Figure 3: Hes5∷CreER^T2-labeled NSCs generate neuroblasts after 21 days.

(a) TAM induction regime once per day for 5 consecutive days before the mice were killed (*) at 7 days (d7) or 21 days (d21) after the end of induction. Quantification of labeled cells on d7 and d21. At d7, the majority are still NSCs, whereas the first rGFP⁺NeuN⁺ neurons start to appear at d21. (b) The majority of the labeled (rGFP⁺) cells at d21 still have NSC morphologies and expressed NSC markers, including GFAP. (c) BrdU induction regime. BrdU was administered once at day 0 (d0) and the mice were killed (*) 1 day (d1), 3 days (d3), or 7 days (d7) later. Quantification of Ascl1⁺ cells labeled with BrdU on d1, d3 and d7. The proportion of BrdU-labeled Ascl1⁺ cells decreased over time. (d) Some Ascl1⁺ cells incorporated BrdU 2 h after induction. (e) At d1 after BrdU application, clusters of BrdU-labeled cells appear, which included a single Ascl1⁺ cell and Dcx⁺ neuroblasts. (f) Graphical representation of the fractions of Ascl1⁺ and Dcx cells among the BrdU-incorporating cell populations. The proportion of the BrdU-labeled cells that were Ascl1⁺ type-2a IPs decreased over time, whereas Dcx⁺ neuroblasts increased. (g) rGFP⁺ cells remain in the SGZ and are in close proximity to Dcx⁺ neuroblasts. NeuN labels mature neurons in the DG granule cell layer. (h) rGFP⁺Dcx⁺ neuroblasts are prominent in the SGZ by d21. (i) A proportion of the rGFP⁺ cells are postmitotic differentiated neurons (arrow). A small population of Dcx⁺ neuroblast remains unlabeled (arrowhead). (j) Tbr2⁺Dcx⁺ early neuroblasts were clustered in close proximity to Ascl1⁺ IPs. (k) Tbr2⁺ cells were proliferating (PCNA⁺) in the adult SGZ, but rGFP⁺Tbr2⁺ cells were rare at d1 after TAM induction. (l) At d21 after TAM induction, recombined rGFP⁺Tbr2⁺ were present in the SGZ and proliferating (PCNA⁺). GCL, granule cell layer; ML, molecular layer; SGZ, subgranular zone; TAM, tamoxifen. Error bars are s.d. Scale bars equals 20 μm, and in (g), equals 100 μm.