May 2006
Volume 47, Issue 13
ARVO Annual Meeting Abstract  |   May 2006
Controlled Photoreceptor Ablation in Transgenic X. Laevis
Author Affiliations & Notes
  • L.M. Hamm
    Ophthalmology, University of British Columbia, Vancouver, BC, Canada
  • B.M. Tam
    Ophthalmology, University of British Columbia, Vancouver, BC, Canada
  • O.L. Moritz
    Ophthalmology, University of British Columbia, Vancouver, BC, Canada
  • Footnotes
    Commercial Relationships  L.M. Hamm, None; B.M. Tam, None; O.L. Moritz, Ariad Pharmaceuticals, Inc, F.
  • Footnotes
    Support  MSFHR, CIHR, FFB–Canada, and the Karl Kirchgessner Foundation
Investigative Ophthalmology & Visual Science May 2006, Vol.47, 803. doi:
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      L.M. Hamm, B.M. Tam, O.L. Moritz; Controlled Photoreceptor Ablation in Transgenic X. Laevis . Invest. Ophthalmol. Vis. Sci. 2006;47(13):803.

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

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Purpose: : Following rod death in retinitis pigmentosa, cone cells die by an uncharacterized secondary pathway, which may be due to direct rod–cone interactions, or indirect effects of rod depletion. In order to study this phenomenon, we have developed a transgenic model with inducible rod cell apoptosis. This model will allow us to precisely control the initiation of rod cell death, and subsequently monitor the affects on neighboring cone cells.

Methods: : We generated primary transgenic X. laevis that express a inducible form of HA epitope–tagged caspase–9 (I–caspase–9) under the control of the X. laevis opsin promoter. I–caspase–9 has binding domains specific for the compound AP20187 (Ariad Pharmaceuticals), which induces its dimerization and autoactivation, resulting in the initiation of the apoptotic cascade. Transgenic animals were treated with AP20187 from 7 to 14 days post fertilization and the localization of I–caspase–9 in photoreceptors was examined by immunofluorescence microscopy. The effect of AP20187 administration on rod viability was examined using confocal microscopy and dot blot assays.

Results: : Immunofluorescence microscopy demonstrated that I–caspase–9 was expressed exclusively in rod photoreceptors. In response to administration of AP20187, we observed complete elimination of the epitope tag signal, indicating cleavage of I–caspase–9. Dot blot analysis indicated that treated animals also had significantly reduced levels of rhodopsin, indicating rod cell death. We also observed morphological evidence of apoptosis in response to administration of AP20178. These effects were not observed in control animals.

Conclusions: : This is a promising model of retinal degeneration that will be useful for a number of applications, including investigations of the mechanisms underlying secondary cone degeneration, such as that seen in retinitis pigmentosa. Our future studies will include investigations of the effects of induced rod ablation on cone viability using biochemical and electrophysiological assays.

Keywords: apoptosis/cell death • photoreceptors • transgenics/knock-outs 

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