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Fig. 3. Exogenous Axin1 promotes hypothalamic and suppresses floorplate marker gene expression. Lateral (A,C,J) and dorsal (B,E,F,K,L,M) views and 14 µm transverse sections (D,G-I) of brains of embryos between 22 and 28 somites in which cells (brown) expressing various constructs (top right) were transplanted into the prospective hypothalamus/anterior floorplate of 60-65% epiboly stage hosts. Sections are at the level of the ventral midbrain (D,G,I) and the hindbrain (H). (A-B) gfp-expressing cells (brown) integrate in the ventral midbrain and hindbrain but do not express nk2.1a. (C-D) Cells overexpressing axin1 (brown, asterisks in the insets) in the midbrain and hindbrain of two different embryos ectopically express nk2.1a (blue, asterisks). The arrow points to the absence of ectopic nk2.1a expression in more dorsal cells overexpressing axin1 (brown) in the midbrain. (E-I) Floorplate marker expression (blue) is downregulated in axin1 overexpressing cells (brown, arrowheads). The expression of foxa2 is shown in the same embryo after in situ hybridization (E) and after immunohistochemistry to reveal the axin1 overexpressing cells (brown, F) and in transverse sections (G,H). The arrowheads point to sites of downregulation of foxa2 or foxa1 in the transplanted cells in the midline of the midbrain (E-G,I) and hindbrain (H). (J-K) axin1 overexpressing cells do not ectopically express foxg1 in the posterior ventral neural tube of the embryo (J) and in higher magnification (K). (L-M) Exogenous axin1 expands shh expression (arrowheads) lateral to the CNS midline shown after in situ hybridization (L) and after immunohistochemistry to reveal the transplanted cells (M). Abbreviations: mb, midbrain; hb, hindbrain; fp, floorplate; pd, posterior diencephalon.

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