Fig. 4

FZD1 knockdown reduces the generation of newborn granule neurons in the dentate gyrus. a Schematic representation of the experimental procedure. Retroviruses expressing shRNAs (shC or shFZD1) and ZsGreen (ZsG) were injected into the dentate gyrus of 2-month-old mice by stereotaxic surgery. Animals were sacrificed 1, 2 or 4 weeks post injection (wpi) to analyze neuronal fate commitment (1 and 2 wpi), generation and morphology of newborn granule neurons (4 wpi). b Quantification of the percentage of ZsG+ cells expressing SOX2/GFAP and the immature neuronal marker DCX 1 wpi. Bars represent mean ± S.E. *p < 0.05, Student’s t-test (N ≥ 4 mice). c Immunostaing of DCX and NeuN in brain sections from animals sacrificed 2 wpi. Images at the right show separated channels of a section of the images shown at the left. Dotted circles indicate transduced cells (ZsG+) in the merged images and in the separated channels. Scale bar: 20 μm. d Quantification of the percentage of ZsG+ cells expressing DCX at wpi. Bars represent mean ± S.E. *p < 0.05, Student’s t-test (N ≥ 3 mice). e Immunostaing of DCX and NeuN in brain sections from animals sacrificed 4 wpi. Images at the right show separated channels of a single z-plane section of the images shown at the left. Dotted circles indicate transduced cells (ZsG+) in the merged images and in each channel. Scale bar: 20 μm. f Quantification of the percentage of ZsG+ cells expressing the mature neuronal marker NeuN at 4 wpi. Bars represent mean ± S.E. *p < 0.01, Student’s t-test (N ≥ 3 mice). g Representative ZsG+ astrocytes and radial glia-like NSCs expressing GFAP. Scale bar: 20 μm. h Quantification of the percentage of ZsG+ cells that show astrocyte or NSCs morphology at 4 wpi. Bars represent mean ± S.E. *p < 0.01, Student’s t-test (N = 4 mice). ML, molecular layer; GCL, granule cell layer; SGZ, subgranular zone