Data_Sheet_1_Evidence That Calcium Entry Into Calcium-Transporting Dental Enamel Cells Is Regulated by Cholecystokinin, Acetylcholine and ATP.PDF

<p>Dental enamel is formed by specialized epithelial cells which handle large quantities of Ca<sup>2+</sup> while producing the most highly mineralized tissue. However, the mechanisms used by enamel cells to handle bulk Ca<sup>2+</sup> safely remain unclear. Our previous work contradicted the dogma that Ca<sup>2+</sup> is ferried through the cytosol of Ca<sup>2+</sup>-transporting cells and instead suggested an organelle-based route across enamel cells. This new paradigm involves endoplasmic reticulum (ER)-associated Ca<sup>2+</sup> stores and their concomitant refilling by store-operated Ca<sup>2+</sup> entry (SOCE) mediated by Ca<sup>2+</sup> release activated Ca<sup>2+</sup> (CRAC) channels. Given that Ca<sup>2+</sup> handling is maximal during the enamel-mineralization stage (maturation), we anticipated that SOCE would also be elevated then. Confirmation was obtained here using single-cell recordings of cytosolic Ca<sup>2+</sup> concentration ([Ca<sup>2+</sup>]<sub>cyt</sub>) in rat ameloblasts. A candidate SOCE agonist, cholecystokinin (CCK), was found to be upregulated during maturation, with Cck transcript abundance reaching 30% of that in brain. CCK-receptor transcripts were also detected and Ca<sup>2+</sup> imaging showed that CCK stimulation increased [Ca<sup>2+</sup>]<sub>cyt</sub> in a dose-responsive manner that was sensitive to CRAC-channel inhibitors. Similar effects were observed with two other SOCE activators, acetylcholine and ATP, whose receptors were also found in enamel cells. These results provide the first evidence of a potential regulatory system for SOCE in enamel cells and so strengthen the Ca<sup>2+</sup> transcytosis paradigm for ER-based transport of bulk Ca<sup>2+</sup>. Our findings also implicate enamel cells as a new physiological target of CCK and raise the possibility of an auto/paracrine system for regulating Ca<sup>2+</sup> transport.</p>