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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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Ultrastructural analysis of the inner retina blood supply in zebrafish. A: Light microscopy image of blood vessels (highlighted by a red square) overlying the inner limiting membrane (ILM) of the retina in adult zebrafish. 40× magnification. B: Ultrastructure of capillary. The basal lamina (BL) is in contact with the ILM enclosing pericytes (PE) and the vascular endothelium. Scale bar: 5 μm. C: Typical structural features of pericytes, e. g. Golgi apparatus (GA), rough endoplasmic reticulum (RER) and large membrane bound vesicles. Scale bar: 1 μm. D: Vitreal space lined by pericyte overlying endothelial cells (EC) which display interdigitating junctional complexes (arrows). Scale bar: 500 nm. E: Multiple vesicles (arrows) from 20 to 250 nm contacting the inner limiting membrane of the retina indicate active interaction between the vessels and the ganglion cell layer (at the Müller endfeet). Scale bar: 1 μm. F: Vascular (top) and ganglion cell layer interface (bottom). A vesicle (left panel) apparently separated from the cell membrane, fuses with the cell membrane when the section is tilted by 43° (right panel) suggesting transcellular transport. Scale bar: 500 nm. G: Interendothelial junctions are significantly more open in the senescent fish. Distance between endothelial cells at tight (asterisks) and adherent (arrows) junctions were measured in different peripheral and central areas of retinas from senescent and young adult fish. Error bars: sem. Inset in H: Histology of a 5 dpf zebrafish eye. 40× magnification. H: EM image of a 5 dpf larval eye illustrating the relationship of hyaloid vessels (encircled by a red line) to both the lens and the retina. Scale bar: 10 μm. I: Higher magnification of the hyaloid vessel confirms its tight attachment to the lens and looser contact to the retina (arrows). Scale bar: 1 μm. MüE: Müller cell endfeet; GC: ganglion cell; GCL: ganglion cell layer; VL: vascular layer; RPE: retinal pigmented epithelium; RET: retina.
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Figure 3: Ultrastructural analysis of the inner retina blood supply in zebrafish. A: Light microscopy image of blood vessels (highlighted by a red square) overlying the inner limiting membrane (ILM) of the retina in adult zebrafish. 40× magnification. B: Ultrastructure of capillary. The basal lamina (BL) is in contact with the ILM enclosing pericytes (PE) and the vascular endothelium. Scale bar: 5 μm. C: Typical structural features of pericytes, e. g. Golgi apparatus (GA), rough endoplasmic reticulum (RER) and large membrane bound vesicles. Scale bar: 1 μm. D: Vitreal space lined by pericyte overlying endothelial cells (EC) which display interdigitating junctional complexes (arrows). Scale bar: 500 nm. E: Multiple vesicles (arrows) from 20 to 250 nm contacting the inner limiting membrane of the retina indicate active interaction between the vessels and the ganglion cell layer (at the Müller endfeet). Scale bar: 1 μm. F: Vascular (top) and ganglion cell layer interface (bottom). A vesicle (left panel) apparently separated from the cell membrane, fuses with the cell membrane when the section is tilted by 43° (right panel) suggesting transcellular transport. Scale bar: 500 nm. G: Interendothelial junctions are significantly more open in the senescent fish. Distance between endothelial cells at tight (asterisks) and adherent (arrows) junctions were measured in different peripheral and central areas of retinas from senescent and young adult fish. Error bars: sem. Inset in H: Histology of a 5 dpf zebrafish eye. 40× magnification. H: EM image of a 5 dpf larval eye illustrating the relationship of hyaloid vessels (encircled by a red line) to both the lens and the retina. Scale bar: 10 μm. I: Higher magnification of the hyaloid vessel confirms its tight attachment to the lens and looser contact to the retina (arrows). Scale bar: 1 μm. MüE: Müller cell endfeet; GC: ganglion cell; GCL: ganglion cell layer; VL: vascular layer; RPE: retinal pigmented epithelium; RET: retina.

Mentions: The ultrastructure of the zebrafish vitreo-retinal vessels was examined to elucidate cellular interactions with the lens and retina (Fig 3). Sections of retinas from adults confirm the presence of a vascular system in direct contact with the inner limiting membrane and the ganglion cell layer (Fig 3A). Vascular endothelial cells are interconnected forming interdigitating junctional complexes (Fig 3B–D). Multiple mature pericytes with Golgi networks and vacuoles (Fig 3B–C) are found associated with the vessels in all young and senescent specimens, but not in the larvae (Fig 3H). The pericytes are located between the vascular endothelium and the basal lamina of the retinal vessels in direct contact with both (Fig 3B–C), as observed in mammalian retinas [5,32]. At the inner limiting membrane, numerous vesicles ranging from ~20–250 nm are coupled to the cell membranes at both the vascular and the ganglion cell sides (Fig 3E). These vesicles fuse with the plasma membranes (Fig 3F) suggesting nourishment by active transcellular transport, equivalent to the vesicle vacuolar organelles (VVO) observed in mammalian retinas [6].


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)

Ultrastructural analysis of the inner retina blood supply in zebrafish. A: Light microscopy image of blood vessels (highlighted by a red square) overlying the inner limiting membrane (ILM) of the retina in adult zebrafish. 40× magnification. B: Ultrastructure of capillary. The basal lamina (BL) is in contact with the ILM enclosing pericytes (PE) and the vascular endothelium. Scale bar: 5 μm. C: Typical structural features of pericytes, e. g. Golgi apparatus (GA), rough endoplasmic reticulum (RER) and large membrane bound vesicles. Scale bar: 1 μm. D: Vitreal space lined by pericyte overlying endothelial cells (EC) which display interdigitating junctional complexes (arrows). Scale bar: 500 nm. E: Multiple vesicles (arrows) from 20 to 250 nm contacting the inner limiting membrane of the retina indicate active interaction between the vessels and the ganglion cell layer (at the Müller endfeet). Scale bar: 1 μm. F: Vascular (top) and ganglion cell layer interface (bottom). A vesicle (left panel) apparently separated from the cell membrane, fuses with the cell membrane when the section is tilted by 43° (right panel) suggesting transcellular transport. Scale bar: 500 nm. G: Interendothelial junctions are significantly more open in the senescent fish. Distance between endothelial cells at tight (asterisks) and adherent (arrows) junctions were measured in different peripheral and central areas of retinas from senescent and young adult fish. Error bars: sem. Inset in H: Histology of a 5 dpf zebrafish eye. 40× magnification. H: EM image of a 5 dpf larval eye illustrating the relationship of hyaloid vessels (encircled by a red line) to both the lens and the retina. Scale bar: 10 μm. I: Higher magnification of the hyaloid vessel confirms its tight attachment to the lens and looser contact to the retina (arrows). Scale bar: 1 μm. MüE: Müller cell endfeet; GC: ganglion cell; GCL: ganglion cell layer; VL: vascular layer; RPE: retinal pigmented epithelium; RET: retina.
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Related In: Results  -  Collection

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Figure 3: Ultrastructural analysis of the inner retina blood supply in zebrafish. A: Light microscopy image of blood vessels (highlighted by a red square) overlying the inner limiting membrane (ILM) of the retina in adult zebrafish. 40× magnification. B: Ultrastructure of capillary. The basal lamina (BL) is in contact with the ILM enclosing pericytes (PE) and the vascular endothelium. Scale bar: 5 μm. C: Typical structural features of pericytes, e. g. Golgi apparatus (GA), rough endoplasmic reticulum (RER) and large membrane bound vesicles. Scale bar: 1 μm. D: Vitreal space lined by pericyte overlying endothelial cells (EC) which display interdigitating junctional complexes (arrows). Scale bar: 500 nm. E: Multiple vesicles (arrows) from 20 to 250 nm contacting the inner limiting membrane of the retina indicate active interaction between the vessels and the ganglion cell layer (at the Müller endfeet). Scale bar: 1 μm. F: Vascular (top) and ganglion cell layer interface (bottom). A vesicle (left panel) apparently separated from the cell membrane, fuses with the cell membrane when the section is tilted by 43° (right panel) suggesting transcellular transport. Scale bar: 500 nm. G: Interendothelial junctions are significantly more open in the senescent fish. Distance between endothelial cells at tight (asterisks) and adherent (arrows) junctions were measured in different peripheral and central areas of retinas from senescent and young adult fish. Error bars: sem. Inset in H: Histology of a 5 dpf zebrafish eye. 40× magnification. H: EM image of a 5 dpf larval eye illustrating the relationship of hyaloid vessels (encircled by a red line) to both the lens and the retina. Scale bar: 10 μm. I: Higher magnification of the hyaloid vessel confirms its tight attachment to the lens and looser contact to the retina (arrows). Scale bar: 1 μm. MüE: Müller cell endfeet; GC: ganglion cell; GCL: ganglion cell layer; VL: vascular layer; RPE: retinal pigmented epithelium; RET: retina.
Mentions: The ultrastructure of the zebrafish vitreo-retinal vessels was examined to elucidate cellular interactions with the lens and retina (Fig 3). Sections of retinas from adults confirm the presence of a vascular system in direct contact with the inner limiting membrane and the ganglion cell layer (Fig 3A). Vascular endothelial cells are interconnected forming interdigitating junctional complexes (Fig 3B–D). Multiple mature pericytes with Golgi networks and vacuoles (Fig 3B–C) are found associated with the vessels in all young and senescent specimens, but not in the larvae (Fig 3H). The pericytes are located between the vascular endothelium and the basal lamina of the retinal vessels in direct contact with both (Fig 3B–C), as observed in mammalian retinas [5,32]. At the inner limiting membrane, numerous vesicles ranging from ~20–250 nm are coupled to the cell membranes at both the vascular and the ganglion cell sides (Fig 3E). These vesicles fuse with the plasma membranes (Fig 3F) suggesting nourishment by active transcellular transport, equivalent to the vesicle vacuolar organelles (VVO) observed in mammalian retinas [6].

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