Article Figures & Data
Figures
- Fig. 1.
Characterization of the cells that express the H-2Z1 transgene in the cerebral cortex: A–G, β-galactosidase activity is visualized as a blue(A, C, F–K) or a fluorescence (white in D,E) reaction product. A, Whole-mount staining of the right telencephalic hemisphere of a P8 H-2Z1 mouse oriented to provide an upright position to the somatotopic body representation disclosed by transgene expression, which is schematized in B [modified from Dawson and Killakey (1987)]. Thein situ orientation is given by thearrows. d, Dorsal; a, anterior. The asterisks mark additional staining, probably located in the secondary somatosensory area. C, Sagittal section of a P8 brain illustrating the layer-specific expression of the H-2Z1 transgene; the arrowhead marks a deep neuron that expresses β-galactosidase. D,E, The diffusible substrate FDG fills the cell bodies of H-2Z1-positive cells; the arrowheadsdelineate a single barrel in D and labeled neurites inE. F, G, β-galactosidase activity is localized to small vesicles in the cell body (arrow in F) that accumulate in one to three locations in the cytoplasm (arrowhead inF) and seem to mark some axons (arrowheads in G). H–J, Transverse sections through the somatosensory area of young adult mice treated by double-labeling for the detection of H-2Z1 expression (inblue, arrowheads inH–K) and GAD 67 (purple,arrow in H), calretinin (brown, arrow inI), parvalbumin (brown,arrow in J), or calbindin (brown, arrow inK). H-2Z1 was never colocalized with GAD 67, calretinin, or parvalbumin. In contrast, some H-2Z1-positive cells coexpressed calbindin. Scale bar (shown in F):A, 400 μm; C, 1 mm; D, 100 μm; E, H, 35 μm; G,K, 20 μm; I, J, 50 μm.
- Fig. 2.
Development of H-2Z1 expression in vitro visualized by the blue detection of β-galactosidase activity. A, Slices and whole cortical explant dissections are schematized on the left of the corresponding figures. The left column illustrates the dissection of a left telencephalic vesicle. Both the hippocampus and ganglionic eminence are removed from whole cortical explants. B, C, Vibratome slices (400 μm thick) of E15.5 H-2Z1 cortex cultivated for 11 d. The agarose embedding was removed inB but left in place in C. The flattening of the slices on the Biopore membrane substrate results in the radial spreading of the H-2Z1 neurons during culture time. The mediolateral extent of the expression domain (between arrowheads) is not markedly modified. d, Dorsal; v, ventral. D, E, Regionalized expression of the H-2Z1 transgene (arrowheads) developed after 12 d in vitro in E13.5 whole cortical explants. E, Transverse vibratome section through one of the explants showing the laminar organization of transgene-expressing cells (arrowhead). Tangential cell mixing is minimal in whole cortical explants. p, Parietal.F, Half cortical explants taken from E13.5 rosa-26 (which express ubiquitously lacZ) and OF-1 wild-type mice were juxtaposed. Cell mixing between the two genotypes is minimal. Scale bar (shown in B): B, 400 μm;C, 250 μm; D, 1.2 mm; E, 200 μm; F, 1 mm.
- Fig. 3.
H-2Z1 expression develops in grafts transplanted heterotopically into an embryonic cortex. A, Schematic representation of the grafting procedure. Parietal (solid circle) or occipital (empty circle) grafts are transplanted in the occipital or parietal regions, respectively, of whole cortical explants. B, C, Heterotopic and isochronic transplantation of parietal (arrow in B) or occipital (arrowhead in C) E13.5 H-2Z1 grafts in nontransgenic whole cortical explants. β-galactosidase is expressed in parietal grafts developing in either location, whereas occipital grafts remain unstained. D, E11.5 H-2Z1 parietal cortex grafts transplanted into the occipital (top explant,arrow) or parietal (bottom explant,arrow) region of E14.5 nontransgenic whole cortical explants develop a robust expression of β-galactosidase. Theasterisks in B–D label the olfactory bulb. E–G, In contrast to what occurs when the explants receive a sharp cut, the exchange of grafts between rosa-26 and OF-1 E15.5 embryos results in widespread cell mixing (arrowsin F, G) between graft and host. Scale bar (shown in B): B, D, 500 μm; C, 250 μm; F,G, 400 μm.
- Fig. 4.
Pioneer neurons develop between E10.5 and E11.5in vitro. A, Calretinin-positive neurons appear in the parietal cortex at E11.5. B, Calretinin neurons develop similarly in E10.5 telencephalic explants maintained in culture for 1 d. C, Schematic representation of the dissection, dissociation, and reimplantation of aggregates.D, E14.5 parietal H-2Z1 reaggregate reimplanted in the anterior cortex of a E15.5 whole cortical explant, cultivated for 11 d, and treated with X-gal. Dissociation of parietal grafts results in a sharp decrease in the number of cells and the level of β-galactosidase expression, which is somewhat over-revealed in this picture. Scale bar (shown in A): A,B, 550 μm; D, 200 μm.
Tables
Donor grafts Host telencephalic vesicles β-gal-positive grafts β-gal-negative grafts % of grafts maintaining presumptive expression E13.5 parietal Parietal E15.5 2 0 100 Occipital E15.5 5 1 83 Occipital E13.5 7 1 88 E13.5 occipital Parietal E13.5 1 7 88 E12.5 parietal Occipital E16.5 7 2 78 E12.5 occipital Parietal E13.5 0 9 100 E11.5 parietal Parietal E14.5 14 1 93 Occipital E14.5 10 6 63 E11.5 occipital Parietal E14.5 1 8 89 Occipital E14.5 1 9 90
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