Role for protein phosphatase in the regulation of Ca2+ influx in parotid gland acinar cells.

Stimulation of Ca2+ (and Mn2+) entry in salivary epithelial cells by carbachol, or thapsigargin, is mediated by an, as yet, unknown mechanism that is dependent on the depletion of Ca2+ from intracellular Ca2+ stores. This study assesses the possible role of protein phosphorylation in the regulation of Ca2+ entry in rat parotid gland acinar cells. Treatment of cells with the protein phosphatase inhibitors okadaic acid, calyculin A, and pervanadate induced a dose-dependent inhibition of carbachol and thapsigargin stimulation of Ca2+ and Mn2+ entry. All three inhibitors decreased carbachol stimulation of internal Ca2+ release, which likely accounts for the inhibition of carbachol-stimulated Ca2+ entry. Thapsigargin-induced internal Ca2+ release was not affected by the treatments. Additionally, all three phosphatase inhibitors decreased Mn2+ entry into cells with depleted internal Ca2+ store(s) (achieved by incubation with either carbachol or thapsigargin in Ca2+-free medium). Treatment of cells with phorbol 12-myristate 13-acetate, 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, or staurosporine did not affect divalent cation entry into unstimulated cells or thapsigargin treated cells. Importantly, when cells with depleted internal Ca2+ store(s) were pretreated with staurosporine, or K-252a, the inhibition of Ca2+ entry by calyculin A and okadaic acid, but not by pervanadate, was attenuated. Although the effect of pervanadate remains to be clarified, these results demonstrate a role for protein phosphorylation in the regulation of divalent cation influx in rat parotid acinar cells.