title: Gucy2f zebrafish knockdown--a model for Gucy2d-related leber congenital amaurosis.
abstract: Mutations in retinal-specific guanylate cyclase (Gucy2d) are associated with Leber congenital amaurosis-1 (LCA1). Zebrafish offer unique advantages relative to rodents, including their excellent color vision, precocious retinal development, robust visual testing strategies, low cost, relatively easy transgenesis and shortened experimental times. In this study we will demonstrate the feasibility of using gene-targeting in the zebrafish as a model for the photoreceptor-specific GUCY2D-related LCA1, by reporting the visual phenotype and retinal histology resulting from Gucy2f knockdown. Gucy2f zebrafish LCA-orthologous cDNA was identified and isolated by PCR amplification. Its expression pattern was determined by whole-mount in-situ hybridization and its function was studied by gene knockdown using two different morpholino-modified oligos (MO), one that blocks translation of Gucy2f and one that blocks splicing of Gucy2f. Visual function was assessed with an optomotor assay on 6-days-post-fertilization larvae, and by analyzing changes in retinal histology. Gucy2f knockdown resulted in significantly lower vision as measured by the optomotor response compared with uninjected and control MO-injected zebrafish larvae. Histological changes in the Gucy2f-knockdown larvae included loss and shortening of cone and rod outer segments. A zebrafish model of Gucy2f-related LCA1 displays early visual dysfunction and photoreceptor layer dystrophy. This study serves as proof of concept for the use of zebrafish as a simple, inexpensive model with excellent vision on which further study of LCA-related genes is possible.
authors.affiliation: Neuro-Ophthalmology Unit, Department of Ophthalmology, Rabin Medical Center, Petah Tikva, Israel. kalishhadas@gmail.com
authors.forename: Hadas
authors.initials: H
authors.lastname: Stiebel-Kalish
authors.forename: Ehud
authors.initials: E
authors.lastname: Reich
authors.forename: Nir
authors.initials: N
authors.lastname: Rainy
authors.forename: Gad
authors.initials: G
authors.lastname: Vatine
authors.forename: Yael
authors.initials: Y
authors.lastname: Nisgav
authors.forename: Anna
authors.initials: A
authors.lastname: Tovar
authors.forename: Yoav
authors.initials: Y
authors.lastname: Gothilf
authors.forename: Michael
authors.initials: M
authors.lastname: Bach
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D004195:Disease Models, Animal; D005260:Female; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D006162:Guanylate Cyclase / Q000235:genetics*; D057130:Leber Congenital Amaurosis / Q000235:genetics* / Q000473:pathology; D008297:Male; D011956:Receptors, Cell Surface / Q000235:genetics*; D058499:Retinal Dystrophies / Q000235:genetics / Q000473:pathology; D014785:Vision, Ocular / Q000235:genetics; D015027:Zebrafish / Q000235:genetics*
chemical_list: D011956:Receptors, Cell Surface; C110715:guanylate cyclase 1; D006162:Guanylate Cyclase