May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
Determining Retinal Cell Birth Order in the Frog, Xenopus laevis, by Clonal Analysis
Author Affiliations & Notes
  • L.L. Wong
    Surgery Dept-Anatomy Div 0604, University California-San Diego, La Jolla, CA, United States
  • D.H. Rapaport
    Surgery Dept-Anatomy Div 0604, University California-San Diego, La Jolla, CA, United States
  • Footnotes
    Commercial Relationships  L.L. Wong, None; D.H. Rapaport, None.
  • Footnotes
    Support  EY11875
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 4299. doi:
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      L.L. Wong, D.H. Rapaport; Determining Retinal Cell Birth Order in the Frog, Xenopus laevis, by Clonal Analysis . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4299.

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

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Abstract

Abstract: : Purpose:The developmental program of the vertebrate eye is well conserved. Studies on retinal cell genesis in monkey, rat, and mouse showed that the seven major cell types are generated in a sequential order, albeit with significant overlap between cell types. Such ordered cell genesis suggests that changes in retinal progenitor cell competence plays a major role in fate acquisition. Xenopus laevis has easily accessible optic vesicles that are highly amenable to targeted gene expression, thus an ideal system to study mechanisms of fate acquisition. However, due to the short duration of retinogenesis (50 h), studies using conventional methods were unable to resolve any order of cell genesis in Xenopus. In addition, the significant overlap of retinal cell type production found in mammals fails to distinguish models between progressive restriction of cell competence that suggests temporal regulation or merely stochastic switching of cell competence. In this study, we maximized temporal resolution by analyzing labeled clones in the developing Xenopus retina. Methods:GFP cDNA was transfected into the presumptive optic vesicle of Stg 20 Xenopus embryos to label clones. Between Stgs 24 to 37, we injected BrdU into these embryos to determine the birth order within each clone. We analyzed coronal sections of Stg 41 embryos when the retina was cytoarchitecturally mature. Majority of clones formed spatially segregated tight clusters. Cell type was determined by laminar location, cell morphology (apparent from GFP filling) and rods were identified by anti-rhodopsin immunoreactivity. Cells that were BrdU- had become post-mitotic before the stage of injection while BrdU+ cells were still mitotic at that time. Results:Our data revealed two striking findings: 1) Xenopus retinal cell type production is not random; cell birthdating within clones is consistent with a specific temporal order of cell type genesis and this order is unidirectional, 2) this sequential order of cell genesis is similar to the proposed sequence found in other vertebrate species, including birds and mammals. Conclusions:The non-overlapping, unidirectional production of retinal cell types suggests that mechanisms for retinal cell fate acquisition involve temporal factors such that a retinal progenitor cell viewed as a developmental unit spins off post-mitotic cells of different types and becomes progressively restricted in competence with each cell division cycle. Further, if the sequence of retinal cell genesis reflects the underlying mechanisms of cell type determination, we found that the mechanisms are well conserved in all vertebrates.

Keywords: retinal development • immunohistochemistry • retina 
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