Modulation of calcium homeostasis in cultured rat aortic endothelial cells by intracellular acidification

Acidosis produces vasodilation in a process that may involve the vascular endothelium. Because synthesis and release of endothelium-derived vasodilatory substances are linked to an increase in cytosolic calcium concentration ([Ca²⁺](i)), we examined the effect of intracellular acidification on cultured rat aortic endothelial cells loaded either with the pH-sensitive probe carboxy-seminaphthorhodafluor-1 or the Ca²⁺-sensitive fluorescent probe indo 1. The basal cytosolic pH (pH(i)) of endothelial monolayers in a 5% CO₂-HCO₃/^- buffer was 7.27 ± 0.02 and that in a bicarbonate-free solution was 7.22 ± 0.03. Acidification was induced either by removal of NH₄Cl (ΔpH(i) = -0.10 ± 0.02), changing from a bicarbonate- free to a 5% CO₂-HCO₃/^--buffered solution at constant buffer pH (ΔpH(i) = -0.18 ± 0.03), or changing from a 5% to a 20% CO₂-HCO₃/^- solution (ΔpH(i) = -0.27 ± 0.07). Regardless of the method used, intracellular acidification increased [Ca²⁺](i) as indexed by indo 1 fluorescence. The increase in [Ca²⁺](i) induced by changing from a 5 to a 20% CO₂-HCO₃/^- solution was not significantly altered by removal of buffer Ca²⁺ either before or after depletion of bradykinin- and thapsigargin-sensitive intracellular Ca²⁺ stores. Thus intracellular acidification of vascular endothelial cells releases Ca²⁺ into the cytosol either from pH-sensitive intracellular buffer sites, mitochondria, or from bradykinin- and thapsigargin-insensitive intracellular stores. This Ca²⁺ mobilization may be linked to endothelial synthesis and release of vasodilatory substances during acidosis.