Fig. 4. Retinoid signaling is required for maintenance of FGF8 and SHH in the forebrain and FNP ectoderm. In situ hybridization of midline sagittal sections 4 hours after bead implantation at stage 10 show that control embryos (A) express FGF8 (red) in the rostrodorsal forebrain (f) and ectoderm of the presumptive FNP (arrows). (E) SHH (yellow) is strongly expressed in the ventral forebrain (f) and notochord (n). (B) Exposure to RAR/RXR antagonists results in a downregulation of FGF8 in the forebrain (asterisk), but not in the isthmus (i), where expression is normal (n=15). (F) At this early stage, SHH expression does not appear to be downregulated (n=15). The change in FGF8 expression precedes any evidence of an increase in programmed cell death. Four hours after bead implantation at stage 10, the amount of programmed cell death (bright green) is equivalent in control (I) and RAR/RXR antagonist-treated (J) embryos (arrows), as determined with a TUNEL assay (n=10). Sections 6 hours after bead implantation at stage 10 show that control embryos continue to express FGF8 (C) and SHH (G) in the ventral forebrain (arrow). (D) RAR/RXR antagonist-treated embryos lack FGF8 in the forebrain and FNP ectoderm (asterisk; n=15). (H) SHH is downregulated along the ventral forebrain (asterisk; n=15). Six hours after bead implantation at stage 10, the amount of programmed cell death is nominally increased in the mesenchyme (asterisk) of RAR/RXR antagonist-treated embryos (L, n=7), compared with that found in controls (K, n=6). Other labeled structures are the hindbrain (h) and midbrain (m). Scale bars: 200 µm.
