June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Defects in cone photoreceptor development and progressive retinal dystrophy in rd3 and abca4b knockout stable zebrafish models
Author Affiliations & Notes
  • Divya Ashok Pidishetty
    Center for Ocular Regeneration (CORE), LV Prasad Eye Institute, Hyderabad, Telangana, India
    Manipal Academy of Higher Education, Manipal, Karnataka, India
  • Praveen Joseph Susaimanickam
    Center for Ocular Regeneration (CORE), LV Prasad Eye Institute, Hyderabad, Telangana, India
  • Santosh Damera
    Center for Ocular Regeneration (CORE), LV Prasad Eye Institute, Hyderabad, Telangana, India
  • Sreedhar Rao Boyenpally
    Ophthalmic Pathology Laboratory, LV Prasad Eye Institute, Hyderabad, Telangana, India
  • Gopal Kushawah
    Centre for Cellular and Molecular Biology CSIR, Hyderabad, Telangana, India
  • Rakesh Mishra
    Centre for Cellular and Molecular Biology CSIR, Hyderabad, Telangana, India
  • Indumathi Mariappan
    Center for Ocular Regeneration (CORE), LV Prasad Eye Institute, Hyderabad, Telangana, India
  • Footnotes
    Commercial Relationships   Divya Pidishetty None; Praveen Joseph Susaimanickam None; Santosh Damera None; Sreedhar Rao Boyenpally None; Gopal Kushawah None; Rakesh Mishra None; Indumathi Mariappan None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 1370 – F0301. doi:
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      Divya Ashok Pidishetty, Praveen Joseph Susaimanickam, Santosh Damera, Sreedhar Rao Boyenpally, Gopal Kushawah, Rakesh Mishra, Indumathi Mariappan; Defects in cone photoreceptor development and progressive retinal dystrophy in rd3 and abca4b knockout stable zebrafish models. Invest. Ophthalmol. Vis. Sci. 2022;63(7):1370 – F0301.

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

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Abstract

Purpose : To knockout zebrafish rd3 and abca4b genes linked to retinal dystrophies and to study the effects of loss of function on retinal development and visual functions

Methods : Zebrafish rd3 and abca4b gene knockout models were generated by CRISPR-Cas9 based genome editing. The edits in the founder animals (F0) was confirmed by T7 endonuclease assay followed by Sanger sequencing. Homozygous mutants were generated by interbreeding of heterozygotes. The wild type, rd3-/-, abca4b-/- mutant fishes were euthanized at 3, 6 and 12 months of age and their retinal morphology was evaluated by immunohistochemistry. Visual assessment of these animals was performed using different functional test paradigms

Results : We report that the retina of rd3-/- mutants at 3 months have underdeveloped cones, with missing outer limiting membrane and lamination defects in the outer nuclear layer. Both rods and cones degenerate at later time points (6 and 12 months), with major loss in cone subtypes. In abca4b-/- mutants, a marked reduction in the cone nuclear layer and a corresponding increase in the number of rod nuclei suggest significant defects in cone precursor cell differentiation and maturation. The single blue cones are completely absent and the UV cones are underdeveloped with rudimentary inner and outer segments. The double cones develop normally, but undergo degeneration in older animals. In experimental paradigms to assess the feed capture response under normal photopic conditions (n=10), the mutant fishes failed to approach their feed/prey and displayed significantly higher latency than the wild type fishes. This visual behavioral deficit corroborates with the cone photoreceptor-specific developmental anomaly and retinal degeneration in mutant fishes

Conclusions : The zebrafish rd3-/- and abca4b-/- mutants show cone developmental defects and degenerative phenotypes that resemble the human disease. Defects in photoreceptor precursor cell differentiation, maturation and lamination indicates possible role(s) for these genes in the early retinal development

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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