Intracellular Mechanisms Involved in D-Glucose–Mediated Amplification of Agonist-Induced Ca²⁺ Response and EDRF Formation in Vascular Endothelial Cells

Prolonged treatment of vascular endothelial cells with pathologically high D-glucose amplifies autacoid-induced Ca²⁺ mobilization and thus formation of nitric oxide. This study investigated the Ca²⁺ source for the change in endothelial Ca²⁺ response on agonist stimulation. Pretreatment with high D-glucose (44 vs. 5 mM) enhanced release of intracellular Ca²⁺ by bradykinin as a result of a 2.0-fold increased formation of inositol 1,4,5-trisphosphate. High D-glucose also amplified Ca²⁺ influx (2.0-fold). In high D-glucose preincubated cells, stimulation with bradykinin significantly increased transplasmalemmal ⁴⁵Ca²⁺ flux (3.2-fold) and caused a 2.0-fold increase in permeability to Mn²⁺, a surrogate for endothelial plasma membrane Ca²⁺ channels. A significant 2.0-fold increase occurred in the maximal slope, suggesting a higher rate of Mn²⁺ (Ca²⁺) influx. Ca²⁺ influx, stimulated by an inositol phosphate-independent depletion of intracellular Ca²⁺ stores with 2,5-di-(tert-butyl)-hydroquinone was also significantly increased 2.4-fold by high D-glucose, with no effect on intracellular Ca²⁺ release. D-glucose failed to modulate resting or stimulated cAMP levels. We suggest that prolonged exposure to pathologically high D-glucose increases formation of inositol polyphosphates, thus increasing Ca²⁺ release. Ca²⁺ entry is increased by amplification of unknown signal transduction mechanisms triggered by Ca²⁺ store depletion.