Fig. 3. Cell proliferation and cell death analysis in wild-type and mutant embryos. (A-C) Cell proliferation analysis, using PH3 antibody labeling. (A,B) Fixed and labeled wild-type brain at 17 and 21 hpf. (C) Quantification comparing MHB region and hindbrain, n=8, P=0.627 at 17 hpf; P=0.008 at 21 hpf. Cell proliferation at 21 hpf is significantly higher, showing an almost twofold increase in proliferation in the hindbrain than in the MHB. (D-G) Ventricle formation after inhibition of cell proliferation by aphidicolin treatment. (D,F) Live control and treated embryos after ventricle injection at 24 hpf; (E,G) same embryos as in D and F, fixed and labeled for cell proliferation. Reduced cell proliferation leads to a decrease in ventricle opening. (H-J) Cell death analysis, using TUNEL labeling with ApopTag kit. (H,I) Fixed and labeled wild-type brain at 17 and 21 hpf. (J) Quantification comparing midbrain, MHB and hindbrain regions, n=10, P=0.575 at 17 hpf; P=0.368 at 22 hpf. Error bars denote standard error. H, hindbrain ventricle; M, midbrain ventricle; MHB, midbrain-hindbrain boundary. Scale bar: 50 µm.