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E. Ivanova, G.-S. Hwang, Z.-H. Pan, D. Troilo; Evaluation of AAV-Mediated Expression of Channelrhodopsin-2 in the Marmoset Retina. Invest. Ophthalmol. Vis. Sci. 2009;50(13):1027.
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© ARVO (1962-2015); The Authors (2016-present)
Converting inner retinal neurons into photosensitive cells by expression of channelrhodopsin-2 (ChR2) offers a new approach for treatment of blinding retinal degenerations. Our previous study has reported ChR2 expression in inner retinal neurons in rodents mediated by recombinant adeno-associated virus (rAAV2/2) with a ubiquitous promoter. Here we evaluated the ability of expressing ChR2 in inner retinal neurons in a New World primate, the common marmoset (Callithrix jacchus).
rAAV2/2 vectors carrying a fusion construct of channelopsin-2 and GFP (Chop2-GFP) under the control of CAG or CMV promoter were injected into the intravitreal space of the eyes of adult marmosets (~7-month-old). The retinas were harvested 3 moths after the virus injection. The expression of GFP was examined in retinal whole-mounts and vertical sections.
Robust expression of Chop2-GFP was observed in the retinas injected with the virus vectors with either CAG or CMV promoter. The fluorescence was observed mostly in the inner retinal layers. The GFP was targeted to the plasma membranes and the morphology of Chop2-GFP-positive cells appeared normal. High concentration of virus (6 x 1012 GP/ml) with CAG promoter resulted in the expression of Chop2-GFP in numerous ganglion and amacrine cells. Some bipolar and horizontal cells and few rods and cones were also labeled. At a low concentration (2 x 1010 GP/ml) individual ganglion cells were labeled. Different types of ganglion cells were found based on their morphological properties. The virus with CMV promoter (1 x 1011 GP/ml) yielded GFP expression predominantly in ganglion and amacrine cells.
Using rAAV2/2 vectors with ubiquitous promoters, the expression of Chop-2-GFP can be achieved in the inner neurons of the marmoset retina. The ability of rAAV2/2 vectors to infect inner retinal neurons in primates offers the possibility of using rAAV-mediated gene delivery of Chop2 as a potential approach for treatment of blinding retinal degenerations in human. In addition, inserting Chop2-GFP into the membrane of individual ganglion cells in the primate retina allows studying their morphological and functional properties.
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