Endothelin-1 augments Na⁺/H⁺ exchange activity in murine pulmonary arterial smooth muscle cells via Rho kinase

• Published in: PloS one Vol. 7; no. 9; p. e46303
• Main Authors: Undem, Clark, Rios, Eon J, Maylor, Julie, Shimoda, Larissa A
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 28-09-2012; Public Library of Science (PLoS)
• Subjects: Activation; Animals; Augmentation; Biology; Cell adhesion & migration; Cell growth; Cell proliferation; Cells, Cultured; Circulatory system; Data processing; Dose-Response Relationship, Drug; Endothelin 1; Endothelin-1 - pharmacology; Enzyme Activation - drug effects; Enzyme inhibitors; Exchanging; Fluoresceins; Fluorescence; Fluorescent Dyes; Homeostasis; Hydrogen; Hydrogen ions; Hydrogen-Ion Concentration; Hypertension; Kinases; Lungs; Male; Medicine; Mice; Mice, Inbred C57BL; Microscopy; Microscopy, Fluorescence; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - drug effects; Muscle, Smooth, Vascular - enzymology; Muscles; Myocytes, Smooth Muscle - cytology; Myocytes, Smooth Muscle - drug effects; Myocytes, Smooth Muscle - enzymology; Na+/H+-exchanging ATPase; pH effects; Protein kinase C; Protein Kinase C - metabolism; Protein Kinase Inhibitors - pharmacology; Pulmonary Artery - cytology; Pulmonary Artery - drug effects; Pulmonary Artery - enzymology; Rho-associated kinase; rho-Associated Kinases - antagonists & inhibitors; rho-Associated Kinases - metabolism; Rodents; Signal transduction; Signal Transduction - drug effects; Signaling; Smooth muscle; Sodium-Potassium-Exchanging ATPase - antagonists & inhibitors; Sodium-Potassium-Exchanging ATPase - metabolism; United States > US; Maryland; Baltimore Maryland
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0046303

Excessive production of endothelin-1 (ET-1), a potent vasoconstrictor, occurs with several forms of pulmonary hypertension. In addition to modulating vasomotor tone, ET-1 can potentiate pulmonary arterial smooth muscle cell (PASMC) growth and migration, both of which contribute to the vascular remodeling that occurs during the development of pulmonary hypertension. It is well established that changes in cell proliferation and migration in PASMCs are associated with alkalinization of intracellular pH (pH(i)), typically due to activation of Na(+)/H(+) exchange (NHE). In the systemic vasculature, ET-1 increases pH(i), Na(+)/H(+) exchange activity and stimulates cell growth via a mechanism dependent on protein kinase C (PKC). These results, coupled with data describing elevated levels of ET-1 in hypertensive animals/humans, suggest that ET-1 may play an important role in modulating pH(i) and smooth muscle growth in the lung; however, the effect of ET-1 on basal pH(i) and NHE activity has yet to be examined in PASMCs. Thus, we used fluorescent microscopy in transiently (3-5 days) cultured rat PASMCs and the pH-sensitive dye, BCECF-AM, to measure changes in basal pH(i) and NHE activity induced by increasing concentrations of ET-1 (10(-10) to 10(-8) M). We found that application of exogenous ET-1 increased pH(i) and NHE activity in PASMCs and that the ET-1-induced augmentation of NHE was prevented in PASMCs pretreated with an inhibitor of Rho kinase, but not inhibitors of PKC. Moreover, direct activation of PKC had no effect on pH(i) or NHE activity in PASMCs. Our results indicate that ET-1 can modulate pH homeostasis in PASMCs via a signaling pathway that includes Rho kinase and that, in contrast to systemic vascular smooth muscle, activation of PKC does not appear to be an important regulator of PASMC pH(i).