Dual actions of nitric oxide in N-methyl-D-aspartate receptor-mediated neurotoxicity in cultured retinal neurons

This study was performed to elucidate the role of nitric oxide (NO) in N-methyl-D-aspartate (NMDA) receptor-mediated glutamate neurotoxicity in the retina. The experiments were done with primary retinal cultures obtained from 17- to 19-day-old rat fetuses. The NOS activity measured by monitoring the conversion of [H-3]arginine to [H-3]citrulline was approximately 5 pmol/min/mg protein. A 10-min exposure of the cultured cells to glutamate (1 mM) or NMDA (1 mM) followed by a 1-h incubation in a normal medium consistently resulted in 60% cell death. The concomitant addition of an inhibitor of NOS, N-omega-nitro-L-arginine (300 mu M), with glutamate or NMDA reduced cell death by 70%. A brief exposure of the cells to sodium nitroprusside (SNP, 500 mu M) or S-nitrosocysteine (SNOC, 500 mu M), NO-generating agents, caused 60% cell death. Depletion of NO by reduced hemoglobin prevented the cell death induced by either glutamate, NMDA, or NO generating agents. Fifty mu M SNOC alone had no effect on the cell viability. However, pretreatment with 50 mu M SNOC as well as simultaneous application of 50 mu M SNOC with NMDA inhibited cell death induced by NMDA. These findings indicate that a low concentration of NO plays a protective role in glutamate neurotoxicity via closing the NMDA receptor gated ion channel. However, elevated concentrations of NO, interacting with oxygen radicals, become toxic and mediate glutamate-induced neurotoxicity in the cultured retinal neurons.