Multiple calcium sources are required for intracellular calcium mobilization during gastric organoid epithelial repair

Calcium (Ca(2+)) is a known accelerator for gastric wound repair. We have demonstrated in vivo and in vitro that intracellular Ca(2+) increases in the gastric epithelial cells directly adjacent to a damaged cell, and that this Ca(2+) rise is essential for the cellular migration that rapidly repairs the epithelium (restitution). While intracellular Ca(2+) has been shown to be an important signaling factor during epithelial restitution, the source from which this intracellular Ca(2+) originates remains unclear. Using gastric organoids derived from mice transgenic for a genetically encoded Ca(2+) indicator, we sought to investigate the potential sources of intracellular Ca(2+) mobilization. During confocal imaging, photodamage (PD) was induced to 1–2 gastric organoid epithelial cells and epithelial restitution measured simultaneously with changes in intracellular Ca(2+) (measured as FRET/CFP ratio in migrating cells adjacent to the damaged area). Inhibition of voltage‐gated Ca(2+) channels (verapamil, 10 µM) or store‐operated calcium entry (YM58483, 20 µM) resulted in delayed repair and dampened intracellular Ca(2+) response. Furthermore, inhibition of phospholipase C (U73122, 10 µM) or inositol trisphosphate receptor (2‐APB, 50 µM) likewise resulted in delayed repair and dampened Ca(2+) response. Results suggest both extracellular and intracellular Ca(2+) sources are essential for supplying the Ca(2+) mobilization that stimulates repair.