Fig. 4. The reduced thickness of neuronal layers in the Dicer-ablated E13.5 dorsal telencephalon is not due to decreased cell cycle progression or division of apical and basal progenitors. (A,B,D,E,G,H) Immunofluorescence microscopy of 10-µm coronal cryosections through the dorsal telencephalon of control (Emx1^Cre/wt Dicer^flox/wt) and conditional Dicer knockout (Dicer KO, Emx1^Cre/wt Dicer^flox/flox) E12.5 (A,B), E13.5 (D,E) and E14.5 (G,H) littermate mouse embryos, showing phosphohistone H3 (PH3, red) and DAPI (white) staining. NL, neuronal layers; white arrows, mitotic apical progenitors; arrowheads, mitotic basal progenitors; yellow arrows, apoptotic nuclei. (C,F,I) Quantification of mitotic (phosphohistone H3-positive) apical and basal progenitors at E12.5 (C), E13.5 (F) and E14.5 (I). Data are the mean of 32 (C) or 28 (F,I) fields counted per condition from two embryos, eight (C) or seven (F,I) cryosections along the rostrocaudal axis per embryo, two fields per cryosection; bars indicate s.d. ^*P<0.05, ^**P<0.01. (J-L) Control (Emx1^Cre/wt Dicer^flox/wt; J,L black triangles) and conditional Dicer knockout (Dicer KO, Emx1^Cre/wt Dicer^flox/flox; K,L white diamonds) E12.5 littermate embryos were subjected to cumulative BrdU labeling in utero and analyzed after 4, 8 and 20 hours. (J,K) Triple immunofluorescence microscopy of 10-µm coronal cryosections through the dorsal telencephalon after 4 hours of cumulative BrdU labeling, showing BrdU (red), Tbr1 (green) and DAPI (blue) staining. NL, neuronal layers. (L) Quantification of BrdU-positive (BrdU+), Tbr1-negative nuclei in the VZ (immunostained as in J,K), expressed as percentage of DAPI-stained nuclei. Data are the mean of two embryos; for each embryo, the average for three fields along the rostrocaudal axis (one field per cryosection) was calculated; bars indicate the variation of the two embryos from the mean (and are often smaller than the size of the symbol used). Horizontal dashed lines indicate the growth fraction. Vertical dotted lines indicate T_C-T_S (control 10.5 hours, Dicer KO 11.0 hours); T_S (length of S phase) was 2.4 hours for control and 3.4 hours for Dicer KO, and T_C (total length of the cell cycle) was 12.9 hours for control and 14.4 hours for Dicer KO. Scale bars: 50 µm.