May 2007
Volume 48, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2007
Evaluation of Virus Mediated Long-Term Expression of Channelrhodopsin-2 in the Mouse Retina
Author Affiliations & Notes
  • E. Ivanova
    Anatomy and Cell Biology, Wayne State Univ Sch of Med, Detroit, Michigan
  • A. Bi
    Anatomy and Cell Biology, Wayne State Univ Sch of Med, Detroit, Michigan
  • Z.-H. Pan
    Anatomy and Cell Biology, Wayne State Univ Sch of Med, Detroit, Michigan
  • Footnotes
    Commercial Relationships E. Ivanova, None; A. Bi, None; Z. Pan, None.
  • Footnotes
    Support EY17130
Investigative Ophthalmology & Visual Science May 2007, Vol.48, 613. doi:
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      E. Ivanova, A. Bi, Z.-H. Pan; Evaluation of Virus Mediated Long-Term Expression of Channelrhodopsin-2 in the Mouse Retina. Invest. Ophthalmol. Vis. Sci. 2007;48(13):613.

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

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Abstract

Purpose:: The feasibility of converting inner retina neurons to photosensitive cells via viral mediated expression of a directly-light sensitive channel, channelrhodopsin-2 (ChR2), offers a new potential approach for treatment of blinding retinal degeneration. Here we evaluated the effects of virus concentration and light conditions on the expression and the potential toxicity of ChR2 in the mouse retina.

Methods:: Adeno-associated virus (AAV2) vectors carrying a fusion construct of channelopsin-2 (Chop2) and GFP (Chop2-GFP) in different concentrations were injected into the intravitreal space of the eyes of wild-type adult and newborn (P1-3) mice. The retinas were harvested up to 20 months after the virus injection for patch-clamp and immunocytochemistry studies. The expression of GFP in different cell types was examined in retinal whole-mounts and vertical sections. The density of the cells located in the ganglion cell layer was estimated based on the DAPI staining and taken as a major criterion for evaluating the potential toxicity of expressing Chop2-GFP. In addition to the normal light conditions (1014 photons cm-2s-1), some injected mice were exposed to strong blue light (1016 photons cm-2s-1; 460 nm) for 12 h, 1 week, and 2 weeks. The retinas of these mice were examined two weeks after the exposure.

Results:: The expression of Chop2-GFP was stable for up to 20 months after the virus injection. Chop-GFP was observed mainly in ganglion cells and AII amacrine cells. Occasionally, Chop2-GFP was observed predominantly in photoreceptors. The infection rate was dependent on virus concentration. The concentration of the virus and the percentage of Chop2-GFP-positive cells have no effect on the density and thus, survival of the cells in the ganglion cell layer. Some retinas had an increased density of the cells in the restricted areas probably caused by the surgery procedure. The morphology of the GFP-positive cells remained normal. The physiological properties of AII amacrine cells expressing ChR2 were similar to those of the uninfected cells. In the experiments where animals were housed under bright light conditions the density of the cells also did not change relatively to the control group.

Conclusions:: Expression of Chop2-GFP did not cause any detectable toxicity and cell death in the retina independent from the virus concentration, infection rate, and duration of the expression. Our results indicate that ChR2 is biocompatible as a light sensor in te retina.

Keywords: retinal degenerations: cell biology • gene transfer/gene therapy • ganglion cells 
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