Extracellular acidosis and very low [Na⁺] inhibit NBCn1- and NHE1-mediated net acid extrusion from mouse vascular smooth muscle cells

Abstract

Aim The electroneutral Na+, cotransporter NBCn1 and Na+/H+ exchanger NHE1 regulate acid–base balance in vascular smooth muscle cells (VSMCs) and modify artery function and structure. Pathological conditions – notably ischaemia – can dramatically perturb intracellular (i) and extracellular (o) pH and [Na+]. We examined effects of low [Na+]o and pHo on NBCn1 and NHE1 activity in VSMCs of small arteries. Methods We measured pHi by 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein-based fluorescence microscopy of mouse mesenteric arteries and induced intracellular acidification by prepulse technique. Results NBCn1 activity – defined as Na+-dependent, amiloride-insensitive net base uptake with CO2/ present – was inhibited equally when pHo decreased from 7.4 (22 mm /5% CO2) by metabolic (pHo 7.1/11 mm : 22 ± 8%; pHo 6.8/5.5 mm : 61 ± 7%) or respiratory (pHo 7.1/10% CO2: 35 ± 11%; pHo 6.8/20% CO2: 56 ± 7%) acidosis. Extracellular acidosis more prominently inhibited NHE1 activity – defined as Na+-dependent net acid extrusion without CO2/ present – at both pHo 7.1 (45 ± 9%) and 6.8 (85 ± 5%). Independently of pHo, lowering [Na+]o from 140 to 70 mm reduced NBCn1 and NHE1 activity <20% whereas transport activities declined markedly (25–50%) when [Na+]o was reduced to 35 mm. Steady-state pHi decreased more during respiratory (ΔpHi/ΔpHo = 71 ± 4%) than metabolic (ΔpHi/ΔpHo = 30 ± 7%) acidosis. Conclusion Extracellular acidification inhibits NBCn1 and NHE1 activity in VSMCs. NBCn1 is equivalently inhibited when pCO2 is raised or []o decreased. Lowering [Na+]o inhibits NBCn1 and NHE1 markedly only below the typical physiological and pathophysiological range. We propose that inhibition of Na+-dependent net acid extrusion at low pHo protects against cellular Na+ overload at the cost of intracellular acidification.