Related Experiment Video
Updated: May 19, 2026
Isolation of Microvascular Endothelial Tubes from Mouse Resistance Arteries
Published on: November 25, 2013
Coordination of intercellular Ca(2+) signaling in endothelial cell tubes of mouse resistance arteries
Matthew J Socha1, Timothy L Domeier, Erik J Behringer
1Department of Medical Pharmacology and Physiology, University of Missouri, Columbia, Missouri 65212, USA.
Objective:
To test the hypothesis that Ca(2+) responses to GPCR activation are coordinated between neighboring ECs of resistance arteries.
Methods:
EC tubes were freshly isolated from superior epigastric arteries of C57BL/6 mice. Intercellular coupling was tested using microinjection of propidium iodide. Following loading with fluo-4 dye, intracellular Ca(2+) responses to ACh were imaged with confocal microscopy.
Results:
Cell-to-cell transfer of propidium iodide confirmed functional GJCs. A 1 μm ACh stimulus evoked Ca(2+) responses (9.8 ± 0.8/min, F/F(0) = 3.11 ± 0.2) which pseudo-line-scan analysis revealed as composed of Ca(2+) waves and spatially restricted Ca(2+) release events. A 100 nm ACh stimulus induced Ca(2+) responses of lower frequency (4.5 ± 0.7/min) and amplitude (F/F(0) = 1.95 ± 0.11) composed primarily of spatially restricted events. The time interval between Ca(2+) waves in adjacent cells (0.79 ± 0.12 s) was shorter (p < 0.05) than that between nonadjacent cells (1.56 ± 0.25 s). Spatially restricted Ca(2+) release events had similar frequencies and latencies between adjacent and nonadjacent cells. Inhibiting intracellular Ca(2+) release with 2-APB, Xestospongin C or thapsigargin eliminated Ca(2+) responses.
Conclusions:
With moderate GPCR stimulation, localized Ca(2+) release events predominate among cells. Greater GPCR stimulation evokes coordinated intercellular Ca(2+) waves via the ER. Calcium signaling during GPCR activation is complex among cells, varying with stimulus intensity and proximity to actively signaling cells.
