3D ultrastructural analysis of the subcellular organisation of inner hair cells and of their innervation during ageing.

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Hearing deteriorates progressively with age. This results from dysfunction of the cells in the cochlea of the inner ear that detect sound (the sensory cells, so-called "hair" cells), and/or the nerves that carry the signals to the brain. The hearing deficits are permanent. An early manifestation is difficulty in understanding speech in noise, then reduced ability to hear quieter and higher frequency sounds that progressively worsens with decreasing sensitivity to louder sounds and lower frequencies. An estimated 70% of those over retirement age suffer some form of hearing impairment. It is a major factor in reducing their quality of life. Amelioration of the condition is currently limited to technological solutions such as hearing aids and cochlear implants. There are no drug treatments. In order to improve the effectiveness of the technologies and to discover pharmacological targets for treatments, a better understanding of the progressive changes to the hair cells and nerves that occur with ageing is essential.Recent work has suggested that a major factor contributing to the deterioration of hearing with age is a slowly progressive loss, beginning quite early in life, of the nerve fibres that normally transmit sound information from the cochlea to the brain (afferent fibres), and which connect with the sensory cell - the inner hair cell (IHC) - that converts sound signals into the electrical signals that stimulate the nerves. There may also be re-organisation of the nerves ("efferent" fibres) that carry information from the brain to the afferent nerve terminals to modulate the auditory signals reaching the brain. In addition there is evidence of changes to the internal organisation of the IHCs themselves. In our current research we have applied two techniques for three-dimensional electron microscopy (3DEM) to characterise of the internal structures of IHC and the distribution of the contacting nerves fibres around it. With these techniques, we have demonstrated a previously unrecognised degree of structural organisation in IHC and have been able to map the distribution of all the afferent nerve terminals around the cell and the internal organisation, a feat not previously accomplished but made possible by the application the 3DEM technologies. We have found a continuous network of cellular membranes and organelles that appears to be structurally related to the synaptic machinery of the cell (the interface between the cell and the nerve where signals are transmitted) and to the position of the nerve terminals. Preliminary work has also shown disruptions to intracellular organisation of IHC that appear to be commensurate with loss nerve fibre loss during ageing. In the proposed project we will extend these techniques to fully characterise age-related changes to the cells and their innervation. We will also develop the use of a third 3DEM technique, array tomography, which offers the potential for routine 3D analysis of tissues without the need for the specialised microscopes that the other two techniques require. The central aim of the project is to map at the cellular and sub-cellular levels the changes to the internal structures of IHCs in the ageing cochlea and to examine the relationship between those and the loss of afferent terminals. We will also identify the compensatory changes to IHC and to the remaining afferent nerves, and the re-organisation of the efferent nerves. The project will provide a comprehensive assessment of the cellular basis of the deterioration of hearing with age.This application also will bring together researchers with expertise in electron microscopy and 3D analysis, cochlear anatomy, cochlear physiology and the molecular and genetic bases of deafness. This co-ordinated approach is designed to generate new approaches and future collaborations to further research into the life-course of the auditory system.