Dataset Information

Spectrum of Fates: a new approach to the study of the developing zebrafish retina.

ABSTRACT: The ability to image cells live and in situ as they proliferate and differentiate has proved to be an invaluable asset to biologists investigating developmental processes. Here, we describe a Spectrum of Fates approach that allows the identification of all the major neuronal subtypes in the zebrafish retina simultaneously. Spectrum of Fates is based on the combinatorial expression of differently coloured fluorescent proteins driven by the promoters of transcription factors that are expressed in overlapping subsets of retinal neurons. Here, we show how a Spectrum of Fates approach can be used to assess various aspects of neural development, such as developmental waves of differentiation, neuropil development, lineage tracing and hierarchies of fates in the developing zebrafish retina.

SUBMITTER: Almeida AD

PROVIDER: S-EPMC3994774 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Spectrum of Fates: a new approach to the study of the developing zebrafish retina.

Almeida Alexandra D AD Boije Henrik H Chow Renee W RW He Jie J Tham Jonathan J Suzuki Sachihiro C SC Harris William A WA

Development (Cambridge, England) 20140409 9

The ability to image cells live and in situ as they proliferate and differentiate has proved to be an invaluable asset to biologists investigating developmental processes. Here, we describe a Spectrum of Fates approach that allows the identification of all the major neuronal subtypes in the zebrafish retina simultaneously. Spectrum of Fates is based on the combinatorial expression of differently coloured fluorescent proteins driven by the promoters of transcription factors that are expressed in ...[more]

PMID: 24718991

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Project description:BackgroundThe zebrafish retina maintains two populations of stem cells: first, the germinal zone or ciliary marginal zone (CMZ) contains multipotent retinal progenitors that add cells to the retinal periphery as the fish continue to grow; second, radial glia (Müller cells) occasionally divide asymmetrically to generate committed progenitors that differentiate into rod photoreceptors, which are added interstitially throughout the retina with growth. Retinal injury stimulates Müller glia to dedifferentiate, re-enter the cell cycle, and generate multipotent retinal progenitors similar to those in the CMZ to replace missing neurons. The specific signals that maintain these two distinct populations of endogenous retinal stem cells are not understood.ResultsWe used genetic and pharmacological manipulation of the ?-catenin/Wnt signaling pathway to show that it is required to maintain proliferation in the CMZ and that hyperstimulation of ?-catenin/Wnt signaling inhibits normal retinal differentiation and expands the population of proliferative retinal progenitors. To test whether similar effects occur during regeneration, we developed a method for making rapid, selective photoreceptor ablations in larval zebrafish with intense light. We found that dephosphorylated ?-catenin accumulates in Müller glia as they re-enter the cell cycle following injury, but not in Müller glia that remain quiescent. Activation of Wnt signaling is required for regenerative proliferation, and hyperstimulation results in loss of Müller glia from the INL as all proliferative cells move into the ONL.Conclusions?-catenin/Wnt signaling is thus required for the maintenance of retinal progenitors during both initial development and lesion-induced regeneration, and is sufficient to prevent differentiation of those progenitors and maintain them in a proliferative state. This suggests that the ?-catenin/Wnt cascade is part of the shared molecular circuitry that maintains retinal stem cells for both homeostatic growth and epimorphic regeneration.

Project description:Midkine is a heparin binding growth factor with important functions in neuronal development and survival, but little is known about its function in the retina. Previous studies show that in the developing zebrafish, Midkine-a (Mdka) regulates cell cycle kinetics in retinal progenitors, and following injury to the adult zebrafish retina, mdka is strongly upregulated in Müller glia and the injury-induced photoreceptor progenitors. Here we provide the first data describing Mdka protein localization during different stages of retinal development and during the regeneration of photoreceptors in adults. We also experimentally test the role of Mdka during photoreceptor regeneration. The immuno-localization of Mdka reflects the complex spatiotemporal pattern of gene expression and also reveals the apparent secretion and extracellular trafficking of this protein. During embryonic retinal development the Mdka antibodies label all mitotically active cells, but at the onset of neuronal differentiation, immunostaining is also localized to the nascent inner plexiform layer. Starting at five days post fertilization through the juvenile stage, Mdka immunostaining labels the cytoplasm of horizontal cells and the overlying somata of rod photoreceptors. Double immunolabeling shows that in adult horizontal cells, Mdka co-localizes with markers of the Golgi complex. Together, these data are interpreted to show that Mdka is synthesized in horizontal cells and secreted into the outer nuclear layer. In adults, Mdka is also present in the end feet of Müller glia. Similar to mdka gene expression, Mdka in horizontal cells is regulated by circadian rhythms. After the light-induced death of photoreceptors, Mdka immuonolabeling is localized to Müller glia, the intrinsic stem cells of the zebrafish retina, and proliferating photoreceptor progenitors. Knockdown of Mdka during photoreceptor regeneration results in less proliferation and diminished regeneration of rod photoreceptors. These data suggest that during photoreceptor regeneration Mdka regulates aspects of injury-induced cell proliferation.

Dataset Information

Spectrum of Fates: a new approach to the study of the developing zebrafish retina.

Publications

Spectrum of Fates: a new approach to the study of the developing zebrafish retina.

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