Fig. 2

Neonatal peripheral inflammatory pain-induced microglia activation and neuronal cell death in the cortex and hippocampus. Microglia cells and neuronal cell death were measured using Iba-1 and TUNEL staining, respectively, in the cortex and hippocampus. a, b Immunohistochemical staining was performed 1 day after the last formalin injection. Iba-1-positive cells (red) were identified as microglia cells in the cortex. Nuclei were stained with Hoechst 33342 (blue). Different shapes of Iba-1-positive microglia cells at resting state (control) and activation state (formalin) are shown in the insets of enlarged magnification. Scale bars = 100 μM. b The ratio of Iba-1+ cells against total Hoechst + cells. Formalin injections significantly increased activated microglia while indomethacin (Indo) attenuated this event. *P < 0.05 vs. control, ^# P < 0.05 vs. formalin, ANOVA plus Bonferroni’s correction; n = 6–8 per group. c TUNEL staining (green) and Hoechst 33342 (blue) were used to evaluate cell death in the cortex and hippocampus. Arrows point to some TUNEL-positive green color cells. Scale bars = 100 μm. d, e Quantified data of TUNEL-positive cells/field in the cortex (d) and hippocampus (e). *P < 0.05 vs. control; n = 6–7 per group. f TUNEL-positive cells/field in the hippocampus in saline control and formalin groups. A marked increase in TUNEL-positive cells was seen in CA2. *P < 0.05 vs. CA1 or CA3, ANOVA plus Bonferroni’s correction; n = 6–7 per group