Optogenetic Manipulation of the Auditory System
by Burak Bali
Date of Examination:2021-01-15
Date of issue:2021-12-14
Advisor:Prof. Dr. Tobias Moser
Referee:Dr. Manuela Schmidt
Referee:Dr. Jens Gruber
Referee:Arezoo PhD Pooresmaeili
Referee:Prof. Dr. Tim Gollisch
Referee:Prof. Dr. Alexander Flügel
Persistent Address:
http://dx.doi.org/10.53846/goediss-9001
Files in this item
Name:PhD_Dissertation-Burak_Bali_withoutCV.pdf
Size:8.47Mb
Format:PDF
The following license files are associated with this item:
Abstract
English
There are millions of people who are affected from sensorineural hearing loss. In this type of deafness, sensory hair cells are typically dysfunctional or lost, and thereby not conveying sound information to brain. The state-of-the-art solution is the cochlear implant (CI) which circumvents the receptor problem via direct electric stimulation of spiral ganglion neurons (SGNs) of that from auditory nerve (AN). Thus, the CI enables hearing and provides open speech comprehension. Yet, for instance in noisy background, in most users, speech comprehension fails likely owing to poor frequency resolution. This results from wide current spread in the the saline-filled cochlea activating large sets of the tonotopically organized SGNs. Here, light can serve as an alternative mode of stimulation since it can be conveniently focused to evoke SGN populations more precisely. Clearly, SGNs are not light sensitive and hence we employ an optogenetic approach. The series of studies presented here gathers under the same roof: How to achieve optogenetic manipulation of the SGNs such that it will suit well to the intrinsic physiological properties of auditory system, be stably maintained for many years, and last but not least be safe? Temporal firing properties of SGNs are uniquely fast allowing fine coding of sound. Therefore, channelrhodopsins (ChRs) used should enable reliable control of light-induced spiking in SGNs. To this end, first, we investigated feasibility of blue-light-gated naturally occurring Chronos with the fastest on-/off-kinetics at the mouse AN with the help of molecular tools, i.e. endoplasmic reticulum exit signal (ES) and membrane trafficking signal (TS), enhancing membrane localization of the ChR. By this way, it was possible to drive the AN up to 1000 Hz stimulation rate. While Chronos suits temporal coding very well, we also sought for red-shifted fast alternatives since chronic exposure to blue light bears risk of phototoxicity in SGNs. Here, the engineered red-shifted very-fast-Chrimson (vf-Chrimson-ES/TS) which has similar fast kinetics as fast as Chronos-ES/TS, evoked fast responses until 500 Hz stimulation rate from the mouse AN. Moreover, we further characterized light intensity encoding of vf-Chrimson-ES/TS in comparison to the slower variant fast-Chrimson (f-Chrimson). Third, we studied the long-term availability and biosafety profile of f-Chrimson, a prominent candidate for use with the optical CI, showing expression in SGNs up to two years following early postnatal AAV-based delivery to the inner ear. There was also a tendency of SGN loss due to combinatorial effect of aging and f-Chrimson expression. Expression spread to the non-injection ear and parts of the brain. The spread was restricted to the vicinity of injection and was not detected in the distal organs such as kidney and spleen. All in all, these three projects that I was involved in at different extents pave the way to future medical optogenetic cochlear implants.
Keywords: channelrhodopsin, gene therapy, auditory nerve, hearing, deafness, cochlear implant
