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H. Tomita, E. Sugano, H. Isago, T. hiroi, Z. Wang, M. Tamai; Visual Responses of Royal College of Surgeons Rats Transferred Modified Volvox Channelrhodopsin-2 Gene. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3465.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the differences of light sensitivity in channelrhodopsin-2 derived from the Clamydomonas and the Volvox (ClChR2 and VChR2), we measured visually evoked potentials (VEPs) in RCS rats transferred either ClChR2 or VChR2.
We made an adeno-associated virus vector type 2 including the ClChR2 or VChR2 gene fused to a fluorescent protein, Venus (AAV-ClChR2V) or pmCherry (AAV-VChR2cherry) respectively. Each AAV vector was injected intravitreally into the eyes of 6-month-old RCS (rdy/rdy) rats. Visual function was evaluated by recording visually evoked potentials (VEPs) with various stimulus patterns. Retinal sections and flat-mounted retinas were made for histological examinations.
VEPs were elicited from RCS rats transferred ClChR2 or VChR2. However amplitudes of VChR2 transferred rats were significantly lower than those of ClChR2 transferred rats. We observed the difference of expression pattern in flat-mounted retina between of the ClChR2 and VChR2 transferred rat. The expression of ClChR2 was mainly observed in plasma membrane of retinal ganglion cells. On the other hand, the diffuse expression of VChR2 was seen in the cell body of retinal ganglion cells and observed as scattered fluorescence. We made a modified VChR2 (mVChR2) by the exchanging the N-terminal fragment of VChR2 into a part of the Clamydomonas ChR1. Amplitudes of VEPs in RCS rats transferred mVChR2 increased. Flat-mounted retina showed decreased scattered fluorescence in the cell body.
These results indicate that light sensitivity can be improved by modifying the channelrhodopsin gene, and mVChR2 is also useful for restoring vision.
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