Opsin basierte Gentherapie zur Wiederherstellung der visuellen Mustererkennung in blinden Mäusen
Anatomie und Physiologie
Zusammenfassung der Projektergebnisse
Degenerative retinal disorders are among the leading causes of blindness in industrial countries. These include a spectrum of conditions ranging from rather common multi-factorial age-related macular degeneration to hereditary diseases like retinitis pigmentosa. Commonly, in these conditions the outer retina including the light-sensing photoreceptors are lost, while the inner retina remains intact. Establishing therapeutic approaches that could hold visual loss or even return visual function that has been lost is one major goal of ophthalmic research. One particularly topical strategy to achieve this goal consists of virally delivering optogenetic tools to the remaining cells of the inner retina to render those light sensitive. Different cellular targets (bipolar versus ganglion cells) and optogenetic tools (channelrhodopsins or mammalian opsins) have been proposed. The principal aim of this DFG Research Fellowship was to establish and apply methods that would allow a direct, quantifiable comparison between different optogenetic tools and cellular targets. I was able to successfully develop a setup (consisting of a high brightness, scalable light source, a pattern generating mirror device and a multi-electrode array) that enabled comparing to what extent different optogenetic strategies were able to restore encoding of key qualities of vision (e.g. brightness, contrast, receptive fields, direction selectivity) on the output level of the (mouse) retina. I have then employed this setup to determine the extent to which responses to complex light stimuli can be restored following viral delivery of red-shifted channelrhodopsin (ReaChR) in retina-degenerate (rd1) mice. While, in brief, time and space encoding was excellent, unfavourable high light intensities and a contrast of at least 30% were required to elicit reliable responses. The most exciting observation, however, was that response fidelity dropped over very short intervals of time (minutes) during sustained stimulation with quasi realistic stimuli. As this observation might contain a cue on why clinical trials using channelrhodopsins for vision restoration have yet not been able to report successful vision restoration in humans we have commenced a more detailed analysis of this phenomenon. While processing of the data is ongoing the current results indicate that this loss in response fidelity might be avoided by specifically targeting one type of retinal neuron: the rod bipolar cells. It was initially thought to complement this organ level analyses also with behavioural data. I therefore engineered an optokinetic drum-like device that would be bright enough to elicit visual responses also in mice treated with weakly light sensitive opsins. Despite long-term iterative optimization of this device, I finally did not manage to evoke any visually guided behavioural responses, neither in treated degenerate mice, nor in healthy sighted mice. The reasons therefore remain unknown. However, it is likely that either the high brightness or the noise of the device evoked fear responses that masked any visually guided behaviour. While pursuing the core objective of the Research Fellowship I was also able to successfully conduct a set of complementary laboratory and clinical projects. The clinical projects could already be converted into publication output. They support a better understanding of disease progression in advanced age-related macular degeneration and inherited macular disorders and help to quantitatively understand the relationship between morphologic disease progression and reading impairment in those patients. These data, in future, may help to elicit patients suitable for optogenetic gene therapeutic trials aiming on high end vision restoration and measure therapeutic success.
Projektbezogene Publikationen (Auswahl)
- (2019). Determinants of Reading Performance in Eyes with Foveal-Sparing Geographic Atrophy. Ophthalmology retina 3, 201-210
Lindner M, Pfau M, Czauderna J, Goerdt L, Schmitz-Valckenberg S, Holz FG & Fleckenstein M
(Siehe online unter https://doi.org/10.1016/j.oret.2018.11.005) - (2019). Foveal Sparing in Central Retinal Dystrophies. Investigative ophthalmology & visual science 60, 3456-3467
Bax NM, Valkenburg D, Lambertus S, Klevering BJ, Boon CJF, Holz FG, Cremers FPM, Fleckenstein M, Hoyng CB, Lindner M
(Siehe online unter https://doi.org/10.1167/iovs.18-26533) - (2019). Retinal imaging including OCT angiography for detecting active choroidal neovascularization in pseudoxanthoma elasticum. Clin Exp Ophthalmol.
Birtel J, Lindner M, Mishra DK, Muller PL, Hendig D, Herrmann P, Holz FG, Fleckenstein M, Gliem M & Issa PC
(Siehe online unter https://doi.org/10.1111/ceo.13385)
