Abstract

The pretreatment of cultured cortical neurons with neurotrophic factors markedly potentiates the cytotoxicity induced by low concentrations of Zn²⁺ or excitotoxins. In the current study, we investigated the mechanism underlying the insulin-like growth factor-I (IGF-I)-induced Zn²⁺ toxicity potentiation. The pretreatment of primary cortical cultures for more than 12 h with 100 ng/ml of IGF-I increased the cytotoxicity induced by 80 μM Zn²⁺ by more than 2-fold. The IGF-I–enhanced cell death was blocked by the COX-2–specific inhibitors N-[2-(cyclohexyloxyl)-4-nitrophenyl]-methane sulfonamide (NS-398; 10–100 μM) and 1-[(4-methylsulfonyl)phenyl]-3-trifluoro-methyl-5-[(4-fluoro)phenyl]pyrazole (SC58125; 10 μM) and by the antioxidant trolox (30 μM). In addition, it was observed that COX-2 expression was increased 12 to 24 h after IGF-I treatment. Preincubation of cortical cultures with IGF-I increased arachidonic acid (AA)-induced cytotoxicity, and AA increased Zn²⁺ toxicity, which suggested the involvement of COX activity in these cellular responses. Moreover, enhanced COX-2 activity led to a decrease in the cell's reducing power, as indicated by a gradual depletion of intracellular GSH. Cortical neurons pretreated with IGF-I and then Zn²⁺ showed consistently enhanced reactive oxygen species production, which was repressed by NS-398 and SC58125. Cortical neurons treated with Zn²⁺ and then AA displayed the increased ROS production, which was also suppressed by NS-398 and SC58125. These results suggest that COX-2 is an endogenous factor responsible for the IGF-I–induced potentiation of Zn²⁺ toxicity and that enhanced COX-2 activity leads to a decrease in the cell's reducing power and an increase in ROS accumulation in primary cortical cultures.