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. 1988 Oct 1;255(1):179-84.
doi: 10.1042/bj2550179.

Evidence that agonists stimulate bivalent-cation influx into human endothelial cells

T J Hallam  1 , R Jacob, J E Merritt
Affiliations

Evidence that agonists stimulate bivalent-cation influx into human endothelial cells

T J Hallam et al. Biochem J. .

Abstract

Human umbilical-vein endothelial cells stimulated with thrombin or histamine show an increase in [Ca2+]i (cytoplasmic free calcium concn.) that is maintained well above the basal pre-stimulated value as long as agonist and a source of extracellular Ca2+ are present. These results provide circumstantial evidence that agonists stimulate influx of Ca2+ across the plasma membrane and into the cytoplasm. Here, we have used Mn2+ as the extracellular bivalent cation which can bind to the fluorescent Ca2+ indicator fura-2 to quench its fluorescence completely. Human umbilical-vein endothelial cells were loaded with fura-2 and, in the presence of extracellular Mn2+, thrombin and histamine were shown to cause quenching of the intracellular dye. This result demonstrates conclusively that agonists can stimulate the influx of bivalent cations. Stimulated discharge of Ca2+ from intracellular stores and influx of Mn2+ were temporally resolved in the same cells to show that release of Ca2+ from intracellular stores clearly precedes influx. Influx of Mn2+ was also demonstrated when extracellular Mn2+ was added after agonist at a time when [Ca2+]i had fallen back to the basal value, showing that influx is not dependent on elevated [Ca2+]i.

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