FIG. 3.

GFP expression within the telencephalon of Tg(Eomes::GFP) embryos at midgestation and early postnatal stages. (a) Onset of GFP fluorescence in the central nervous system (CNS) at E10.0. (b,c) Wholemount brightfield and GFP overlays depicting GFP fluorescence in the developing telencephalon at E12.5 (b) and E14.5 (c). Discrete foci of GFP fluorescence in both forelimbs and hindlimbs (white arrowheads). (d) By postnatal day zero (P0), GFP fluorescence was markedly reduced. (e–h) Fluorescent staining with Eomes antibody on vibratome sections revealed colocalization of GFP with Eomes protein, although the domain of GFP fluorescence was broader than that of the endogenous protein. (i–l) High magnification rendered confocal images of sectioned brains at E14.5 (boxed region in g) showing nuclear counterstain (i), GFP fluorescence (j), and Eomes protein expression (k). Three channel merge (l) reveals that GFP is not expressed in some cells with Eomes protein (yellow arrowhead) while other cells have both GFP and Eomes protein expression (white arrowhead). (m–p) Staining with T-brain1 (Tbr1) antibody on similar sections reveals colocalization of GFP and Tbr1, illustrating that GFP fluorescence is extended into regions of postmitotic neurons. (q–t) Pax6 antibody staining reveals minimal overlap between Pax6 expression in ventricular zone (vz) and GFP expression in subventricular zone (svz) to marginal zone (mz). (u–x) Staining with the phosphohistone-H3 shows colocalization with some GFP expressing cells (white arrowheads). Scale bars = 100 μm. vz, ventricular zone; svz, subventricular zone; iz, intermediate zone; sp, subplate; cp, cortical plate; mz, marginal zone; fl, forelimb; hl, hindlimb.