ZFIN ID: ZDB-FIG-210324-54
Halabi et al., 2021 - Semaphorin 3fa Controls Ocular Vascularization From the Embryo Through to the Adult. Investigative ophthalmology & visual science   62:21 Full text @ Invest. Ophthalmol. Vis. Sci.
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Gene:
Antibodies:
Fish:
Anatomical Terms:
Stage Range: Day 5 to Days 14-20
PHENOTYPE:
Fish:
Observed In:
Stage Range: Day 5 to Days 14-20

Fig. 6 No obvious disruption of the RPE and outer limiting membrane with the loss of Sema3fa. (A, B) Transverse sections of 8 dpf WT and sema3faca304 mutant light-adapted retinas labeled with hematoxylin and eosin. A higher power view of the RPE is shown in A’ and B’. No obvious gaps are seen in the RPE, and apical microvilli (arrows) are evident in both genotypes (N = 3, n = 9 at 72 hpf for both genotypes; N = 2, n = 10 at 8 dpf for both genotypes). (C, D) Immunolabeling of the RPE with the Zpr2 antibody is comparable in WT (N = 2, n = 9) and mutant (N = 2, n = 9) retinas. (E–H) No obvious disruption of the outer limiting membrane (OLM; arrows), as stained with an antibody for the adhesive junction protein ZO-1 (N = 2, n = 10/10 WT eyes; n = 7/7 sema3faca304 eyes) and phalloidin that labels F-actin (N = 3, n = 15/15 WT eyes; n = 16/16 sema3faca304 eyes). (I, J) Glutamine synthetase immunolabeling of the Müller glia in WT (I; N = 2, n = 11/11) and mutant (J: N = 2, n = 10/10) 7 dpf retinas, revealing labeling of the apical Müller glia endfeet at the OLM (arrows) in both genotypes. (K, L) Labeling at the OLM (arrows) of F-actin (phalloidin) and Müller glia endfeet in a Tg(gfap:egfp) background in WT (n = 7/7) and sema3faca304 (n = 8/8) 15 dpf retinas. (M, N) Working model of roles for Sema3fa in regulating vascularization of the neural retina. Choroid vasculature (M): In WT, the choroid plexus (CP) remains outside of the RPE and Bruch's membrane (bm) through the release of Sema3fa (purple hexagon) by cells of the inner nuclear layer and the RPE. If Sema3fa signaling is perturbed, vessels pass through the RPE and leak into the nuclear layers containing the cone and rod (C/R) photoreceptors, horizontal cells (HC), and bipolar cells (BC). Hyaloid/retinal vasculature (N): Sema3fa (purple hexagon) maintains spacing and reduces the density of the blood vessel networks in WT retinas. In the absence of Sema3fa blood vessels overgrow. We propose that in both models Sema3fa acts via Nrp2b (yellow rectangle) and blood vessel growth is supported but not driven by the pro-angiogenic Vegf.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
EGFP nt11Tg standard conditions Days 14-20 Muller cell IFL
sema3faca304/ca304; nt11Tg standard conditions Days 14-20 Muller cell IFL
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Anatomy
Ab1-glul IHC WT standard conditions Days 7-13 Muller cell
IHC sema3faca304/ca304 standard conditions Days 7-13 Muller cell
Ab1-tjp1 IHC WT standard conditions Day 5 outer limiting membrane cell-cell junction
IHC sema3faca304/ca304 standard conditions Day 5 outer limiting membrane cell-cell junction
zpr-2 IHC WT standard conditions Days 7-13 retinal pigmented epithelium
IHC sema3faca304/ca304 standard conditions Days 7-13 retinal pigmented epithelium
Phenotype Details
Fish Conditions Stage Phenotype
sema3faca304/ca304 standard conditions Day 5 outer limiting membrane morphology, normal
Days 7-13 retinal pigmented epithelium morphology, normal
sema3faca304/ca304; nt11Tg standard conditions Days 14-20 outer limiting membrane morphology, normal
Acknowledgments:
ZFIN wishes to thank the journal Investigative ophthalmology & visual science for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Invest. Ophthalmol. Vis. Sci.