Restoring sight with the help of optogenetics

Back in December, I talked a bit about the history of optogenetics and some of its promising applications. Optogenetics is a biological technology that induces light sensitivity in cells. Depending on which optogenetic molecule you express, light can cause different reactions. Recently, some ingenious scientists crafted a new optogenetic tool and have made strides in bringing sight back to blind mice suffering from retinal degeneration. Retinal degenerative diseases such as retinitis pigmentosa and age-related macular degeneration affect 1 out of every 300 people. These diseases cause rods and cones, the photo-receptive cells in the retina, to slowly die. Luckily, cells downstream of the photo-receptors like bipolar cells and retinal ganglion cells remain healthy and can be targets for treatments. Michiel van Wyk and his colleagues from the University of Bern expressed their optogenetic molecule in bipolar cells to manipulate them into sensing light.

Using optogenetics to cure blindness is not a novel idea. Researchers in the past have expressed a popular optogenetic tool named channelrhodopsin in bipolar cells. Although this allowed the normally photo-insensitive cells to respond to light, the technique had some big caveats. Channelrhodopsin requires an immense amount of light to become active. Bipolar cells expressing channelrhodopsin needed roughly 10¹⁵ photons/s*cm to be activated. To put that into perspective, it is equivalent to being outside in a field of snow on a sunny day. That much light isn't sustainable or healthy for the cells. Not only does it require bright light, but the scientists had to introduce molecules into the bipolar cells that weren't normally there. The molecular machinery necessary to make channelrhodopsin work isn't normally present in bipolar cells. It is far less efficient to introduce novel molecules and pathways into a cell than it is to hijack part of the cells natural processes.

To overcome these obstacles, researchers engineered a new optogenetic molecule called Opto-mGluR6. This tool combines halves of two proteins: melanopsin and mGluR6. Melanopsin is a photosensitive protein found in mammals that isn't used for vision, but regulates the sleep-wake cycle. The other half of the protein, mGluR6, is a metabotropic glutamate receptor. At its most basic, mGluR6 responds to the neurotransmitter glutamate by activating downstream signaling networks. One downstream effect of mGluR6 activation is the closing of cation channels. MGluR6 signaling has the added advantage of amplifying the signal it receives. One mGluR6 protein can activate multiple downstream targets, starting a cascade of signaling. By combining melanopsin with mGluR6, they created a chimeric protein able to react to light by closing excitatory cation channels. On the left, you can see a diagram of the protein as it sits inside of a cell membrane. The researchers created a mouse that would express this protein only in bipolar cells within the retina. Normally, bipolar cells carry signals from photoreceptors in the back of the retina towards retinal ganglion cells (displayed in the diagram above). Expressing opto-mGluR6 in healthy bipolar cells allows them to percieve light and project directly to the retinal ganglion cells.

With this new molecule, the researchers were able to address the problems found in previous studies. The least amount of light that created a viable signal from the bipolar cells was 5x10¹¹ photons/s*cm², which is roughly equivalent to the amount of light at dusk. This is a drastic improvement compared to using channelrhodopsin. Also, the signaling activated by the mGluR6 portion of the protein is found naturally in bipolar cells. So, not only does the signal amplify small amounts of photo-activation, but all of the mechanisms are present in the cell already. These improvements make the technique far easier and more efficient to use.

To show that this technique could be used to improve vision, they ran several tests in rd1 mutant mice with and without opto-mGluR6 expression. Rd1 mice have early onset retinal degeneration and make for a realistic disease model. First, the scientists visualized the blood oxygen levels in visual cortex of opto-mGluR6 expressing rd1 mice when shown visual stimuli. Increased blood oxygen suggests that region of the brain has increased activity, which correlates to visual recognition. This is shown in the figure to the right, with darker colors indicating more activation. To test the mice's visual acuity in a real-life scenario, they used a visually guided swim task. Mice were put into a pool of opaque water with a submerged platform signaled by LED lights. If a mouse were unable to see, one would expect them to swim around at random searching for a platform to rest on. If the mice could see, they would learn that the light signals where the platform is and swim directly towards it. The scientists found that blind rd1 mice with opto-mGluR6 could use the light to find the platform just as well as visually capable mice. The blind rd1 mice without opto-mGluR6, however, had severe difficulties in finding the platform. Data from this experiment is shown to the left. The black lines show examples of the swimming path of a single mouse. Mice expressing Opto-mGluR6 swam far less distance to find the platform because they could see the indicating light. These tests show that formerly blind mice could now perform tasks just as well as visually competent mice.

The authors of this paper made fantastic progress in optimizing optogenetic techniques and improving the vision of blind mice. This is an enormous accomplishment. Before this could become a realistic option for those suffering from retinal degenerative diseases, we would need to ensure there are no aversive effects and understand what exactly the brain is visualizing. It is difficult to tell at this point what the mice are seeing exactly, and just how much acuity the engineered bipolar cells have. Further study should look into what aspects of vision this technique recovers. Can the mice perceive color, movement, or basic shapes? By studying this in depth it will also help optimize the process, and point us towards more efficient ways to use optogenetic tools. It is exciting to see a basic science technology that felt novel so recently begin to have such broad impacts in the medical world.

References:

Doroudchi, M., Greenberg, K., Liu, J., et al. Virally delivered Channelrhodopsin-2 safely and effectively restores visual function in multiple mouse models of blindness. Molecular Therapy, 19, 1220-1229 (2011).

http://www.medicaldaily.com/three-blind-mice-no-more-optogenetics-restores-vision-lab-rats-could-soon-cure-332772

Lagali, P., Balya, D., Awatramani, G., et al. Light-activated channels targeted to ON bipolar cells restore visual function in retinal degeneration. Nature Neuroscience 11, 667-675 (2008).

Lin, B., Koizumi, A., Tanaka, N., Panda, S., Masland, R. Restoration of visual function in retinal degeneration mice by ectopic expression of melanopsin. PNAS, 105, 16009-16014 (2008).

Wyk, M.V., Pielecka-Fortuna, J., Lowel, S. & Kleinloel, S. Restoing the ON Switch in Blind Retinas: Opto-mGluR6, a Next-Generation, Cell-Tailored Optogenetic Tool. PLOS Biology, 13, 1-30 (2015).

Image credits:

The top image comes from millipores retinal antibody page (http://www.emdmillipore.com/US/en/life-science-research/antibodies-assays/neuroscience/differentiation/vision/K16b.qB.bvQAAAFCgcBYZna_,nav?bd=1) and is augmented by the author

the bottom 3 images are augmented from the M.V. Wyk et al. paper cited above.