Fig. 1

Fig. 1

a TUNEL assay (A, B) and HuC/D (C, D) immunostaining of PTU-treated normal Sb and pnt^tq209 mutant zebrafish at 72 hpf to detect apoptotic cells and differentiated neurons, respectively. The merge between the two fluorescent signals is shown in E and F. b Quantification of apoptotic bodies, detected with the TUNEL assay, in Sb and pnt^tq209 zebrafish, showing a significant increase in the total number of apoptotic bodies in the brain and retina of mutant larvae (number of apoptotic bodies (TUNEL-positive puncta) in the brain, Sb: 31.10 ± 2.35, pnt^tq209: 57.53 ± 3.12; in the retina, Sb: 5.33 ± 0.85, pnt^tq209: 21.80 ± 1.52; results are reported as mean ± S.E.M. (n = 15–20)). c apoE in situ hybridization (A, B) and NR staining (C–F) were carried out in PTU-treated, 72 hpf zebrafish to visualise microglia. d, e Quantification of apoE-positive (d) and NR-positive (e) cells in the whole CNS. A significant increase in the number of microglial cells characterizes pnt^tq209 zebrafish ((number of apoE-positive puncta, Sb: 13.10 ± 1.28, pnt^tq209: 25.20 ± 1.38; number of NR-positive puncta, Sb: 17.50 ± 1.18, pnt^tq209: 23.13 ± 1.35; results are reported as mean ± S.E.M. (apoE: n = 10; NR: n = 24)). f Analysis of the size of NR puncta. pnt^tq209 zebrafish showed a significant increase in the size of microglia when compared to Sb (microglial area (NR-positive pixels), Sb: 210.45 ± 15.65, pnt^tq209: 379.35 ± 26.73; results are reported as mean ± S.E.M. (n = 15)). g, h Evaluation of the number of cells stained with apoE (g) and NR (h) in the brain and retina, separately. The results demonstrate that, in pnt^tq209 larvae, the increase in microglial cell population is also extended to the retina ((number of apoE-positive puncta in the brain, Sb: 4.40 ± 0.56, pnt^tq209: 9.60 ± 1.38; in the retina, Sb: 8.70 ± 1.58, pnt^tq209: 15.60 ± 1.73; number of NR-positive puncta in the brain, Sb: 11.19 ± 1.10, pnt^tq209: 16.50 ± 1.21; in the retina, Sb: 5.44 ± 0.93, pnt^tq209: 7.94 ± 1.23; results are reported as mean ± S.E.M. (apoE: n = 10; NR: n = 15)). FB forebrain, HB hindbrain, MB midbrain, R retina