Fig. 2
Disruption of GC oscillations in 3-month-old Nestin-GFP mice induces reversible NSPC quiescence. a Representative example of Nestin-GFP (green), GFAP (white) and Ki67 (red) immunoreactivity in the DG of 3-month-old Nestin-GFP mice. The boxed area shows a cluster of Nestin-GFP+/GFAP+/Ki67+ NSPC. b Magnification of area boxed in (a). b’ Same area further magnified with channels split and Z-stacked, showing the expression of individual markers. Arrow: cell soma. b” The dashed white line shows a transversal cell section. c Histogram of the transversal section in (a”), showing fluorescent intensity signals for DNA (blue), GFP (green), GFAP (black), and Ki67 (red). Representative Z-stacked confocal images of NSPC in the DG of mice treated for 7 days with d 0, e 12.5, f 25 mg/kg/day [CORT] pellets or allowed to recover for 2 days after removal of a g 12.5 and h 25 mg/kg/day [CORT] pellet. Arrows: Type-1 cells; arrowheads: Type-2a/2b cells. Scale bars represent 20 μm (a, b and d, h), 15 μm (b’) and 10 μm (b”). i AM and PM plasma CORT levels after the treatments indicated in the graph legends. Bars are mean ± SEM [CORT] and red circles individual data-points (animals). Statistical comparisons were done using one-way analysis of variance test with Tukey’s post hoc test for multiple comparisons (n = 4 mice, ***p < 0.001, AM vs. PM in 0 mg/kg/day, ns p > 0.05, AM vs. PM in both 12.5 and 25 mg/kg/day and after 2 day recovery; #p < 0.001, 25 mg/kg/day AM and PM vs. 0 mg/kg/day AM and PM, respectively; $p < 0.05, 25 mg/kg/day AM and PM vs. 12.5 mg/kg/day AM and PM, respectively; §p < 0.001, both AM and PM in 12.5 and 25 mg/kg/day vs. 2-day recovery). j Percentages of Ki67+ (full bars and full circles) or Ki67− (dashed bars and open circles) of Type-1 (red), −2a (green) and −2b (blue) NSPC, 7 days postimplantation with 0, 12.5, 25 mg/kg/day [CORT] pellets and 25 mg/kg/day [CORT] +2-day recovery. Ki67+ NSPC were not observed in animals treated with [CORT] 12.5 and 25 mg/kg/day. Bars are mean ± SEM and circles individual data points (animals) (n = 4 mice, *p < 0.05, ***p < 0.001, vs. 0 mg/kg/day, one-way ANOVA or ###p < 0.001, 7-day treatment vs. 7-day treatment +2-day recovery with the same [CORT], one-way ANOVA). Further information in Fig. S4A. ML molecular layer, SGZ subgranular zone, GCL granule cell layer
