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Nitric oxide inhibits capacitative Ca 2+ entry and enhances endoplasmic reticulum Ca 2+ uptake in bovine vascular endothelial cells
Author(s) -
Dedkova Ele.,
Blatter Lothar A.
Publication year - 2002
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2001.013258
Subject(s) - nitric oxide , endoplasmic reticulum , sodium nitroprusside , chemistry , intracellular , calcium , cytosol , nitric oxide synthase type iii , fura 2 , endothelial stem cell , biophysics , biochemistry , nitric oxide synthase , medicine , enos , endocrinology , biology , enzyme , organic chemistry , in vitro
In vascular endothelial cells, elevation of cytosolic free calcium concentration ([Ca 2+ ] i ) causes activation of nitric oxide synthase (NOS) and release of nitric oxide (NO). The goal of the study was to characterize the interplay between [Ca 2+ ] i and NO production in this cell type. Simultaneous measurements of [Ca 2+ ] i and intracellular NO concentration ([NO] i ) in cultured bovine vascular endothelial cells (CPAE cell line) with the fluorescent indicators fura‐2 and DAF‐2, respectively, revealed that Ca 2+ influx following agonist‐induced intracellular Ca 2+ store depletion (capacitative Ca 2+ entry, CCE) represents the preferential Ca 2+ source for the activation of the Ca 2+ ‐calmodulin‐dependent endothelial NOS (eNOS). Exposure to the NO donor sodium nitroprusside (SNP) showed that high NO levels suppressed CCE and had an inhibitory effect on Ca 2+ extrusion by the plasmalemmal Ca 2+ ‐ATPase. This inhibitory effect on CCE was mimicked by the membrane‐permeant cGMP analogue 8‐bromo‐cGMP, but was reversed by the NO scavenger haemoglobin and prevented by the inhibitor of the NO‐sensitive guanylate cyclase ODQ. Brief exposure to SNP reduced the peak of ATP‐induced Ca 2+ release from the endoplasmic reticulum (ER) and accelerated Ca 2+ reuptake into the ER. Prolonged incubation with SNP resulted in enhanced Ca 2+ loading of the ER, as revealed by direct measurements of store content with the ER‐entrapped low‐affinity Ca 2+ indicator mag‐fura‐2. The results suggest that in vascular endothelial cells, NO synthesis is under autoregulatory control that involves NO‐dependent [Ca 2+ ] i regulation. Via cGMP‐dependent inhibition of CCE and acceleration of Ca 2+ sequestration into the ER, NO can lower [Ca 2+ ] i and therefore exert an autoregulatory negative feedback on its own Ca 2+ ‐dependent synthesis.