Regulation of Ca2+ signals in a parotid cell line Par-C5

The Ca2+ signaling system in an established immortalized rat parotid acinar cell line, Par-C5, was examined using the Ca2+-sensitive fluorescent indic ator fura-2 and by measuring inositol 1,4,5-trisphosphate (IP3) formation. Agonist-induced increase in intracellular Ca2+ ([Ca2+](i)) by mobilization of intracellular stores and influx across the cell membrane was stimulated by acetylcholine (ACh) and ATP, whereas noradrenaline-(NA)-induced a small [Ca2+], increase mediated primarily by release from intracellular Ca2+ stor es. [Ca2+](i), increase by ACh and ATP was meditated through the phosphoino sitide signal pathway since both agonists significantly increased 1,4,5-IP3 formation and Ca2+ mobilization was abolished by the phospholipase C inhib itor U73122. In Ca2+-free medium, ACh or ATP discharged the IP3-sensitive C a2+ store and essentially abolished subsequent [Ca2+](i) response to thapsi gargin (TG). Exposure to ionomycin and monensin after TG induced a further mobilization of Ca2+, suggesting IP3-insensitive stores are present. Furthe rmore, depletion of IP3-sensitive Ca2+ stores by TG, ACh and ATP enhanced p lasmalemmal Ca2+-entry pathways. Exposure to tumor necrosis factor-alpha (T NF-alpha), a cytokine associated with lymphocyte invasion of salivary epith elial cells in autoimmune disorders, significantly reduced ACh-stimulated C a2+ mobilization. TNF-alpha inhibitory effect on Ca2+ mobilization was not directly due to an interaction on muscarinic receptors since ACh-induced 1, 4,5-IP3 formation was not altered. These results in the Par-C5 cell line in dicate 1) [Ca2+](i) is regulated by muscarinic and P2Y-nucleotide receptors and partly by alpha (1)-adrenergic receptors; 2) IP3-sensitive and -insens itive Ca2+ stores exist; 3) Ca2+ influx activated by ACh, ATP or TG is medi ated by the store-operated Ca2+ entry pathway; and 4) muscarinic agonist-st imulated Ca2+ mobilization is altered by the cytokine TNF-alpha. (C) 2001 E lsevier Science Ltd. All rights reserved.