June 2015
Volume 56, Issue 7
ARVO Annual Meeting Abstract  |   June 2015
Endothelial cells provide paracrine signals to regulate retinal neurogenesis in zebrafish embryos.
Author Affiliations & Notes
  • Susov Dhakal
    University of Idaho, Moscow, ID
  • Deborah L Stenkamp
    University of Idaho, Moscow, ID
  • Footnotes
    Commercial Relationships Susov Dhakal, None; Deborah Stenkamp, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2015, Vol.56, 1482. doi:
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      Susov Dhakal, Deborah L Stenkamp; Endothelial cells provide paracrine signals to regulate retinal neurogenesis in zebrafish embryos.. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):1482.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: The role of vascular endothelial cells during retinal neurogenesis and development of the retina remains poorly understood. We tested the role of the vascular endothelial cells in the development of the retina in zebrafish embryos.

Methods: We utilized Gal4-UAS system of gene expression by crossing two transgenic lines of zebrafish, Tg(cdh5:gal4) and Tg(UAS-E1b:Eco.NfsB-mCherry) to create doubly transgenic embryos such that the Nfsb-mCherry reporter was expressed specifically in cells expressing cadherin5 (endothelial cells). When the doubly transgenic embryos were treated with a pro-drug, Metronidazole (Mtz), Mtz reacted with E. coli nitroreductase-B enzyme to produce a toxic byproduct, which killed the cells expressing the transgene so that selective ablation of endothelial cells was achieved. The embryos were then evaluated for eye and lens size, retinal lamination, and neuronal and non-neuronal cell differentiation using specific microscopic and histological techniques. We also analyzed the retinal phenotype of silent heart mutant zebrafish, which have a mutation in troponin2a gene- resulting in a noncontractile heart but intact vasculature- in order to test whether circulating factors have any role in regulating retinal neurogenesis.

Results: Experimental destruction of endothelial cells during retinal neurogenesis caused microphthalmia and disrupted retinal lamination. Retinal progenitor proliferation, as well as differentiation of specific retinal cell types was also reduced by the experimental destruction of endothelial cells. Our results also indicate that the retinal phenotype observed in the experimentally avascular embryos was not due to increased cell death. The retinal phenotype was normal in silent heart mutants.

Conclusions: Our results suggest that a regulatory relationship exists between the ocular vasculature and retinal progenitors. Lack of an abnormal retinal phenotype in silent heart mutants suggests that endothelial cells specifically, and not circulating factors, provide paracrine signals to the developing retina.


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