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Genetic determinants of hyaloid and retinal vasculature in zebrafish.

Alvarez Y, Cederlund ML, Cottell DC, Bill BR, Ekker SC, Torres-Vazquez J, Weinstein BM, Hyde DR, Vihtelic TS, Kennedy BN - BMC Dev. Biol. (2007)

Bottom Line: Similar to the transient hyaloid vasculature in mammalian embryos, vessels are first found attached to the zebrafish lens at 2.5 days post fertilisation.Finally, we identify 9 genes with cell membrane, extracellular matrix and unknown identity that are necessary for zebrafish hyaloid and retinal vasculature development.Zebrafish have a retinal blood supply with a characteristic developmental and adult morphology.

View Article: PubMed Central - HTML - PubMed

Affiliation: UCD School of Biomolecular, and Biomedical Sciences, University College Dublin, Dublin 4, Ireland. yolanda.alvarez@ucd.ie

ABSTRACT

Background: The retinal vasculature is a capillary network of blood vessels that nourishes the inner retina of most mammals. Developmental abnormalities or microvascular complications in the retinal vasculature result in severe human eye diseases that lead to blindness. To exploit the advantages of zebrafish for genetic, developmental and pharmacological studies of retinal vasculature, we characterised the intraocular vasculature in zebrafish.

Results: We show a detailed morphological and developmental analysis of the retinal blood supply in zebrafish. Similar to the transient hyaloid vasculature in mammalian embryos, vessels are first found attached to the zebrafish lens at 2.5 days post fertilisation. These vessels progressively lose contact with the lens and by 30 days post fertilisation adhere to the inner limiting membrane of the juvenile retina. Ultrastructure analysis shows these vessels to exhibit distinctive hallmarks of mammalian retinal vasculature. For example, smooth muscle actin-expressing pericytes are ensheathed by the basal lamina of the blood vessel, and vesicle vacuolar organelles (VVO), subcellular mediators of vessel-retinal nourishment, are present. Finally, we identify 9 genes with cell membrane, extracellular matrix and unknown identity that are necessary for zebrafish hyaloid and retinal vasculature development.

Conclusion: Zebrafish have a retinal blood supply with a characteristic developmental and adult morphology. Abnormalities of these intraocular vessels are easily observed, enabling application of genetic and chemical approaches in zebrafish to identify molecular regulators of hyaloid and retinal vasculature in development and disease.

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Adult-onset retinal vasculature phenotypes in Plexin D1 mutants. A-B: Pseudo-coloured adult retinas from (A) wild type and (B) obd adult fish to facilitate quantification of the vessel branches. A black circumferential line demarks half the distance between the optic disc and the annular peripheral vein at different retinal regions (MID- retina). C: The number of main branches radiating from the optic disc and the number of vessels crossing the mid-retina are higher in obd mutants (n = 18) than in wild type retinas (n = 24). D: The average number of branch points per vessel is significantly decreased in obd (n = 5) versus wildtype (n = 5) retinas. The number of branch points per 9 randomly chosen vessels in each retina was counted from outside in. To avoid bias, the nearest vessel was chosen every 40° of retinal circumference, starting from the most ventral poin. E: A significant reduction in the distance from the optic disc branches to the secondary branches, but not to the tertiary branches, is observed in obd vessels (n = 40) versus wildtypes (n = 22).
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Figure 5: Adult-onset retinal vasculature phenotypes in Plexin D1 mutants. A-B: Pseudo-coloured adult retinas from (A) wild type and (B) obd adult fish to facilitate quantification of the vessel branches. A black circumferential line demarks half the distance between the optic disc and the annular peripheral vein at different retinal regions (MID- retina). C: The number of main branches radiating from the optic disc and the number of vessels crossing the mid-retina are higher in obd mutants (n = 18) than in wild type retinas (n = 24). D: The average number of branch points per vessel is significantly decreased in obd (n = 5) versus wildtype (n = 5) retinas. The number of branch points per 9 randomly chosen vessels in each retina was counted from outside in. To avoid bias, the nearest vessel was chosen every 40° of retinal circumference, starting from the most ventral poin. E: A significant reduction in the distance from the optic disc branches to the secondary branches, but not to the tertiary branches, is observed in obd vessels (n = 40) versus wildtypes (n = 22).

Mentions: The plexin D1 receptor is mutated in out of bounds (obd) mutants resulting in patterning defects of intersegmental vessels [27,42]. In obd larvae, patterning defects of the hyaloid vessels are subtle at 3 dpf but by 5 dpf the vessels distinctly form abnormal branches and extra interconnections (Fig 4E). The phenotype is variable in juvenile (~30 dpf) where the hyaloid vasculature is often undistinguishable from wild type. However, in adult obd zebrafish increased tortuosity of the retinal vasculature is consistently observed, with highly contorted looping not found in wild type retinas (Fig 4F and Additional File 2). Compared to wild types, adult obd retinae have a significantly higher number of vessels at the mid-retina (Fig 5C and Additional File 2). The obd phenotype is not a result of more frequent vessel branching across all of the retina, as the average number of branches per vessel is not increased, but in fact is significantly decreased (Fig 5D). The higher number of retinal vessels in obd mutants stems from a significantly increased number of primary branches radiating from the optic disc (Fig 5C) and a significantly reduced distance to the secondary branch (Fig 5E).


Genetic determinants of hyaloid and retinal vasculature in zebrafish.

Alvarez Y, Cederlund ML, Cottell DC, Bill BR, Ekker SC, Torres-Vazquez J, Weinstein BM, Hyde DR, Vihtelic TS, Kennedy BN - BMC Dev. Biol. (2007)

Adult-onset retinal vasculature phenotypes in Plexin D1 mutants. A-B: Pseudo-coloured adult retinas from (A) wild type and (B) obd adult fish to facilitate quantification of the vessel branches. A black circumferential line demarks half the distance between the optic disc and the annular peripheral vein at different retinal regions (MID- retina). C: The number of main branches radiating from the optic disc and the number of vessels crossing the mid-retina are higher in obd mutants (n = 18) than in wild type retinas (n = 24). D: The average number of branch points per vessel is significantly decreased in obd (n = 5) versus wildtype (n = 5) retinas. The number of branch points per 9 randomly chosen vessels in each retina was counted from outside in. To avoid bias, the nearest vessel was chosen every 40° of retinal circumference, starting from the most ventral poin. E: A significant reduction in the distance from the optic disc branches to the secondary branches, but not to the tertiary branches, is observed in obd vessels (n = 40) versus wildtypes (n = 22).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2169232&req=5

Figure 5: Adult-onset retinal vasculature phenotypes in Plexin D1 mutants. A-B: Pseudo-coloured adult retinas from (A) wild type and (B) obd adult fish to facilitate quantification of the vessel branches. A black circumferential line demarks half the distance between the optic disc and the annular peripheral vein at different retinal regions (MID- retina). C: The number of main branches radiating from the optic disc and the number of vessels crossing the mid-retina are higher in obd mutants (n = 18) than in wild type retinas (n = 24). D: The average number of branch points per vessel is significantly decreased in obd (n = 5) versus wildtype (n = 5) retinas. The number of branch points per 9 randomly chosen vessels in each retina was counted from outside in. To avoid bias, the nearest vessel was chosen every 40° of retinal circumference, starting from the most ventral poin. E: A significant reduction in the distance from the optic disc branches to the secondary branches, but not to the tertiary branches, is observed in obd vessels (n = 40) versus wildtypes (n = 22).
Mentions: The plexin D1 receptor is mutated in out of bounds (obd) mutants resulting in patterning defects of intersegmental vessels [27,42]. In obd larvae, patterning defects of the hyaloid vessels are subtle at 3 dpf but by 5 dpf the vessels distinctly form abnormal branches and extra interconnections (Fig 4E). The phenotype is variable in juvenile (~30 dpf) where the hyaloid vasculature is often undistinguishable from wild type. However, in adult obd zebrafish increased tortuosity of the retinal vasculature is consistently observed, with highly contorted looping not found in wild type retinas (Fig 4F and Additional File 2). Compared to wild types, adult obd retinae have a significantly higher number of vessels at the mid-retina (Fig 5C and Additional File 2). The obd phenotype is not a result of more frequent vessel branching across all of the retina, as the average number of branches per vessel is not increased, but in fact is significantly decreased (Fig 5D). The higher number of retinal vessels in obd mutants stems from a significantly increased number of primary branches radiating from the optic disc (Fig 5C) and a significantly reduced distance to the secondary branch (Fig 5E).

Bottom Line: Similar to the transient hyaloid vasculature in mammalian embryos, vessels are first found attached to the zebrafish lens at 2.5 days post fertilisation.Finally, we identify 9 genes with cell membrane, extracellular matrix and unknown identity that are necessary for zebrafish hyaloid and retinal vasculature development.Zebrafish have a retinal blood supply with a characteristic developmental and adult morphology.

View Article: PubMed Central - HTML - PubMed

Affiliation: UCD School of Biomolecular, and Biomedical Sciences, University College Dublin, Dublin 4, Ireland. yolanda.alvarez@ucd.ie

ABSTRACT

Background: The retinal vasculature is a capillary network of blood vessels that nourishes the inner retina of most mammals. Developmental abnormalities or microvascular complications in the retinal vasculature result in severe human eye diseases that lead to blindness. To exploit the advantages of zebrafish for genetic, developmental and pharmacological studies of retinal vasculature, we characterised the intraocular vasculature in zebrafish.

Results: We show a detailed morphological and developmental analysis of the retinal blood supply in zebrafish. Similar to the transient hyaloid vasculature in mammalian embryos, vessels are first found attached to the zebrafish lens at 2.5 days post fertilisation. These vessels progressively lose contact with the lens and by 30 days post fertilisation adhere to the inner limiting membrane of the juvenile retina. Ultrastructure analysis shows these vessels to exhibit distinctive hallmarks of mammalian retinal vasculature. For example, smooth muscle actin-expressing pericytes are ensheathed by the basal lamina of the blood vessel, and vesicle vacuolar organelles (VVO), subcellular mediators of vessel-retinal nourishment, are present. Finally, we identify 9 genes with cell membrane, extracellular matrix and unknown identity that are necessary for zebrafish hyaloid and retinal vasculature development.

Conclusion: Zebrafish have a retinal blood supply with a characteristic developmental and adult morphology. Abnormalities of these intraocular vessels are easily observed, enabling application of genetic and chemical approaches in zebrafish to identify molecular regulators of hyaloid and retinal vasculature in development and disease.

Show MeSH
Related in: MedlinePlus