Peptide growth factors protect against ischemia in culture by preventing nitric oxide toxicity

Reduction or elimination of nitric oxide (NO) production in cortical neurons by NO synthase (NOS) inhibitors during glutamate toxicity in vitro or during focal cerebral ischemia in vivo can prevent neuronal cell death. In contrast, growth factors can prevent neuronal degeneration induced by treatment with glutamate or potassium cyanide. We have determined whether NO mediates hippocampal cell death during anoxia in vitro and whether the peptide growth factors basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) can prevent hippocampal neuronal death during anoxia or NO exposure. Both bFGF and EGF increased hippocampal neuronal survival from about 35% in anoxic cultures to about 65% in treated cultures during an 8 hr period of anoxia. Inhibition of NOS by NG-monomethyl-L-arginine, a competitive inhibitor of NOS, rescued 65-70% of the neurons that would normally die during an 8 hr anoxic incubation, and this effect was reversed by L-arginine, a precursor for NO. Thus, hippocampal neuronal death following anoxia is, at least in part, mediated by NO. NO, generated by either nitroprusside or 3-morpholino-sydnonimine, was toxic to hippocampal neurons. Pretreatment of cultures with either bFGF (10 ng/ml) or EGF (10 ng/ml) prior to NO exposure increased survival from approximately 40% in untreated cultures to 80% in treated cultures, yet the effect of combining bFGF and EGF was not greater than treatment with either of the growth factors alone. Knowledge that the growth factors bFGF and EGF are neuroprotective against NO toxicity provides insights into the mechanisms of ischemic neuronal death that may direct future therapeutic modalities for cerebrovascular disease and neurodegenerative disorders.