Calcium handling in porcine coronary endothelial cells by gastrin-17

In porcine coronary artery endothelial cells (PCAEC), gastrin-17 has recently been found to increase nitric oxide (NO) production by the endothelial NO synthase (eNOS) isoform through cholecystokinin 1/2 (CCK1/2) receptors and the involvement of protein kinase A (PKA), PKC and the b₂-adrenoreceptor-related pathway. As eNOS is the Ca^2C-dependent isoform of the enzyme, we aimed to examine the effects of gastrin-17 on Ca^2C movements. Thus, experiments were performed in Fura-2-acetoxymethyl-ester-loaded PCAEC, where changes of cytosolic Ca^2C ([Ca^2C]_c) caused by gastrin-17 were analysed and compared with those of CCK receptors and b₂-adrenoreceptors agonists/antagonists. In addition, some experiments were performed by stimulating cells with gastrin-17 in the presence or absence of cAMP/PKA activator/inhibitor and of phospholipase C (PLC) and Ca^2C-calmodulin dependent protein kinase II (CaMKII) blockers. The results have shown that gastrin-17 can promote a transient increase in [Ca^2C]_c mainly originating from an intracellular pool sensitive to thapsigargin and from the extra cellular space. In addition, the response of cells to gastrin-17 was increased by the adenylyl cyclase activator and the b₂-adrenoreceptor agonists and affected mainly by the CCK2 receptor agonists/antagonists. Moreover, the effects of gastrin-17 were prevented by b₂-adrenoreceptors and CaMKII blockers and the adenylyl cyclase/PKA and PLC inhibitors. Finally, in PCAEC cultured in Na^C-free medium or loaded with the plasma membrane Ca^2C pump inhibitor, the gastrin-17-evoked Ca^2C transient was long lasting. In conclusion, this studyshowsthatgastrin-17affected intracellularCa^2Chomeostasisin PCAEC by both promoting a discharge of an intracellular pool and by interfering with the operation of store-dependent channels through mainly CCK2 receptors and PKA/PLC- and CaMKII-related signaling downstream of b₂-adrenoreceptor stimulation. ^© 2013 Society for Endocrinology.