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Figure 7


Fig. 7. Summary of HH function in the ventral midbrain. (A-D) Whole mounts; (E) cross-section. (A-C) A three-step temporal patterning of the ventral midbrain by HH blockade. (A) Electroporation at or before H&H stage 11: anterior-medial rFP patterning is complete or HH-independent. The caudo-medial rFP, lateral rFP and all other midbrain cell fates still require HH signaling for their specification. Increased cell spread is noted. (B) Electroporation at H&H stages 11-13: medial rFP specification is complete. Lateral rFP and arcuate cell fates, represented here by PHOX2A (blue), PAX6 (purple) and EVX1 (yellow), continue to be specified. The caudo-medial midbrain requires HH signaling for a longer time than does the rostral midbrain. HH is also required for forming a coherent arc pattern. (C) Electroporation after H&H stage 13: midbrain patterning is complete, with the exception of the lateral rFP and cell fates associated with this region (e.g. arc 2). Ectopic cell spread is noted only in this region. (D) HH signaling regulates midbrain boundaries. Disruption of the boundaries of the midbrain results in the non-autonomous spread of PAX7+ (green) cells at the DV boundary and of WNT1+ (blue) cells and FGF8+ (brown) cells at the MHB. Ectopic PAX7 expression (arrow) is also seen in ventral midbrain progenitors as a result of re-specification into dorsal phenotypes. (E) PHOX2A expression, demonstrating that Ptc1{Delta}loop2 electroporations (right of the vertical line) result in increased cell proliferation (expanded ventricular layer) and in reduced differentiation compared with controls (left of the vertical line). The effects are seen within columns of cytoplasmically connected midbrain cells (white), which line up ventricular to the Ptc1{Delta}loop2+ cells (blue circles). Non-autonomous cell spread (*) is also seen. 1, first arc; 2, arc 2; III, third ventricle; E1, EVX1; EP, electroporated; P6, PAX6; H&H, embryonic stages according to Hamburger and Hamilton (Hamburger and Hamilton, 1951); MHB, midbrain-hindbrain boundary; rFP: rostral floor plate; TEC, tectum.