RGC-specific Gal4 expression in the adult zebrafish brain. a–d: Expression of GFP and GFAP in the adult Tg(gfap:Gal4FF;UAS:GFP) telencephalon. Sections of the telencephalon of Tg fish were stained with anti-GFAP (red) antibodies. Nuclei were stained with (Hoechst blue). a′–d′: Magnified images of the white-boxed areas above. e-h: Expression of GFP and BLBP in the adult Tg(gfap:Gal4FF;UAS:GFP) telencephalon. Sections of the telencephalon of Tg fish were stained with anti-BLBP (red) antibody. Nuclei were stained with Hoechst (blue). e′–h′: Magnified images of the white-boxed areas above. i: Colocalization of GFP with BLBP in the adult telencephalon. The percentage of GFP(+)/BLBP(+) cells is 97.51 ± 2.49% in the Dm domain, 98.11 ± 1.89% in the Dl domain, 98.85 ± 1.15% in the Vd domain, and 94.31 ± 5.69% in the Vv domain. Data are expressed as mean ± SEM; n = 3. Scale bars, 200 µm in a and e, 20 µm in a′ and e′.

RGC-specific NTR expression in the adult zebrafish brain. a–d: Expression of mcherry and GFAP in the adult Tg(gfap:Gal4FF;UAS:nfsB-mcherry) telencephalon. Sections of the telencephalon of Tg fish were stained with anti-GFAP (green) antibody. Nuclei were stained with Hoechst (blue). a′–d′: Magnified images of the white-boxed areas above. e–h: Expression of mcherry and BLBP in the adult Tg(gfap:Gal4FF;UAS:nfsB-mcherry) telencephalon. Sections of the telencephalon of Tg fish were stained with anti-BLBP (green) antibody. Nuclei were stained with Hoechst (blue). e′–h′: Magnified images of the white-boxed areas above. i: Colocalization of mcherry with BLBP in the telencephalon. The percentage of mcherry(+)/BLBP(+) is 91.27 ± 2.02% in the Dm domain, 9.61 ± 1.47% in the Dl domain, 43.61 ± 4.51% in the Vd domain, and 8.71 ± 5.98% in the Vv domain. Data are expressed as mean ± SEM; n = 3. Scale bars, 200 µm in a and e, 20 µm in a′ and e′.

NTR-mediated RGC ablation in Tg(gfap:Gal4FF;UAS:nfsB-mcherry) adult zebrafish. a: Mtz treatment schedule for RGC-specific ablation. b–d: Expression of mcherry and GFAP in the telencephalon of adult Tg zebrafish after Mtz treatment. Sections of the telencephalon of Tg fish were stained with anti-GFAP (green) antibody. Nuclei were stained with Hoechst (blue). e–g: Expression of mcherry and GFAP in the telencephalon of adult Tg(gfap:Gal4FF;UAS:nfsB-mcherry) zebrafish. Sections of the telencephalon of Tg fish were stained with anti-GFAP (green) antibody. Nuclei were stained with Hoechst (blue). i–l: The number of mcherry-positive cells in the adult telencephalon with or without the Mtz treatment. Dm, Dl, and Vd. *P < 0.05. Scale bars, 200 µm in b and e, 100 µm in b′ and e′.

RGC-ablation reduces proliferative RGCs in the adult zebrafish brain. a: Mtz treatment schedule for RGC-specific ablation. b–g: Expression of PCNA and GFAP in WT without (b–d) or with Mtz treatment (e–g). Sections of the telencephalon of WT fish brains were stained with anti-PCNA (green) and anti-GFAP (red) antibodies. Nuclei were stained with Hoechst (blue). h–m: Expression of PCNA and GFAP in Tg(gfap:Gal4FF;UAS:nfsB-mcherry) zebrafish without (h–j) or with Mtz treatment (k–m). Sections of the telencephalon of Tg fish were stained with anti-PCNA (green) and anti-GFAP (red) antibodies. Nuclei were stained with Hoechst (blue). n–q: Quantification of PCNA(+)/GFAP(+) cells in WT and Tg zebrafish with or without Mtz treatment. *P < 0.05, **P < 0.01, and ***P < 0.001. Data are expressed as mean ± SEM; n = 3. Scale bars, 100 µm in b, e, h, and k.