Figure 2.

EP1 receptor blockade and knockout reduces hemin neurotoxicity. Primary postnatal neurons from WT and EP1^−/− pups cultured in serum-free Neurobasal media were treated with hemin (12.5–100 μM) for 18 h. (A) Hemin-induced cell viability was measured by LDH assay [LDH/LDH_Max %] from WT and EP1^−/− neurons. ^***p < 0.001 vs. control; ^#p < 0.01 and ^##p < 0.001 compared to the effect of hemin (50 and 75 μM) in neurons cultured from EP1^−/− pups. (B) Cultured WT and EP1^−/− neurons were treated with vehicle control or hemin (50 and 75 μM) for 18 h and phase-contrast images were captured using a 20× DIC objective. (C) Cultured WT neurons were pre-treated with EP1 receptor antagonist, SC51089 (10 μM), COX-2 antagonist, NS-398 (10 μM) and NMDAR antagonist, MK-801 (10 μM) for 15 min and were then co-treated with hemin (75 μM). After 18 h of treatment, cell viability was measured using LDH assay. LDH assay data represent means ± SEM of duplicate measures from triplicate wells from four to five experiments. Statistical analysis was carried out using One-Way ANOVA, with Bonferroni's multiple comparison tests. ^***p < 0.001 vs. control; ^*p < 0.05 vs. hemin alone.