Search

Cochlear implants restore hearing to people with hearing impairments all over the world. Although this technology has drastically improved their quality of life, users still struggle to understand speech in noisy environments and appreciate music. This is due to current spread throughout the cochlea – a fundamental limitation of electrical stimulation – and research into alternative stimulation methods is necessary to achieve better outcomes for cochlear implant users. Alternatives include the use of light stimulation enabled by optogenetics, either on its own or combined with electrical stimulation. Much research to date has been focused on the spatial precision of cochlear stimulation, but temporal precision is also important for encoding key features of sounds, including pitch and timbre. Research to date has demonstrated that optogenetic stimulation has significantly poorer temporal precision than electrical stimulation, and it is unreliable at the high rates of stimulation needed for cochlear implants. It remains unclear whether combined optogenetic-electrical stimulation has high temporal precision and reliability at the stimulation rates needed to encode pitch and timbre. This thesis aims to establish a baseline of the temporal precision of auditory nerve responses for these new stimulation technologies and considers the impact of important factors for clinical translation. Namely, the studies here consider the effects of auditory nerve degeneration following deafness, viral transduction efficiency, and opsin characteristics. The results of the studies presented in this thesis demonstrate that, although optogenetic stimulation has poor temporal precision and reliability at 100 pulses per second (pps) and above, combined stimulation remains reliable and demonstrates electrical-like temporal precision up to 400 pps. Nerve degeneration caused by chronic deafness resulted in poorer reliability for optogenetic stimulation and higher power requirements for combined

Bei der Presbyakusis oder Altersschwerhörigkeit handelt es ich um einen irreversiblen, progressiven, bilateralen symmetrischen sensorineuralen Hörverlust, der auf eine Degeneration verschiedener Innenohrstrukturen zurückzuführen ist. Aufgrund der hohen Prävalenz und der negativen Auswirkung auf die Lebensqualität ist es dringend erforderlich, die Ätiologie der Presbyakusis genauer zu verstehen. Ziel dieses Projektes ist es, den Zusammenhang von Calcium bindenden Proteinen und der Altersschwerhörigkeit genauer zu analysieren. Calcium bindende Proteine regulieren die intrazelluläre Calciumkonzentration in Neuronen und schützen die Zellen vor einem Calciumüberschuss. Im Innenohr zeigt sich eine heterogene Verteilung von Calretinin, Calbindin und Parvalbumin. Calcium abhängige neuronale Prozesse spielen eine wichtige Rolle bei der neuronalen Alterung und Degeneration. In dieser Studie werden altersbedingte Veränderungen der Immunreaktivität von Calretinin, Calbindin und Parvalbumin in Spiralganglienzellen untersucht. In diese Studie wurden 16 C57BL/6J- und 16 129/SvJ-Mäuse unterschiedlichen Alters (2, 4, 7 und 12 Monate) eingeschlossen. Die Hörschwellen wurden anhand der auditorischen Hirnstammableitung gemessen. Anschließend wurden die Cochleae histologisch aufgearbeitet und eine semi-quantitative Immunhistochemie für die zuvor genannten Calcium bindenden Proteine durchgeführt. Die Hörschwellen von C57BL/6J- und 129/SvJ-Mäusen stiegen im Alter von 7 Monaten signifikant an. Die durchschnittliche Immunreaktivität von Calbindin stieg sowohl mit dem Alter als auch mit der relativen Anzahl positiver Zellen signifikant an, jedoch wurden keine signifikanten Veränderungen bei Calretinin und Parvalbumin beobachtet. Die Hochregulierung von Calbindin könnte als Schutz dienen, um Funktionsdefizite auszugleichen, die mit dem Alter auftreten. Die Expression von Calretinin und Parvalbumin scheint sich bei murinen Innenohren bis zu einem Alter von 12 Monaten in C57BL/6J und 129/SvJ ni, Age-related hearing loss is an irreversible progressive bilateral symmetrical sensorineural hearing loss resulting from degeneration of different inner ear structures. No preventative or curative measures are known at this time. Because of the high prevalence and the impact on life quality there is an urgency to understand the aetiologies of hearing loss. The aim of this project is to understand more about the impact of calcium binding proteins on hearing function. Calcium binding proteins in neurons buffer intracellular free Ca2+ ions, which interacts with proteins controlling enzymatic and ion channel activity. The heterogeneous distribution of Calretinin, Calbindin, and Parvalbumin influence calcium homeostasis and calcium related neuronal processes play an important role in neuronal aging and degeneration. This study evaluates age related changes in Calretinin, Calbindin, and Parvalbumin immune reactivity in spiral ganglion cells. In this study 16 C57BL/6J and 16 129/SvJ mice at different ages (2, 4, 7 and 12 months) were included. Hearing thresholds were assessed using auditory brainstem response before inner ears were excised for further evaluation. Semi-quantitative immunohistochemistry for the aforementioned calcium-binding proteins were performed at the cellular level. Hearing thresholds of C57BL/6J and 129/SvJ mice increased significantly by 7 months of age. The average immune reactivity of Calbindin increased significantly with aging as well as the relative number of positive cells, but no significant alterations in Calretinin and Parvalbumin were observed. Upregulation of Calbindin could serve as protection to compensate for functional deficits that occur with aging. Expression of both Calretinin and Parvalbumin seems to be stabilizing factors in murine inner ears up to an age of 12 months in C57BL/6J and 129/SvJ., submitted by Dr. med. univ. Natalie Fischer, Zusammenfassung in deutscher Sprache, Dissertation Medical University of Innsbruck 2019

The loss of auditory neurons that occurs with profound hearing loss is irreversible in humans. Stem cell therapy for auditory neuron replacement thereby offers potential for hearing restoration in patients with profound deafness. This project aims to investigate and quantify the formation of new synapses between human embryonic stem cell-derived neurons and target peripheral and central auditory tissues in vitro. Sensory neuron-like cells were generated from human embryonic stem cells over 21 days in vitro and then co-cultured with developing sensory hair cells or cochlear nucleus slices for a further 2 weeks. Synapse formation was examined using four-channel immunofluorescence and confocal microscopy, and analysed using ImageJ software. Sites of apparent contact between stem cell-derived neurons and target auditory tissues were compared to synapses in the developing mammalian peripheral and central auditory tissues. Stem cell-derived neurons expressed synapsin 1 and vesicular glutamate transporter-1 at sites of innervation with both hair cells and cochlear nucleus neurons. This observation is consistent with results obtained from control co-cultures using early postnatal auditory neurons and auditory tissues. Significantly more synaptic contacts were observed in stem cell co-cultures in comparison to control co-cultures (P <0.05). Moreover, potential synapses from stem cell-derived neurons corresponded anatomically with early synaptogenesis in the developing mammalian auditory system in vivo. This assay describes the characterisation, timing and quantification of potential synaptic connections both by stem cell-derived neurons in vitro and in the developing mammalian auditory system.

Release of neurotrophin-3 (NT3) and brain-derived neurotrophic factor (BDNF) from hair cells in the cochlea is essential for the survival of spiral ganglion neurons (SGNs). Loss of hair cells associated with a sensorineural hearing loss therefore results in degeneration of SGNs, potentially reducing the performance of a cochlear implant. Exogenous replacement of either or both neurotrophins protects SGNs from degeneration after deafness. We previously incorporated NT3 into the conducting polymer polypyrrole (Ppy) synthesized with para-toluene sulfonate (pTS) to investigate whether Ppy/pTS/NT3-coated cochlear implant electrodes could provide both neurotrophic support and electrical stimulation for SGNs. Enhanced and controlled release of NT3 was achieved when Ppy/pTS/NT3-coated electrodes were subjected to electrical stimulation. Here we describe the release dynamics and biological properties of Ppy/pTS with incorporated BDNF. Release studies demonstrated slow passive diffusion of BDNF from Ppy/pTS/BDNF, with electrical stimulation significantly enhancing BDNF release over 7 days. A 3-day SGN explant assay found that neurite outgrowth from explants was 12.3-fold greater when polymers contained BDNF (p < 0.001), although electrical stimulation did not increase neurite outgrowth further. The versatility of Ppy to store and release neurotrophins, conduct electrical charge, and act as a substrate for nerve-electrode interactions is discussed for specialized applications such as cochlear implants.