Journal: Frontiers in Cellular Neuroscience

Article Title: Angiopoietin 1 and integrin beta 1b are vital for zebrafish brain development

doi: 10.3389/fncel.2023.1289794

Figure Lengend Snippet: Transgenic expression of zebrafish angpt1 increases proliferation and HuC-positive cells but reduces GABAergic neurons. 5-dpf dissected brains were collected from Turku wild-type embryos injected with Tol2 constructs, including control or angpt1 transgene driven by h2afx , elavl3 , and gfap promoter at the one-cell stage. Brain samples were stained with anti-HuC and anti-GABA antibodies following the EdU assay. EdU-proliferation images shown in (A) control (B) h2afx : angpt1 (C) elavl3 : angpt1 (D) gfap : angpt1 (E) Quantification of EdU-positive cell numbers in the Hc region [ N = 7–11 per injected group, F (3, 31) = 10.75 P < 0.0001]. HuC-immunostaining images shown in (F) control (G) h2afx : angpt1 (H) elavl3 : angpt1 (I) gfap : angpt1 (J) Quantification of HuC-positive cell numbers in the Hc region [ N = 7–11 per injected group, F (3, 31) = 11.58, P < 0.0001]. GABA-immunostaining images show (K) control (L) h2afx : angpt1 (M) elavl3 : angpt1 (N) gfap : angpt1 (O) Quantification of GABA-positive cell numbers in the Hc region [ N = 7–11 per injected group, F (3, 31) = 33.92, P < 0.0001]. (A’–N’) Show the high magnification images corresponding to the white rectangular area in the Hc region shown in (A) and equivalent in (B–N) . EdU-positive cells are displayed in magenta, HuC-positive cells in cyan, and GABA-positive cells in green. Hc, caudal hypothalamus. Statistical analysis is shown in mean ± SD by an ordinary one-way ANOVA with the Tukey multiple comparisons test. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001. Scale bar is 200 μm.

Article Snippet: 5′-entry vectors, including pENTR5′- ubi (ubiquitin promoter) (plasmid #27230), p5E- elavl3 (plasmid #75025), and p5E- gfap (plasmid #75024), were obtained from Addgene ( ; ), and p5E- h2afx was from the Tol2Kit.

Techniques: Transgenic Assay, Expressing, Injection, Construct, Control, Staining, EdU Assay, Immunostaining

Journal: Frontiers in Cellular Neuroscience

Article Title: Angiopoietin 1 and integrin beta 1b are vital for zebrafish brain development

doi: 10.3389/fncel.2023.1289794

Figure Lengend Snippet: Zebrafish angpt1 regulates neurogenesis in a tek-independent manner. 5-dpf dissected brains were collected from tek +/+ and tek –/– embryos injected with Tol2 constructs including control, angpt1 transgene driven by h2afx , elavl3 , or gfap promoter at one-cell stage. The brain samples were stained with anti-HuC and anti-GABA antibodies following the EdU assay. EdU-proliferation images shown in (A) control of tek +/+ (B) control of tek –/– (C) tg ( elavl3 : angpt1 ) in tek +/+ (D) tg ( elavl3 : angpt1 ) in tek –/– (E) Quantification of EdU-positive cell numbers in the Hc region ( N = 7 per transgene-genotype group, genotype factor F (1, 24) = 23.43, P < 0.0001; transgene factor F (1, 24) = 82.89, P < 0.0001). HuC-immunostaining images show (F) control of tek +/+ (G) control of tek –/– (H) tg ( elavl3 : angpt1 ) in tek +/+ (I) tg ( elavl3 : angpt1 ) in tek –/– (J) Quantification of HuC-positive cell numbers in the Hc region [ N = 7 per transgene-genotype group, genotype factor F (1, 23) = 4.501, P = 0.0449; transgene factor F (1, 23) = 92.70, P < 0.0001]. GABA-immunostaining images show (K) control of tek +/+ (L) control of tek –/– (M) tg ( elavl3 : angpt1 ) in tek +/+ (N) tg ( elavl3 : angpt1 ) in tek –/– (O) Quantification of GABA-positive cell numbers in the Hc region [ N = 7–11 per transgene-genotype group, genotype factor F (1, 24) = 0.5009, P = 0.4859; transgene factor F (1, 24) = 98.17, P < 0.0001]. (A’–N’) Show the high magnification images corresponding to the white rectangular area in the Hc region shown in (A) and equivalent in (B–N) . EdU-positive cells in magenta, HuC-positive cells in cyan, and GABA-positive cells in green. Hc, caudal hypothalamus. Statistical analysis is shown in mean ± SD by an ordinary two-way ANOVA with Sidak’s multiple comparisons test, with a single pooled variance. ** p < 0.01 and **** p < 0.0001. Scale bar is 200 μm.

Article Snippet: 5′-entry vectors, including pENTR5′- ubi (ubiquitin promoter) (plasmid #27230), p5E- elavl3 (plasmid #75025), and p5E- gfap (plasmid #75024), were obtained from Addgene ( ; ), and p5E- h2afx was from the Tol2Kit.

Techniques: Injection, Construct, Control, Staining, EdU Assay, Immunostaining