Expression of tcf and top:dgfp mRNA in the 50 hpf embryonic zebrafish hypothalamus. (A–F) Lateral views of whole-mount brains at 50 hpf. (G–L) Ten-micron plastic cross sections of the posterior hypothalamus. Sectioned regions are indicated in panels (A–F). Circled regions in (G–L) indicate domains of specific gene expression. top:dgfp and all tcf genes except tcf7l2 are expressed in the posterior hypothalamus. T, telencephalon; Ha, habenula; TeO, tectum opticum; DT, dorsal thalamus (thalamus); Hy, hypothalamus. Scale bars: 100 μm.

Expression of tcf genes and top:dgfp in the larval and adult zebrafish hypothalamus. (A–C) Ventral view of 10 dpf larval brains; orientation is indicated in panel (A). (D–F) Higher power images of the fields indicated in (A–C). top:dgfp Expression encompasses the combined domains of lef1 and tcf7. (G–I) Ventral views of 6mpf adult brains. (J) Cross section through adult hypothalamus at the level indicated in panel (G). (K) GFP antibody staining on an adult TOP:dGFP hypothalamus cross section at the same level as panel (J). (L) axin2 expression on an adult hypothalamus cross section at the same level as panel (J). lef1 and GFP/axin2 occupy different regions of the periventricular zone. pHy, periventricular hypothalamus. Scale bars: (A, D) 100 μm; (G) 500 μm; (J) 200 μm.

Immunohistochemical identification of Wnt-responsive cells in the 50 hpf embryonic hypothalamus. (A) Lateral view of TOP:dGFP whole-mount brain stained with GFP antibody, observed in a 100-μm confocal projection. (B, C) Triple labeling for TOP:dGFP, HuC/D or PCNA, and Topro3, observed in a single confocal slice. (D–I) Twelve-micron cryosections immunostained for the markers listed in each panel. Positions of cross sections are indicated in panel (A). Boxed region is shown at higher magnification in lower right corner, and circled cells are double labeled. GFP staining partially overlaps with HuC/D, PCNA, and 5-HT, but not with TH, GABA, or GFAP. T, telencephalon; TeO, tectum opticum; DT, dorsal thalamus (thalamus); Hy, hypothalamus. Scale bars: (A, B) 100 μm; (D) 50μm.

Immunohistochemical identification of Lef1- expressing cells in the 50 hpf embryonic hypothalamus. (A–E) Twelve-micron cryosections immunostained for the markers listed in each panel. Positions of cross sections for panels (A) and (C–E) are at the same level as Figure 3E, and panel (B) is at the same level as Figure 3G. Boxed region is shown at higher magnification in lower right corner, and circled cells are double labeled. Lef1 staining partially overlaps with TOP:dGFP, HuC/D, PCNA, and dlx5/6:gfp, but not with GFAP. (F) Lateral view of dlx5/6:gfp whole-mount brain, observed with a 100-μm confocal projection. (G–L) Twelve-micron cryosections immunostained for the markers listed in each panel. Positions of cross sections are indicated in panel (F). Boxed region is shown at higher magnification in lower right corner, and circled cells are double labeled. GFP staining partially overlaps with HuC/D, PCNA, and GABA, but not with TH, 5-HT, or GFAP. T, telencephalon; TeO, tectum opticum; DT, dorsal thalamus (thalamus); Hy, hypothalamus. Scale bars: (A, G) 50 μm; (F) 100 μm.

Immunohistochemical identification of Wnt-responsive and Lef1-expressing cells in the adult hypothalamus. (A) GFP antibody staining on a sagittal section through the midline of a TOP:dGFP adult brain. Specific expression of GFP is observed in the optic tectum (white box) and the periventricular hypothalamus (red box). (B–D) Twenty-five-micron hypothalamic cross sections immunostained for the markers listed in each panel. GFP-positive cells do not express HuC/D, PCNA, or GFAP. Scale bars: (A) 300 μm; (B) 100 μm.