Cholinergic-induced Ca2+ elevation in rat lacrimal gland acini is negatively modulated by PKC delta and PKC epsilon

PURPOSE. To investigate the role of protein kinase C (PKC) in cholinergic a gonist-induced Ca2+ elevation in lacrimal gland acini. METHODS. Lacrimal gland acini were prepared by collagenase digestion, and c hanges in intracellular Ca2+ ([Ca2+](i)) were measured using fura-2 as a fl uorescent probe. RESULTS. Preactivation of PKC by phorbol 12-myristate 13-acetate (PMA), or inhibition of protein phosphatase type 1/2A (PP1/2A) by calyculin A, decrea sed both the [Ca2+](i) transient and the plateau of [Ca2+](i) induced by in creasing concentrations of carbachol, a cholinergic agonist. Staurosporine, an inhibitor of PKC, completely reversed the effect of PMA. Inhibition of the Ca2+-independent PKC isoforms PKC delta and -epsilon but not the Ca2+-d ependent isoform PKC alpha substantially reversed the inhibitory effect of PMA on cholinergic agonist-induced Ca2+ elevation. The inhibitory effect of PMA was obtained only in the presence of extracellular Ca2+, suggesting th at PKC inhibits the influx of Ca2+. PMA completely inhibited the cholinergi c agonist-induced plateau of [Ca2+](i). PMA and calyculin A decreased both the [Ca2+](i) transient and the plateau of [Ca2+](i) induced by thapsigargi n, further supporting the idea that PKC modulates the entry of Ca2+. CONCLUSIONS. In the lacrimal gland, agonist-induced changes in [Ca2+](i) ar e negatively regulated by PKC-dependent phosphorylation of a target protein (s) that is sensitive to PP1/2A.