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Keplinger, Stefan Florian (2016): Establishing optogenetic tools in the auditory system of the Mongolian Gerbil. Dissertation, LMU München: Fakultät für Biologie



The Mongolian Gerbil (Meriones unguiculatus) serves as a popular and widely used model organism for the human auditory system. Its hearing range largely overlaps with that of human’s and even extends below 1 kHz, frequencies very important for human hearing. Like humans, gerbils can localize sounds based on their interaural time difference (ITD) or interaural level difference (ILD) and also show perceptual suppression of the spatial source of reverberations (precedence effect). The auditory circuitries underlying the computation of ITDs and ILDs are very well described in the gerbil, although the exact mechanisms for the extraction of ITDs are still under debate. The contribution of the medial nucleus of the trapezoid body (MNTB) in tuning neurons sensitive to ITDs is still unclear. Similarly, the precedence effect is well known and thought to greatly facilitate listening in reverberant environments, yet the neural substrate of the precedence effect is still elusive. A circuitry that might subserve the precedence effect is hypothesized to be formed by the dorsal nucleus of the lateral lemniscus (DNLL) and the inferior colliculus (IC). However, a precise and reversible manipulation of the DNLL-IC circuitry or the ITD circuitry has not been possible due to the lack of technical means. With the advent of optogenetics, tools are becoming available that would allow to specifically activate and silence nuclei within both circuitries. Yet, transgenic lines or genetic tools are neither disposable nor established for the Mongolian Gerbil. Hence, in order to express optogenetic tools in the gerbil auditory brainstem and midbrain, a reliable and neuron specific gene delivery system needs to be established as a major prerequisite. Only when this important first step is taken, the actual optogenetical tools can be applied and tested. In this study, the first hurdle of gene delivery into the Mongolian Gerbil was successfully cleared by using recombinant adeno-associated viruses (rAAV) as vectors. Via the stereotactic injection of rAAVs into the DNLL, IC and MNTB, not only reliable and efficient transduction of neurons was achieved but also neuronal specific expression of transgenes was attained. As a second accomplishment, the channelrhodopsin mutant CatCH as well as the halorhodopsin NpHR3.0 were characterized in acute brain slices by performing whole cell patch-clamp recordings of transduced neurons. As a final step and proof of principle experiment, sound evoked neural responses in the DNLL and IC were successfully manipulated with light in vivo, as could be demonstrated by single cell extracellular recordings from anaesthetized animals. In sum, this study successfully adapted and established gene delivery and optogenetic tools in the auditory system of the Mongolian Gerbil. This represents a fully functional and highly versatile toolbox that not only paves the way to further elucidate the ITD as well as the DNLL-IC circuitry but is also applicable to other questions.