Fig. 2: Protective effects of DR-Ab against excitotoxic injury in primary cultured cortical neurons.

a Concentration-dependent effects of glutamate on cell viability. Cells were treated with glutamate + glycine (10:1) for 24 h, n = 6/group. One-way ANOVA followed by Bonferroni’s test, **p < 0.01, ***p < 0.001 vs control group. b Neurons from E17 NKAα1^+/− mice exhibited lower cell viability when compared to those from NKAα1^+/+ mice when subjected to glutamate (50 µM) + glycine (5 μΜ) for 24 h, n = 6/group. Two-way ANOVA followed by Bonferroni’s test, **p < 0.01 with respect to NKAα1^+/+ group. c Concentration-dependent protective effect of DR-Ab against glutamate-induced neuron injury. Cells were pretreated with DR-Ab (0.075, 0.15, 0.3 mg/ml) or IgG (0.3 mg/ml) purified from normal rat serum 1 h before glutamate (100 µM) + glycine (10 μΜ) stimulation for 24 h. n = 6/group. One-way ANOVA followed by Bonferroni’s test, **p < 0.01, ***p < 0.001. d Pretreatment with peptide (40 µM, 1 h) used to raise the DR-Ab significantly attenuated the protective effect of DR-Ab, n = 6/group. One-way ANOVA followed by Bonferroni’s test, **p < 0.01, ***p < 0.001. e, f Representative DAPI immunostaining images (e) and group data (f) showing that DR-Ab reduced cell apoptosis caused by glutamate, n = 6/group. One-way ANOVA followed by Bonferroni’s test, ***p < 0.001. g DR-Ab decreased ROS production stimulated by glutamate treatment for 30 min, n = 6/group. One-way ANOVA followed by Bonferroni’s test, **p < 0.01, ***p < 0.001. h, i Western blots showing that DR-Ab inhibited glutamate-induced cytochrome c release (h), n = 7/group, and caspase 3 cleavage (i), n = 6/group. One-way ANOVA followed by Bonferroni’s test, *p < 0.05, **p < 0.01, ***p < 0.001. DR: DR-Ab, Glu: glutamate (100 µM) + glycine (10 μΜ) for 24 h, NS: IgG purified from normal rat serum