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Figures
- Fig. 1.
DLAV plexus morphogenesis in zebrafish (A–P) and mouse (Q–U) embryos. (A–D) Left lateral views (anterior, left; dorsal, up) of primary Se and DLAV development at 24 (A), 28 (B) and 32 hpf (C–D; D is a magnified detail of C). (E–M′″,O–P) Lateral (E,K,L–M; L is a 3D rendering), dorsal (G–I, O–P) and cross-sectional (F, from E; M′–M′″ correspond to levels m′–m′″ in M) views of DLAV plexus morphogenesis in the mid (E–M′″) and anterior (O–P) trunk regions at 38 (E–G), 46 (H), 48 (I,O), 56 (J–L), 60 (M–M′″) and 72 hpf (P). Anterior, left (lateral and dorsal views); dorsal, up (lateral and cross-sectional views). White asterisks (F–L,Q), examples of contra-lateral DLAV interconnections. White arrowhead (J–L), midline dorsal vessel). (N) Development of contra-lateral DLAV interconnections at 36–46 hpf). The x-axis shows their anteroposterior location with regards to consecutive Se vessels pairs (anterior-most Se vessel usually found at the third somite boundary level); n = 9–11 animals per stage. (A–P) Data from Tg(fli:EGFP)y1 embryos [M–M′″ includes Tg(flk1:ras-mCherry)s896]. Endothelium, green (A–K, M–M′″,O–P) and red (M–M′″) or gray (L). Somite boundaries, red (A–E,G–I,O–P). Scale bars: (A–M′″,O–P): 30 µm (white); 20 µm (red). (Q–U) 3D-rendered dorsal views of DLAV plexus morphogenesis in mouse embryos at the 22, 26, 27, 32, and 34-somite stage, respectively. Endothelium, gold. Anterior, left. Scale bars: 250 µm. See the Materials and Methods for details.
- Fig. 2.
Endothelial cell abundance and behaviors during DLAV plexus morphogenesis. (A) Quantification of endothelial cell abundance within the developing DLAVs and DLAV plexus at 34, 48 and 72 hpf in the anterior and mid trunk (regions spanning somites 3–5 and 8–11, respectively); n = 10–11 animals per stage. Error bars show the s.e.m. ***P<0.001. (B,C) Dorsal views of the mid trunk (region spanning somites 8–12) DLAVs and DLAV plexus at 48 (B) and 72 (C) hpf; anterior, left. (A–C) Data from Tg(flk1:EGFP-NLS); Tg(flk1:ras-mCherry)s896 embryos. (D–F′) Trunk vasculature (details) of a chimeric embryo from a Tg(flk1:EGFP-NLS) donor and a Tg(flk1:ras-mCherry)s896 host at 32 (D,D′), 56 (E,E′), and 72 (F,F′) hpf. Anterior, left. White asterisks, endothelial cell nuclei of donor origin found within DLAV plexus. (B–F′) Endothelium, red; endothelial cell nuclei, green. Scale bars: 30 µm.
- Fig. 4.
Circulatory flow and VEGF signaling make additive contributions to DLAV plexus morphogenesis. (A–D′) Lateral views of the mid trunk vasculature of 56 hpf Tg(flk1:EGFP-NLS); Tg(flk1:ras-mCherry)s896 embryos under the indicated four conditions. Anterior, left; dorsal, up. Endothelium, red; endothelial cell nuclei, green; somite boundaries, blue (A,B,C,D). Scale bars: 30 µm. (E–G) Quantifications of the effect of treatments (see Materials and Methods) on the vascular; n = 6–10 animals per treatment and stage. Error bars, s.e.m. ***P<0.001. The normalized endothelial cell abundance within the whole trunk vasculature (E) or specific subsets of it (F–G) was determined. (H) Diagram showing a cross-sectional view of the trunk blood vasculature (∼56 hpf). Endothelium color-coded according to its arterial (red) or venous (blue) lineage; vessel lumens, white. Dorsal, up; left side, left. (I) Favored model of how DLAV plexus morphogenesis and maintenance are positively regulated by circulatory flow and VEGF signaling. It is likely that additional flow-dependent signals (‘?’) are involved in this process.
- Fig. 3.
Maturation of DLAV and DLAV plexus circulatory function during development. (A–J′) Mid trunk DLAV and DLAV plexus vasculature (details). Endothelium is shown in green [Tg(fli:EGFP)y1]. Microangiograms of circulatory plasma flow is shown in red (Qdot-605). Anterior, left; dorsal, up (lateral views). Scale bars: 30 µm.
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