Endotoxin impairs agonist-stimulated intracellular free calcium (Cai) responses in freshly dispersed aortic endothelial cells

Joyce J. Jones, Michael Sturek, H. Richard Adams, Janet L. Parker

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Impairment in endothelial cell intracellular free calcium (Cai) mobilization mechanisms may contribute to decreased nitric oxide (NO) biosynthesis and impaired vasorelaxation responses of endotoxemic guinea pigs to endothelium-dependent vasodilators. We tested this hypothesis using fura-2 microfluorometry to compare agonist-stimulated Cai responses of aortic endothelial cells freshly dispersed from guinea pigs 16 h after intraperitoneal injection of Escherichia coli endotoxin (lipopolysaccharide, LPS; 4 mg/kg) or saline (CON). In the presence of normal extracellular Ca2+ (2 mmol/L), basal (non-stimulated) endothelial Cai (340/380 nm fluorescence ratio, R) was not different between CON and LPS cells (1.1 ± 0.03 and 1.1 ± 0.03, respectively). However, exposure to ADP (10 μmol/L) produced a biphasic increase in Cai that was markedly decreased in cells from LPS-treated animals (P <0.0001). Peak ADP-stimulated Cai responses averaged 2.2 ± 0.21 in CON cells and 1.5 ± 0.11 (P <0.01) in cells dispersed from LPS-treated animals. Exposure to acetylcholine (ACh; 10 μmol/L) produced sustained increases in Cai (R = 1.4 ± 0.13) in CON cells; however, LPS abolished Cai responses to ACh. Exposure of endothelial cells to substance P (100 nmol/L) produced a biphasic increase in Cai that was not different between groups. In the absence of extracellular Ca2+ (plus 10 μmol/L EGTA), exposure to ADP (10 μmol/L) produced transient increases in Cai (Ca2+ release) that were decreased in cells from LPS-treated versus CON animals. Exposure to ACh in zero Ca2+ (10 μmol/L) produced smaller increases in Cai (peak R = 1.3 ± 0.12) in CON cells (when compared to ADP); however, Cai responses to ACh remained absent in cells from LPS-treated animals. Re-exposure to Ca2+ produced sustained ACh-induced Cai responses (Ca2+ influx) in cells from CON, but not LPS-treated animals; LPS markedly impaired (P <0.05) ADP-induced sustained Cai responses. Our data demonstrate that in vivo LPS exposure elicits decreased agonist-stimulated endothelial Cai responses primarily involving impaired Ca2+ influx mechanisms. Known dependence of endothelial agonist-stimulated NO synthesis on Cai suggests that defects in cell Ca2+ mobilization may contribute to LPS-induced impaired NO biosynthesis and decreased endothelium-dependent relaxation.

Original languageEnglish (US)
Pages (from-to)386-391
Number of pages6
JournalShock
Volume15
Issue number5
StatePublished - May 2001
Externally publishedYes

Fingerprint

Endotoxins
Endothelial Cells
Calcium
Adenosine Diphosphate
Nitric Oxide
Guinea Pigs
Cytophotometry
Endothelium-Dependent Relaxing Factors
Fura-2
Egtazic Acid
Substance P
Intraperitoneal Injections
Vasodilation
Acetylcholine
Endothelium
Lipopolysaccharides
Fluorescence

Keywords

  • Acetylcholine
  • ADP
  • Bacterial lipopolysaccharide
  • Bradykinin
  • Endothelium-dependent

ASJC Scopus subject areas

  • Physiology
  • Critical Care and Intensive Care Medicine

Cite this

Endotoxin impairs agonist-stimulated intracellular free calcium (Cai) responses in freshly dispersed aortic endothelial cells. / Jones, Joyce J.; Sturek, Michael; Adams, H. Richard; Parker, Janet L.

In: Shock, Vol. 15, No. 5, 05.2001, p. 386-391.

Research output: Contribution to journalArticle

Jones, Joyce J. ; Sturek, Michael ; Adams, H. Richard ; Parker, Janet L. / Endotoxin impairs agonist-stimulated intracellular free calcium (Cai) responses in freshly dispersed aortic endothelial cells. In: Shock. 2001 ; Vol. 15, No. 5. pp. 386-391.
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