ROLE FOR PROTEIN PHOSPHATASE IN THE REGULATION OF CA2-GLAND ACINAR-CELLS( INFLUX IN PAROTID)

Stimulation of Ca2+ (and Mn2+) entry in salivary epithelial cells by c arbachol, or thapsigargin, is mediated by an, as yet, unknown mechanis m that is dependent on the depletion of Ca2+ from intracellular Ca2+ s tores. This study assesses the possible role of protein phosphorylatio n in the regulation of Ca2+ entry in rat parotid gland acinar cells. T reatment 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 thr ee 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 b y the treatments. Additionally, all three phosphatase inhibitors decre ased Mn2+ entry into cells with depleted internal Ca2+ store(s) (achie ved by incubation with either carbachol or thapsigargin in Ca2+-free m edium). Treatment of cells with phorbol 12-myristate 13-acetate, 1-(5- isoquinolinylsulfonyl)-2-methylpiperazine, or staurosporine did not af fect divalent cation entry into unstimulated cells or thapsigargin-tre ated cells. Importantly, when cells with depleted internal Ca2+ store( s) were pretreated with staurosporine, or K-252a, the inhibition of Ca 2+ 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.