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Manipulating IP 3 R‐mediated calcium release in permeabilized endothelial cell tubes of resistance arteries
Author(s) -
Socha Matthew J.,
Domeier Timothy L.,
Segal Steven S.
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.1058.8
Subject(s) - thapsigargin , endoplasmic reticulum , endothelium , calcium , chemistry , egta , biophysics , inositol , microbiology and biotechnology , endocrinology , medicine , receptor , biology , biochemistry
Calcium signaling is integral to endothelial cell (EC) function and is governed by stimuli originating both within the vessel lumen and from surrounding cells. Our goal was to gain insight into the intrinsic properties of inositol trisphosphate receptor (IP 3 R)‐mediated Ca 2+ release of resistance artery endothelium under conditions in which the endoplasmic reticulum functions as an isolated signaling organelle. Intact EC tubes were isolated from superior epigastric arteries of male C57BL/6 mice, permeabilized with saponin, and imaged using laser‐scanning confocal microscopy. Exposure to internal solution containing 100 nM free Ca 2+ (with 40 μM Fluo‐4) evoked Ca 2+ release events at a frequency of 3.9 ± 0.5 events/EC/min; these typically spread along entire ECs at ~15 μm/s (vs. ~17 μm/s for intact EC tubes). Release events were suppressed ( P <0.05) by removing free Ca 2+ with 500 μM EGTA (0 events), depleting ER Ca 2+ stores with 10 μM thapsigargin (0 events), or by inhibiting IP 3 R with 100 μM 2‐APB (0.4 ± 0.2 events/EC/min; P <0.05). Remarkably, increasing IP 3 levels in the internal solution tended to decrease Ca 2+ release events (5 nM: 2.1 ± 0.5; 50 nM: 1.5 ± 0.4; 10 μM: 0.5 ± 0.5 events/EC/min). We conclude that IP 3 R‐mediated Ca 2+ signaling can be manipulated in permeabilized endothelium of resistance arteries to evaluate key determinants of EC function in health and disease. (NIH R37HL041026, R01HL086483, F32HL107050)