Figure 1. Characterization of primary hippocampal cultures and concentration-response curves.

(A) Hippocampal cultures harvested from the postnatal rat brain were immunocytochemically stained on day in vitro 7 (DIV7) for the neuronal marker microtubule-associated protein 2 (MAP2, green) and the astrocyte marker glial fibrillary acidic protein (GFAP, red). Nuclei were stained blue with the Hoechst reagent. Photomicrographs were captured using a 20× objective on an epifluorescent microscope. (B–M) Primary hippocampal cultures were treated with MG132 or paraquat on DIV5 (1^st hit) or DIV6 (2^nd hit). Viability was assayed on DIV7 by (B, D, F, H) the In-Cell Western assay for MAP2 levels and (C, E, G, I) the Cell Titer Glo luminescent assay for ATP. Representative MAP2 In-Cell Western images are shown in panels J–M. Shown are the mean and SEM of 3–4 independent experiments, each performed in triplicate. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 versus 0 μM MG132 or paraquat, one-way ANOVA followed by Bonferroni post hoc correction.