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Z.-H. Pan, E. Ivanova; Cell Type-Specific Targeting of Channelrhodopsin-2 in the Retina Through AAV-Mediated Cre/LoxP Recombination in Cre-Transgenic Mouse Lines. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3466.
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© ARVO (1962-2015); The Authors (2016-present)
Converting inner retinal neurons to photosensitive cells by expression of channelrhodopsin-2 (ChR2) opens the possibility of a new approach for treating blindness caused by retinal degeneration. The use of ChR2 to gain optical control of neuronal activity in the retina could also offer new tools for basic retinal research. In this study, we examined the ability and efficiency of achieving cell type-specific expression of ChR2 in the retina through adeno-associated virus (AAV)-mediated Cre/LoxP recombination.
Adeno-associated virus type 2 (AAV2) vectors carried a Cre-dependent switch (FLEX) and a reverse Chop2-mCherry fusion construct, AAV2-FLEX-rev-Chop2-mCherry, with ubiquitous promoter of CMV or EF1α. The AAV2 vectors were injected intravitreally into the eyes of several Cre-transgenic mouse lines in which the expression of Cre recombinase was found in certain specific inner retinal neurons. The expression of Chop2-mCherry was examined by fluorescence microscopy. Electrophysiological recordings were performed to study the ChR2-mediated light responses.
The use of AAV2-FLEX-rev-Chop2-mCherry vectors with EF1α promoter could achieve robust expression of Chop2-mCherry in Cre-expressing third-order retinal neurons but not in retinal bipolar cells. The combined use of CMV promoter and an AAV2 capsid mutant (Y444F) markedly increased the expression efficiency in retinal bipolar cells. Targeted expression of Chop2-mCherry was achieved in several specific types of third-order neurons, including starburst amacrine cells and bistratified ganglion cells. ChR2-mediated light responses were recorded from the targeted cells.
Targeted expression of ChR2-mCherry can be achieved in the retina through AAV2-mediated Cre/LoxP recombination in retinal cell type-specific Cre transgenic mice. The ability of achieving cell type-specific targeting of ChR2 in the retina enables to generate valuable animal models for evaluating ChR2-based gene therapy as well as for dissecting the structure and function of retinal circuits.
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