Figure 6

Absence of bone after inhibition of NO formation. (A,B). Live staining of 5 dpf zebrafish with DAF-FM DA for 3 h in the dark after 90 h of treatment with the NOS inhibitor TRIM (A) or its solvens (0.1% DMSO) (B). Note the presence of opercular bone (arrow) and cleithrum (arrowhead) in the control, which are clearly absent in the TRIM-treated specimen. (C,D). Alizarin red S whole mount staining of TRIM-treated embryos likewise shows the complete absence of bones (C) compared to their presence in control embryos (D). Arrow: opercular bone; arrowhead: cleithrum. (E–H′). Semithin sections at comparative cross-sectional levels show the absence of opercular bone and cleithrum ((E,F), and higher magnification in (E′,F′)) compared to their distinct presence in control embryos ((G,H), and their higher magnification in (G′,H′)). Opercular bone: arrow; cleithrum: arrowhead; asterisks indicate the location of absent bones. Scale bars in (A,B) = 200 µm, in (E–H) = 100 µm, in (E′–H′) = 50 µm.