Phenytoin-Induced Elevation of the Intracellular Calcium Concentration by Stimulation of Calcium-Sensing Receptors in Gingival Fibroblasts

Background: The mechanism concerning gingival overgrowth as a side effect of phenytoin, a therapeutic drug for epilepsy has been still unclear. As one of mechanisms, by measuring the intracellular calcium concentration ([Ca]i) of the gingival fibroblasts, it has been advocated that there is relationship between gingival overgrowth and phenytoin-induced alterations in the [Ca]i in gingival fibroblasts. To confirm that phenytoin elevates the [Ca]i, and if so, to find out its mode of action. Methods: The [Ca]i was measured with the Ca-sensitive fluorescent dye fura-2/AM. Cells were soaked in a flexiperm chamber and perfused by a saline. Drugs at appropriate concentrations were added to the perfusate. Results: Phenytoin concentration-dependently elevated the [Ca]i. NPS2390, a calcium-sensing receptor (CaSR) blocker, significantly suppressed the phenytoin-induced [Ca]i elevation. U73122, a phospholipase C (PLC) inhibitor, inihibited the phenytoin-induced [Ca]i elevation. TMB-8, a blocker of inositol triphophate (IP3) receptors in ER, significantly depressed the phenytoin-induced [Ca]i elevation. m-3M3FBS, a PLC activator, enhanced the phenytoin-induced [Ca]i elevation. From the findings obtained, it is discussed as follows: The Ca-free saline and NPS2390, a CaSR antagonist, inhibited the phenytoin-induced [Ca]i rise; These results indicate that CaSRs exist in gingival fibroblasts and that CaSRs are involved in the phenytoin-induced [Ca]i rise; U73122 and TMB-8 depressed the phenytoin-induced [Ca]i elevation and furthermore, m-3M3FBS enhanced the phenytoin-induced [Ca]i elevation, showing that the Ca release from the ER is involved in the phenytoin-induced [Ca]i elevation. Conclusion: We have concluded that phenytoin elevates the [Ca]i by activating CaSRs and enhancing the Ca release from the Ca stores in gingival fibroblasts.