Figure 1.

Calpain activity is required for synaptic NMDAR-dependent neuroprotection against starvation and oxidative stress. A, Trophic deprivation for 3 d induced cultured cortical neuron death. Incubation with 20 μm Bic and 100 μm 4-AP during starvation period (Bic + 4-AP) or before starvation (Bic + 4-AP, then MK801 + TTX) reduced neuronal death. Coapplication of 10 μm CI-III with Bic and 4-AP blocked the neuroprotective effect of Bic and 4-AP. Incubation of neurons under either normal condition or starvation with 10 μm CI-III alone for 3 d did not cause cell death. Neuronal death was observed and quantified by Hoechst staining; 300–500 cells were counted for each group in each independent experiment. *p < 0.05; ns, not significantly different; one-way ANOVA followed by Bonferroni test. n = 3. Error bar indicates SEM. B, Similar experimental design as in A except that neuronal death was induced by a 24 h-H₂O₂ insult (20 μm). In all cases, results are means ± SEM of three independent experiments with 300–500 cells for each group in each independent experiment. *p < 0.05; ns, not significantly different; one-way ANOVA followed by Bonferroni test. C, Representative photos of Hoechst staining. Hoechst brightly stains condensed and/or fragmented nuclei of apoptotic neurons but only dimly stains normal nuclei of healthy neurons. Scale bar, 10 μm.