Fig. 4

Caudal fin bony ray development and innervation progress in the absence of endothelium. (A) Diagram illustrating experimental scheme of VEGFR inhibition in developing zebrafish. (B-G) Confocal projections of fins expressing reporters for osteoblasts [Tg(sp7:mCherry-NTR)] and endothelium [Tg(fli1a:EGFP)] and immunostained for axons (acetylated tubulin antibody). Treatments and stages as indicated. Yellow arrowhead denotes the future position of the hypural diastema (HD). Note that despite the severe reduction in endothelial growth into the fin following PTK787 treatment, early osteoblast and axon patterning appear unperturbed. (H-K) Quantification of developmental metrics and ray innervation between treatment groups. Lines represent linear regression (H,I,K) or local polynomial smoothing (J). SL/flexion angle slope between the two independent trials of DMSO control fish and PTK-treated fish was not significantly different (P=0.158; ANCOVA). Number of bony rays/flexion angle slope between the two independent trials of DMSO control fish and PTK treated fish was not significantly different (P=0.913; ANCOVA). Number of innervated rays/flexion angle slope between the two independent trials of DMSO control fish and PTK treated fish was significantly different above a flexion angle of 13.06 degrees (P<0.05; Johnson–Neyman technique). For H,I,K, a total of n=24 PTK787-treated (mean SL=5.14±0.74 s.d.) and n=18 DMSO-treated fish (mean SL=5.28±0.73 s.d.) are plotted. For J, a total of n=18 PTK787-treated (mean SL=5.49±0.58 s.d.) and n=17 DMSO-treated fish (mean SL=5.72±0.61 s.d.) are plotted. Data are representative of at least three biological replicates. Scale bars: 100 μm (B-G).