The Na +/Ca 2+ exchanger-mediated Ca 2+ influx triggers nitric oxide-induced cytotoxicity in cultured astrocytes

Abstract

Nitric oxide (NO) is involved in many pathological conditions including neurodegenerative disorders. We have previously found that sodium nitroprusside (SNP), an NO donor, stimulates mitogen-activated protein kinases (MAPKs) such as extracellular signal-regulating kinase (ERK), c-jun N-terminal protein kinase (JNK) and p38 MAPK, leading to caspase-independent apoptosis in cultured astrocytes. In view of the previous observation that NO stimulates the activity of the Na +/Ca 2+ exchanger (NCX), this study examines the involvement of NCX in cytotoxicity. The specific NCX inhibitor SEA0400 blocked SNP-induced phosphorylation of ERK, JNK and p38 MAPK, and decrease in cell viability. SNP-induced phosphorylation of ERK, JNK and p38 MAPK was blocked by removal of external Ca 2+, and SNP treatment caused an increase in 45Ca 2+ influx. This increase in 45Ca 2+ influx was blocked by SEA0400, but not the Ca 2+ channel blocker nifedipine. In addition, SNP-induced 45Ca 2+ influx and cytotoxicity were reduced in NCX1-deficient cells which were transfected with NCX1 siRNA. Inhibitors of intracellular Ca 2+-dependent proteins such as calpain and calmodulin blocked SNP-induced ERK phosphorylation and decrease in cell viability. Furthermore, the guanylate cyclase inhibitor LY83583 and the cGMP-dependent protein kinase inhibitor KT5823 blocked SNP-induced cytotoxicity. These findings suggest that NCX-mediated Ca 2+ influx triggers SNP-induced apoptosis in astrocytes, which may be mediated by a cGMP-dependent pathway.