All-optical recording and stimulation of retinal neurons in vivo in retinal degeneration mice

Abstract

Here we demonstrate the application of a method that could accelerate the development of novel therapies by allowing direct and repeatable visualization of cellular function in the living eye, to study loss of vision in animal models of retinal disease, as well as evaluate the time course of retinal function following therapeutic intervention. We use high-resolution adaptive optics scanning light ophthalmoscopy to image fluorescence from the calcium sensor GCaMP6s. In mice with photoreceptor degeneration (rd10), we measured restored visual responses in ganglion cell layer neurons expressing the red-shifted channelrhodopsin ChrimsonR over a six-week period following significant loss of visual responses. Combining a fluorescent calcium sensor, a channelrhodopsin, and adaptive optics enables all-optical stimulation and recording of retinal neurons in the living eye. Because the retina is an accessible portal to the central nervous system, our method also provides a novel non-invasive method of dissecting neuronal processing in the brain.

Adaptive optics: principles and applications in ophthalmology

“…AO-SLO detected shortened cone outer segments in the blue cone monochromacy mouse model, which was significantly restored after opsin gene augmentation therapy (134). Functional fluorescent AO imaging also showed extensive functional loss in photoreceptor-degenerated rd10 mice as compared to wild-type mice (135). More recently, Miller et al used multimodal imaging including AO-SLO to characterize short-term and long-term cellular interactions in the retina after focal photoreceptor damage (136).…”

Theoretical optimization of high-frequency optogenetic spiking of red-shifted very fast-Chrimson-expressing neurons

“…The vf-Chrimson opens up promising new possibilities to provide high-fidelity, high-frequency operation with high-spatiotemporal resolution in a wide range of biomedical applications in which red-shifted opsins have been used. [26][27][28][29][30][31][32][33] The proposed model of vf-Chrimson can also be integrated with circuit models of other types of cells to check their response in a quick way.…”

“…10 Moreover, red-shifted opsins have been used for a wide range of biomedical applications, including generation of behavior responses in mechanosensory neurons; increase in firing rates in primary visual cortex of mice that result in suppressing endogenous gamma; bimodal neural excitation of Caenorhabditis elegans; dual color control using low-noise multishank optoelectrodes; retinal degeneration in mice; restoration of vision in blind mice, macaque, and human retina; and defibrillation therapy for human heart using LED arrays and also for Ca 2þ imaging. [26][27][28][29][30][31][32][33] Recently, f-Chrimson has been used to develop optical cochlear implants to restore auditory nerve activity in deaf mice. 10 It is extremely important to develop a theoretical understanding of the biophysical mechanism of the photosensitization agents and their behavior in different cells, to design and control optogenetic devices and circuits.…”

Optogenetic Therapy for Visual Restoration

“…Tyrosine-mutated AAVs are generated through rational mutagenesis to improve intracellular nuclear trafficking, based on the finding that phosphorylation of surface-exposed tyrosine residues on AAV leads to ubiquitination and subsequent proteasome-mediated degradation. Intravitreal injection of AAV2 quadruple mutants (Y272, 444, 500, 730F; 4YF) has been demonstrated to transduce bipolar cells using either the Grm6 [ 16 , 20 , 72 ] or L7-5 promoter [ 24 , 51 ] in rodent models. When AAV2(4YF) was used with the hSyn-1 promoter, optogenetic gene expression was dominant in RGCs [ 70 ].…”