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K. P. Greenberg, A. Pham, A. Delwig, F. S. Werblin; Genetically Reconstructed Center-Surround Opponency by Targeting Channelrhodopsin-2 and Halorhodopsin to Ganglion Cell Somata and Dendrites. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3896.
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The insertion of directly light sensitive ion channels into normally non-photosensitive retinal neurons is a promising approach for restoring sight to profoundly blind individuals. Retinal degeneration results in the eventual deafferentation of the inner retina and remodeling of microcircuits underlying center-surround receptive field architecture, however ganglion cell somata and dendrites appear to remain intact. Center-surround antagonism is a fundamental property of the visual stream from the retina through the visual cortex. Therefore, we sought to restore center-surround antagonism directly to ganglion cells in a retina lacking photoreceptor input.
Two directly photosensitive microbial rhodopsins, channelrhodopsin-2 (ChR2) and halorhodopsin (eNpHR), were genetically targeted differentially to the soma and dendrites in rabbit ganglion cells, creating spatially distinct excitatory and inhibitory subcellular domains. These fluorescently tagged rhodopsins were delivered to intact rabbit retinae via biolistic gene transfer and subcellular localization was evaluated by confocal microscopy. Patterned light stimuli were used to map excitation and inhibition at the soma and dendrites of transfected ganglion cells, as measured by whole-cell patch clamp.
Robust ChR2-mediated excitatory currents were elicited under 460nm illumination at the soma, while 560nm illumination elicited eNpHR-mediated inhibitory currents at the dendrites. Both currents had a dynamic range of ~2 log units and their relative amplitudes were tunable by modulating the intensities of the two wavelengths of light. Somatic-targeted ChR2 exhibited significantly sharper Gaussian-shaped spatial tuning profiles (HWHM=60µm) when compared to ChR2 expressed throughout the entire membrane (HWHM=400µm). Opposing currents were measured with discrete spatial regions when ChR2 was targeted to the soma and eNpHR was targeted to the dendrites. We are evaluating the neural image generated from populations of these neurons in response to natural scenes through modeling and direct measurement.
We demonstrate that genetically reconstructed center-surround receptive field function can be implemented directly in ganglion cells by targeting opposing rhodopsins differentially to the soma or dendrites. Imparting direct concentric antagonistic light sensitivity directly to ganglion cells should convey enhanced contrast and edge sensitivity along the visual pathway to higher visual centers and may lead to a future therapy for those that have suffered substantial photoreceptor loss.
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