Gbx2 plays a cell-nonautonomous role in controlling the
thalamic lineage boundaries. (A-D) In situ hybridization analysis
of Cdh6 and Efna5 expression on the coronal brain sections
of Gbx2CreER/+ and Gbx2CreER/- embryos
at E14.5. The arrowhead indicates the expression in the thalamus; the asterisk
marks the diminished Cdh6 and lost Efna5 expression in the
mutant embryos. (E-H) X-gal and Fast Red staining of sagittal brain
sections of E16.5 chimeric embryos composed of wild-type (blue) and ES-derived
cells (pink) of genotype Gbx2+/- (E) or
Gbx2-/- (F). The arrow indicates aggregates of
Gbx2-/- cells in the cerebellum; the arrowheads indicate
the sharp thalamic borders. (G,H) Magnified view of the thalamus in E and F.
Note that the Gbx2-/- cells, like
Gbx2+/-, extensively intermingle with the host cells in
the thalamus. (I) Schematic diagram illustrating the generation of
genetic mosaics using CreER-mediated deletion of Gbx2 in the thalamus
of Gbx2CreER/F; R26R embryos. The arrows indicate the
primers for PCR analysis to detect the floxed (1.7 kb) and deletion (0.4 kb)
alleles of Gbx2. (J) PCR analysis of microdissected thalamus
of Gbx2CreER/F; R26R embryos. Note that the thalamus
contains both Gbx2CreER/F and
Gbx2CreER/- cells after tamoxifen administration, but only
Gbx2CreER/F cells without tamoxifen. (K,L)
X-gal staining of coronal brain sections of two
Gbx2CreER/F; R26R embryos with different levels
of β-gal activity at E18.5 after tamoxifen administration at E10.5. The
arrowheads mark the sharp borders of the fate-mapped Gbx2 lineage.
(M-P) In situ hybridization of Efna5 on sagittal brain
sections of Gbx2CreER/+ (M),
Gbx2-/-wild-type chimera (N),
Gbx2CreER/FGbx2CreER/- mosaic (O), and
Gbx2CreER/- (P) embryos at E16.5. Inset in O shows X-gal
staining of a sagittal section of the mosaic embryo. The arrows indicate the
restored expression of Efna5 in the chimeric and mosaic embryos; the
asterisk indicates the absence of Efna1 expression in the thalamus of
Gbx2-/- embryo. ET, epithalamus; Ncx, neocortex; PT,
pretectum; PTh, prethalamus; TH, thalamus; WT, wild type. Scale bars: 400
μm in A-D; 450 μm in E,F; 29 μm in G,H; 380 μm in K; 200 μm in
L-O.