Characterization of EPT and D89E mice used in this study. (A) Representative images of the V-SVZ, RMS, and OB of adult Dcx-CreER^T2; R26-tdTomato (control) mice at 3 dpi (n = 3 mice) stained for Dcx, DsRed, and NeuN. tdTomato is expressed in the subset of Dcx⁺ cells in the V-SVZ-RMS (10.2 ± 1.4%, n = 3 mice pooled from two independent experiments), and OB (0.7 ± 0.3%, n = 3 mice). These tdTomato⁺ cells in the V-SVZ-OB were positive for Dcx (85.1 ± 0.8%, n = 3 mice) or NeuN (14.8 ± 0.6%, n = 3 mice), but negative for GFAP or Iba1, suggesting that some of the tdTomato⁺ Dcx⁺ cells were differentiated into tdTomato⁺ NeuN⁺ OB neurons within 3 d. (B) Representative images of the GCL of the OB in the adult control, Dcx-CreER^T2; R26-EPT; R26-tdTomato (EPT), and Dcx-CreER^T2; R26-D89E; R26-tdTomato (D89E) mice at 28 dpi (n = 3 mice each, pooled from six independent experiments) stained for flag (white), Iba1 (cyan), and GFAP (blue). tdTomato⁺ signals (red) are directly observed without staining. Arrows indicate flag⁺ signals in tdTomato⁺ granule cells in the OB. (C) Representative images of the EPL of the OB in EPT and D89E mice at 28 dpi stained for DsRed (red; n = 3 mice each). Injected PSVue480 is shown in white. (D) Proportion of PS⁺tdTomato⁺ spines in the EPL of EPT and D89E mice at 28 dpi (n = 3 mice each, pooled from three independent experiments; t₍₄₎ = −3.0, P = 0.042, unpaired t test). (E) Distributions (left graph, bars; and right graph) and their fitting curves (left graph, lines) of PS intensity in EPT and D89E mice at 28 dpi. Arrows and arrowheads (left graph) indicate the onset and peak of fitting curves in EPT (pink) and D89E (blue) mice (EPT, AIC = −55.2; D89E, AIC = −33.2; also see Materials and methods). While the onset of the fitting curve is similar between the two transgenic mice, its peak is shifted rightward in D89E mice (arrowheads, EPT, 8.8; D89E, 10.5), suggesting that PS is initially exposed on spines similarly in EPT and D89E mice, but accumulated in D89E mice due to the failure of the microglial spine pruning. PS intensity at tdTomato⁺ spines in D89E mice (n = 24 events from three mice) is significantly higher than that in EPT mice (n = 22 events from three mice, pooled from three independent experiments; t₍₄₄₎ = −2.0, P = 0.048, unpaired t test). (F–H) No effect of MFG-E8^EPT and MFG-E8^D89E on the migration of tdTomato⁺ cells toward the OB. Representative images (F; n = 3 mice each) and proportions (G and H; pooled from four independent experiments) of tdTomato⁺ cells (red in F) in the GL, GCL, anterior RMS (aRMS), and posterior RMS (pRMS) of control, EPT, and D89E mice at 3 (F and G) and 14 (F and H) dpi (n = 3 mice each) are shown. (I and J) No effect of MFG-E8^EPT and MFG-E8^D89E on the number of Iba1⁺ microglia in the adult OB. Representative images (I; n = 3 mice each) and numbers (J; pooled from six independent experiments) of Iba1⁺ cells (green in I) in the EPL and GCL of the OB in control, EPT, and D89E mice (n = 3 mice each) at 28 dpi are shown. RMS, rostral migratory stream; GCL, granule cell layer; EPL, external plexiform layer; GL, glomerular layer; AU, arbitrary unit. Scale bars: A and I, 20 µm; B, 5 µm; C, 1 µm; F, 50 µm. *, P < 0.05. Data shown are mean ± SEM.