Faculty Collaboration Database

Simultaneous in situ monitoring of intracellular Ca2+ and NO in endothelium of coronary arteries. Am J Physiol Heart Circ Physiol 2002 Dec;283(6):H2725-32

Date

10/22/2002

Pubmed ID

12388315

DOI

10.1152/ajpheart.00428.2002

Scopus ID

2-s2.0-0036889893 (requires institutional sign-in at Scopus site)   37 Citations

Abstract

We developed an in situ assay system to simultaneously monitor intracellular Ca(2+) concentration ([Ca(2+)](i), fura 2 as indicator) and nitric oxide (NO) levels [4,5-diaminofluorescein as probe] in the intact endothelium of small bovine coronary arteries by using a fluorescent microscopic imaging technique with high-speed wavelength switching. Bradykinin (BK; 1 microM) stimulated a rapid increase in [Ca(2+)](i) followed by an increase in NO production in the endothelial cells. The protein tyrosine phosphatase inhibitor phenylarsine oxide (PAO; 10 microM) induced a gradual, small increase in [Ca(2+)](i) and a slow increase in intracellular NO levels. Removal of extracellular Ca(2+) and depletion of Ca(2+) stores completely blocked BK-induced increase in NO production but had no effect on PAO-induced NO production. However, a further reduction of [Ca(2+)](i) by application of BAPTA-AM or EGTA with ionomycin abolished the PAO-induced NO increase. These results indicate that a simultaneous monitoring of [Ca(2+)](i) and intracellular NO production in the intact endothelium is a powerful tool to study Ca(2+)-dependent regulation of endothelial nitric oxide synthase, which provides the first direct evidence for a permissive role of Ca(2+) in tyrosine phosphorylation-induced NO production.

Author List

Yi FX, Zhang AY, Campbell WB, Zou AP, Van Breemen C, Li PL

Author

William B. Campbell PhD Professor in the Pharmacology and Toxicology department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Animals
Arsenicals
Bradykinin
Calcium
Cattle
Chelating Agents
Coronary Vessels
Endothelium, Vascular
Enzyme Inhibitors
Fluorescent Dyes
In Vitro Techniques
Intracellular Fluid
Ionophores
Microscopy, Fluorescence
Nitric Oxide
Protein Tyrosine Phosphatases