Fig. 9. Whole-mount in situ hybridization of ligated embryos with arterial markers. Ligations were performed for the indicated time periods, and embryos were hybridized with the indicated antisense riboprobes. Arrows (A-H) show ligation site. (A-F) Arterial markers. Note arterial expression on the left side of all embryos and progressive downregulation of NRP1 (A-D) and ephrinB2 (E), as well as the absence of binding of EphB4-Fc (F) on the ligated side. (G-J) Venous markers. (G,H) No upregulation of venous marker expression is detectable after 1 hour of ligation. (I,J) 12 hours after ligation. Tie2 and NRP2 mRNA are not expressed in the anterior (I) or posterior (J) left vitelline arteries (A) that cross over the midline, although they are both expressed in venularized arteries (arrowheads) on the ligated side. Scale bars: 530 µm in A,C,D; 510 µm in B; 725 µm in E; 230 µm in F; 700 µm in G,J; 880 µm in H; 435 µm in I.