Figure 3

Increasing levels of CREB protects against apoptosis and excitotoxicity. (A) Analysis of cell death in uninfected hippocampal neurons and in hippocampal neurons infected with rAAVs-hrGFP, rAAV-CREB or rAAV-mCREB. CREB and mCREB were detected with the M2 Flag antibody; hrGFP was detected with an antibody to hrGFP. Cell death was determined by analyzing Hoechst stained nuclei with shrunken cell body and large round chromatin clumps. For mCREB expression we obtained very weak Flag immunoreactivity in the death-inducing conditions, which is most likely due to compromised cell health following mCREB expression. Representative images are shown. Scale bar, 10 μm. Quantitative analysis of cell death induced by 10 nM staurosporine (B), growth factor withdrawal (C), or bath application of 20 μM NMDA (D). Bars represent means ± SEM (n = 4). Statistical significance was determined by Two-way ANOVA, Bonferroni post hoc test; statistically significant differences are indicated (***p < 0.001, **p < 0.01, *p < 0.05 compared to rAAV-hrGFP infected neurons in groups induced to undergo cell death; ###p < 0.001, ##p < 0.01 compared to rAAV-hrGFP infected neurons in untreated groups).