Modulation by alkalosis of basal leak Ca2+ entry and store-depletion-induced Ca2+ entry was investigated in the vascular endothelial cell line ECV 304. Ca2+ entry was monitored as the increase in the intracellular free Ca2+ concentration ([Ca2+]i) induced by elevation of the extracellular Ca2+ concentration. When ECV 304 cells were challenged with 100 nM thapsigargin in nominally Ca2+-free solution, [Ca2+]i increased transiently, and the increase in [Ca2+]i during a subsequent cumulative elevation of extracellular Ca2+ (from nominally Ca2+-free up to 5 mM) was markedly enhanced compared with non-stimulated cells (i.e. basal Ca2+ leak). Prolonged elevation of the extracellular pH (pHo) from 7.4 to 7.9 did not affect resting [Ca2+]i or the thapsigargin-induced [Ca2+]i transient evoked in nominally Ca2+-free solution, but increased leak Ca2+ entry as well as store-depletion-activated Ca2+ entry significantly. Basal Ca2+ leak and store-depletion-activated Ca2+ entry were enhanced either by acute elevation of pHo from 7.4 to 7.9 or by chronic alkalosis (pHo = 7.9). Stimulation of Ca2+ entry by extracellular alkalosis was observed both in normal and in high extracellular K+ (110 mM) solution, suggesting that the effects of alkalosis are independent of membrane potential. The intracellular pH (pHi) increased slightly during both acute and chronic extracellular alkalosis (from 7.22±0.01 to 7.37±0.04 and 7.45±0.05 respectively). Elevation of pHi to 7.60±0.06 at constant pHo by administration of 20 mM NH4Cl failed to stimulate, and in fact inhibited, store-depletion-activated Ca2+ entry. Our results demonstrate that a decrease in the extracellular but not the intracellular proton concentration promotes both basal and stimulated Ca2+ entry into endothelial cells.
Divergent effects of extracellular and intracellular alkalosis on Ca2+ entry pathways in vascular endothelial cells
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Ichiro WAKABAYASHI, Klaus GROSCHNER; Divergent effects of extracellular and intracellular alkalosis on Ca2+ entry pathways in vascular endothelial cells. Biochem J 15 April 1997; 323 (2): 567–573. doi: https://doi.org/10.1042/bj3230567
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