during metabolic depression: a (31)P-NMR study in invertebrate (Sipunculus nudus) isolated muscle J. Exp. Biol., January 8, 2000; 203(16): 2417 - 2428. [Abstract] [PDF]

				A. Bobik, A. Agrotis, P. Kanellakis, R. Dilley, A. Krushinsky, V. Smirnov, E. Tararak, M. Condron, and G. Kostolias Distinct Patterns of Transforming Growth Factor-ß Isoform and Receptor Expression in Human Atherosclerotic Lesions : Colocalization Implicates TGF-ß in Fibrofatty Lesion Development Circulation, June 8, 1999; 99(22): 2883 - 2891. [Abstract] [Full Text] [PDF]

				A. Pushkin, N. Abuladze, I. Lee, D. Newman, J. Hwang, and I. Kurtz Cloning, Tissue Distribution, Genomic Organization, and Functional Characterization of NBC3, a New Member of the Sodium Bicarbonate Cotransporter Family J. Biol. Chem., June 4, 1999; 274(23): 16569 - 16575. [Abstract] [Full Text] [PDF]

				R. M. Touyz, G. He, L.-Y. Deng, and E. L. Schiffrin Role of Extracellular Signal-Regulated Kinases in Angiotensin II–Stimulated Contraction of Smooth Muscle Cells From Human Resistance Arteries Circulation, January 26, 1999; 99(3): 392 - 399. [Abstract] [Full Text] [PDF]

				G.L. Smith, C. Austin, C. Crichton, and S. Wray A review of the actions and control of intracellular pH in vascular smooth muscle Cardiovasc Res, May 1, 1998; 38(2): 316 - 331. [Abstract] [Full Text] [PDF]

				M. R. Ward, A. Agrotis, P. Kanellakis, R. Dilley, G. Jennings, and A. Bobik Inhibition of Protein Tyrosine Kinases Attenuates Increases in Expression of Transforming Growth Factor-ß Isoforms and Their Receptors Following Arterial Injury Arterioscler Thromb Vasc Biol, November 1, 1997; 17(11): 2461 - 2470. [Abstract] [Full Text]

				H. Wang, D. Singh, and L. Fliegel The Na+/H+ Antiporter Potentiates Growth and Retinoic Acid-induced Differentiation of P19 Embryonal Carcinoma Cells J. Biol. Chem., October 17, 1997; 272(42): 26545 - 26549. [Abstract] [Full Text] [PDF]

				M. Christ, K. Douwes, C. Eisen, G. Bechtner, K. Theisen, and M. Wehling Rapid Effects of Aldosterone on Sodium Transport in Vascular Smooth Muscle Cells Hypertension, January 1, 1995; 25(1): 117 - 123. [Abstract] [Full Text]