Fig. 3.
Genetic crosses of ne-Emx2 transgenic and Emx2+/− heterozygous mice restore cortical Emx2 expression to wt levels and restore cortical area sizes toward wt sizes. (A–D) Tangential sections through layer 4 of flattened mouse cortices at postnatal day 7, stained for cytochrome oxidase to reveal area patterning landmarks. Sections are from four different genotypes that express different levels of EMX2 encoding transcripts in progenitor cells of the cortex during development (see Fig. 2B). (A) Wild type. (B) ne-Emx2+/+ transgenics. (C) Emx2+/− heterozygotes. (D) ne-Emx2+/+; Emx2+/− double mutants. CO staining patterns reveal differences in sizes and positions of cortical areas between genotypes. Horizontal lines across the sections support visualizing those changes. M, motor cortex; S, somatosensory cortex; PMBSF, posteromedial barrel subfield of somatosensory cortex; V, visual cortex. For a schematic of the area size changes, see Fig. 2A; for quantification, see F–I. (E) Schematic of flattened cortical hemisphere: indicated area landmarks, as revealed by CO and serotonin staining (serotonin sections not shown), were used for the area measurements and quantifications shown in F–I. Gray shaded area marks nonneocortical regions (olfactory bulb, rhinal cortex). (F–I) Histograms of cortical sizes, cortical area ratios, and cortical length ratios at postnatal day 7 of wt, TG (ne-Emx+/+), E+/− (Emx2+/−), and TG;E+/− (ne-Emx+/+; Emx2+/−) mice. Schematics below histograms relate to the schematic in E and indicate the specific measurements performed. (F) Overall surface area of neocortex in millimeters. Overall surface areas are not significantly different from wt (wt: 35.3 ± 2.1, n = 17; TG: 32.9 ± 0.9, n = 13; E+/−: 34.0 ± 1.1, n = 9; TG;E+/−: 32.6 ± 0.7, n = 13). All P values are >0.14 (unpaired Student's t test). (G) Frontal length ratio, defined as the ratio of FL (length between the rostral edge of PMBSF and the rostral pole of neocortex) to TL (total length of neocortex from rostral pole to occipital pole). Compared with wt (n = 13), the frontal ratio is significantly decreased in TG (−15 ± 0.8%∗∗, n = 12), significantly increased in E+/− (+9 ± 1.42%∗∗, n = 8), and reverts toward wt in TG;E+/− (−6 ± 1.17%∗, n= 11). (H) PMBSF area ratio, defined as the ratio of PA (area of PMBSF) to TA (area of the entire neocortex). Compared with wt (n + 8), the PMBSF ratio is significantly decreased in TG (−21 ± 2.3%∗∗, n = 6), significantly increased in E+/− (+11 ± 3.7%∗∗, n = 8), and reverts toward wt in TG;E+/− (−6 ± 2.6%, P = 0.0771; n = 8). (I) Frontal area ratio, defined as the ratio of MA [area covered by the rostral (motor) cad8 expression domain] to TA (area of entire cortex). Compared with wt (n = 6), the size of the rostral motor domain is significantly reduced in TG (−34 ± 1.7%∗∗, n = 9), significantly increased in E+/− (+15 ± 1.5%∗∗, n = 6), and reverts towards wt in TG;E+/− (−18 ± 2.7%∗∗, n = 9). Measurements were performed on whole mounts of cad8 in situ hybridization without flattening of the hemisphere. (Scale bar in A for A–D: 1 mm.) Bars in the histograms represent the means, and error bars represent SEM. Symbols used in this legend indicate the following: %, percent increase or decrease relative to wt; ±, SEM; n, number of cases: ∗, P < 0.05; ∗∗, P < 0.01 (unpaired Student's t tests).