The specific Na(+)/Ca(2+) exchange inhibitor SEA0400 prevents nitric oxide-induced cytotoxicity in SH-SY5Y cells

The Na(+)/CA(2+) exchanger (NCX) plays a role in the regulation of intracellular CA(2+) levels, and nitric oxide (NO) is involved in many pathological conditions including neurodegenerative disorders. We have previously found that sodium nitroprusside (SNP), an NO donor, causes apoptotic-like cell death in cultured glial cells via NCX-mediated pathways and the mechanism for NO-induced cytotoxicity is cell type-dependent. The present study examined using the specific NCX inhibitor 2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline (SEA0400) whether NCX is involved in NO-induced injury in cultured neuronal cells. The treatment of neuroblastoma SH-SY5Y cells with SNP resulted in Apoptosis and the cytotoxicity was blocked by the mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK) kinase inhibitor U0126 and the p38 MAP kinase (MAPK) inhibitor SB203580, but not by the c-Jun N-terminal kinase (JNK) inhibitor SP60012. SNP increased CA(2+) influx and intracellular CA(2+) levels. In addition, SNP increased ERK and p38 MAPK phosphorylation, and production of Reactive Oxygen Species (ROS) in an extracellular CA(2+)-dependent manner. These effects of SNP were prevented by SEA0400. SNP-induced cytotoxicity was not affected by inhibitors of the CA(2+), Na(+) and store-operated/capacitative channels. Moreover, SNP-induced increase in intracellular CA(2+) levels, ROS production and decrease in cell viability were blocked by a cGMP-dependent protein kinase (PKG) inhibitor. These results suggest that CA(2+) influx via the reverse of NCX is involved in the cascade of NO-induced neuronal Apoptosis and NO activates the NCX through Guanylate Cyclase/PKG pathway.