Bioluminescent Multi-Characteristic Opsin for simultaneous optical stimulation and large-scale monitoring of the visual system

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The optogenetic stimulation strategy, which is an alternative to electrical stimulation and now under clinical trial for vision restoration in retinal degenerative diseases, involves simple intravitreal injection of safe AAV-carrying opsin to photosensitize higher order retinal neurons. Compared with other strategies, optogenetic method is cell-specific and provides higher resolution. Our optogenetic studies in dry-AMD mice model suggests that degenerated retina can be made highly photosensitive, enabling vision restoration at low light level. Furthermore, our preclinical studies have shown that the visually guided behavior deteriorated with progressive degeneration, which recovered to more non-degenerated level after optogenetic treatment. Though primary visual cortex is known to maintain its retinotopy in subjects with retinal degeneration despite prolonged visual loss, detailed knowledge of how optogenetic sensitization of higher order neurons manifests in restoration of visual cortical activity is currently lacking. Thus, there is a need for mapping changes in the visual cortical activities as progression of retinal degeneration and subsequent vision restoration by optogenetic sensitization of retina occurs. The goal of this study is to develop and characterize tools for simultaneous optical modulation and imaging of retinal and cortical activities using spectrally-separated activation and detection channels. To address this goal, we propose to use newly developed voltage-sensitive bioluminescence assay instead of fluorescence in order to allow simultaneous long-term cortical imaging upon visual stimulation using multi- characteristic opsin (MCO). This will enable us to modulate neural activity in retina and image visual cortex with high temporal and spatial resolution, providing details about disease progression and restoration. The innovativeness of our proposal includes bioluminescent membrane voltage indicator to measure neural activity in long-term studies with high temporal resolution. This new bioluminescent technique does not require an additional potentially phototoxic external excitation source (as used for fluorescence). Further innovativeness includes simultaneous measurements of cortical activities and behavior in response to visual stimulation during retinal degeneration and vision restoration. Towards this goal we will: (i) Demonstrate functioning of Bioluminescent MCO and hardware for stimulation and bioluminescent neural activity monitoring; (ii) Quantify changes in visual cortical activities during progression of retinal photoreceptor degeneration by bMCO using head-mountable sCMOS-camera; and (iii) Evaluate restoration of visual cortical activities upon re- photosensitization of degenerated retina by intravitreal injection of MCO. Success of this proposal will lead to development of a modular and scalable interface system with the capability to serve a multiplicity of applications to modulate and monitor large-scale activity in the nervous system. In the long term, we aim to demonstrate a clinically viable optical stimulation device that can reliably provide stimulation inputs to the visual cortex and simultaneous recording of cortical activities.