Background-Disturbances in pH affect artery function, but the mechanistic background remains controversial. We investigated whether Na ϩ ,HCO 3 Ϫ cotransporter NBCn1, by regulating intracellular pH (pH i ), influences artery function and blood pressure regulation. Methods and Results-Knockout of NBCn1 in mice eliminated Na ϩ ,HCO 3 Ϫ cotransport and caused a lower steady-state pH i in mesenteric artery smooth muscle and endothelial cells in situ evaluated by fluorescence microscopy. Using

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... phy, arteries from NBCn1 knockout mice showed reduced acetylcholine-induced NO-mediated relaxations and lower rho-kinase-dependent norepinephrine-stimulated smooth muscle Ca 2ϩ sensitivity. Acetylcholine-stimulated NO levels (electrode measurements) and N-nitro-L-arginine methyl ester-sensitive L-arginine conversion (radioisotope measurements) were reduced in arteries from NBCn1 knockout mice, whereas relaxation to NO-donor S-nitroso-Nacetylpenicillamine, acetylcholine-induced endothelial Ca 2ϩ responses (fluorescence microscopy), and total and Ser-1177 phosphorylated endothelial NO-synthase expression (Western blot analyses) were unaffected. Reduced NO-mediated relaxations in arteries from NBCn1 knockout mice were not rescued by superoxide scavenging. Phosphorylation of myosin phosphatase targeting subunit at Thr-850 was reduced in arteries from NBCn1 knockout mice. Evaluated by an in vitro assay, rho-kinase activity was reduced at low pH. Without CO 2 /HCO 3 Ϫ , no differences in pH i , contraction or relaxation were observed between arteries from NBCn1 knockout and wild-type mice. Based on radiotelemetry and tail-cuff measurements, NBCn1 knockout mice were mildly hypertensive at rest, displayed attenuated blood pressure responses to NO-synthase and rho-kinase inhibition and were resistant to developing hypertension during angiotensin-II infusion. Conclusions-Intracellular acidification of smooth muscle and endothelial cells after knockout of NBCn1 inhibits NO-mediated and rho-kinase-dependent signaling in isolated arteries and perturbs blood pressure regulation. (Circulation. 2011;124:1819-1829.) Key Words: pH Ⅲ hypertension Ⅲ blood pressure Ⅲ nitric oxide Ⅲ rho-kinase B lood pressure dysregulation is a major cause of human disease. Hypertension is a risk factor for development of coronary heart disease, stroke, and peripheral vascular disease 1-3 whereas hypotension is related to syncope and falls. 4,5 Both hyper-and hypotension increase overall mortality. 6 -8 Editorial see p 1806 Clinical Perspective on p 1829 Arterial tone regulation is important for blood pressure control and is modulated by local and systemic factors. Sustained changes in intracellular pH (pH i ) of vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) occur physiologically and pathologically, but have been difficult to investigate experimentally, and little is known about their vascular effects. 9 It has, however, been proposed that endothelial enzymes (eg, endothelial nitric oxide synthase [eNOS] 10 and endothelin converting enzyme 11 ), vascular ion channels (eg, Ca 2ϩ -activated K ϩ channels 12,13 and L-type Ca 2ϩ channels 14 ) and the Ca 2ϩ sensitivity of the contractile machinery 9,15,16 are important pH i -affected targets. Nitric oxide signaling in the cardiovascular system affects blood pressure regulation, 17-20 development of atherosclerosis, 20 and thrombocyte aggregation. 21 The catalytic activity of the isolated NO synthase displays a bell-shaped pH depen-The online-only Data Supplement is available with this article at http://circ.ahajournals.org/lookup/suppl/