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Characterization of DLV formation defects in <italic>vegfab</italic> and <italic>vegfc</italic> mutants at distinct larval stages and visualization of perfused cranial vasculature in WT and <italic>vegfab</italic> mutant larvae.(A–D) Dorsal views of 102 hpf WT (A), vegfab-/- (B), vegfc-/- (C), and vegfd-/- (D) cranial vasculature visualized by Tg(kdrl:EGFP) expression. A significant fraction of vegfab-/- and vegfc-/- larvae lacked the DLV at 102 hpf. (E and F) Percentage of the fish of indicated genotype with and without the DLV at 102 (E) and 154 (F) hpf. A significantly higher fraction of vegfab-/- and vegfc-/- fish lacked the DLV at 102 hpf (47% and 40%, respectively) than WT (0%) (n = 13 for WT, n = 19 for vegfab-/-, n = 25 for vegfc-/-, and n = 19 for vegfd-/- fish examined at 102 hpf). At 154 hpf, while 63% of vegfab-/- fish exhibited the ‘No DLV’ phenotype, only 13% of vegfc-/- fish showed this phenotype (n = 16 for WT, n = 27 for vegfab-/-, n = 23 for vegfc-/-, and n = 26 for vegfd-/- fish examined at 154 hpf). Only a small fraction of vegfd-/- fish examined lacked the DLV at 102 and 154 hpf (5% and 0%, respectively). (G–H") Dorsal views of 96 hpf vegfab+/+ (G–G") and vegfab-/- (H–H") heads of larvae that carry the Tg(kdrl:EGFP) transgene (G and H) and that were injected with streptavidin-conjugated Qdot 655 nanocrystals (G’ and H’) (merged images shown in G" and H"). In vegfab+/+ fish (n = 10), the DLV is visualized by both Tg(kdrl:EGFP) expression (G) and blood-circulating Qdot 655 nanocrystals (G’). In contrast, in the vegfab-/- larvae that lack the Tg(kdrl:EGFP)-positive DLV (H, n = 8), blood-circulating Qdot 655 nanocrystals did not label the DLV (H’), showing the lack of a perfused, functional DLV. Yellow arrows point to the presence and absence of the DLV in vegfab+/+ and vegfab-/- larvae, respectively. Scale bars: 50 µm.
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