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

Calcium (Ca ) is a known accelerator for gastric wound repair. We have demonstrated in vivo and in vitro that intracellular Ca increases in the gastric epithelial cells directly adjacent to a damaged cell, and that this Ca rise is essential for the cellular migration that rapidly repairs the epithelium (restitution). While intracellular Ca has been shown to be an important signaling factor during epithelial restitution, the source from which this intracellular Ca originates remains unclear. Using gastric organoids derived from mice transgenic for a genetically encoded Ca indicator, we sought to investigate the potential sources of intracellular Ca 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 (measured as FRET/CFP ratio in migrating cells adjacent to the damaged area). Inhibition of voltage-gated Ca channels (verapamil, 10 µM) or store-operated calcium entry (YM58483, 20 µM) resulted in delayed repair and dampened intracellular Ca 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 response. Results suggest both extracellular and intracellular Ca sources are essential for supplying the Ca mobilization that stimulates repair.