Structural organisations underlying auditory sensitivity

Most cellular functions depend upon assemblies of interacting molecules, and upon the ways in which those complexes are organised within a cell. Moreover, cells themselves are organised to compartmentalise different activities into different regions while at the same time enabling the integration of those activities that is necessary for the cell to function as it should. There is, thus, a very close relationship between cellular architecture and cell function. Consequently, crucial to understanding how different molecules are integrated into a cell to support its function, and how cells go wrong when molecular function is disrupted during disease, is knowledge of the three-dimensional cellular architecture at levels of resolution that enable visualisation of macromolecular assemblies and the cellular context in which they normally function. Electron microscopy reveals such structural details across a range from the approximate dimension of macromolecules up to the level of the whole cell. Three-dimensional information can be obtained from tissue sections examined by electron microscopy by the application of now established methods for 'electron tomography'. In this technique, a collection of images of the same structure is taken from many different angles. When these images are assembled together, a three-dimensional view of the structure is obtained. From this it is possible to identify how structures are associated with one another, for example how one molecular complex interacts with another, or at a lower level of resolution, how organelles within a cell are distributed and whether there is continuity between them. If tissue is prepared for microscopy by means that preserve the natural state, which can be achieved by freezing them very rapidly before processing, it becomes possible to obtain details of sub-cellular structures in a close-to-life condition in their true context inside the cell. This project will apply these modern methods to assess features of the cellular architecture of the sensory tissues of the inner ear. These tissues are responsible for the sense of hearing and maintenance of balance and they are remarkably three-dimensional in their architecture. Individual sensory 'hair' cells are cylindrical and bear at their top ends an organised bundle of projections, deflections of which in response to sound vibrations (hearing) or motion (balance) lead to signalling to the nerves at the bottom of the cell. Each hair cell is surrounded by cells that provide structural support. The project will explore the organisation of structures crucial to the cell's ability to faithfully generate appropriate neural output in response to mechanical input. It will determine the structure and relationships of macromolecular complexes that, with deflections of the hair bundle, control the opening and closing of channels through which a current that triggers the neural stimulation flows. It will determine the organisation of the structural components within the supporting cells that create the rigid framework that is necessary to ensure that small vibrations from quiet sounds produce deflections of the bundle and signal detection. And it will define the pathway within a hair cell by which the chemical that is released to stimulate the nerve travels from where it is made at the top end of the cell to the bottom end. Abnormalities of the molecules associated with these activities cause hair cell dysfunction. Hair cell dysfunction and loss are the main causes of deafness and balance disequilibrium, major disabling conditions that are particularly prevalent in the elderly. By elucidating details of the relationships between structure and function, this project will contribute to understanding the fundamental bases underlying hair cell dysfunction. From such knowledge means to ameliorate the resultant physiological deficits -deafness and/or balance disequilibrium- will ensue.

Electron tomography will be applied to sensory tissues of the inner ear to obtain high resolution 3-D images of the architecture of hair cells and supporting cells. Tissue will be prepared using high pressure freezing to obtain rapid freeze-fixation and preservation in a close-to-life condition, followed by freeze-substitution. The structure of the tip-link between stereocilia, and of its interactions at the site of the transduction channel at the bottom end and with the presumed adaptation complex at the top end, will be explored. Comparisons of tip link structures between auditory and vestibular hair cells and between mammals and non-mammals will be made, to test the hypothesis that the tip link maybe adapted for differing physiological demands. The studies are aimed at resolving current controversies over tip link structure and how the link can act as a gating spring to control opening of the transduction channel. The structure and interactions of lateral links between stereocilia will also be examined to determine how these links contribute to hair bundle stiffness. The 3-D organisation of the microtubule bundles in pillar cells of the organ of Corti will be determined to gain insight into their contribute to the micromechanical properties of the sensory epithelium. In addition, the project will examine the architectural organisation of intracellular membrane system and the cytoskeleton in inner hair cells to identify the possible pathways through which the population of neurotransmitter vesicles associated with ribbon synapses is maintained. It will determine whether there are continuous membrane compartments through the inner hair cells from the apical supranuclear region to the basal region from which the neurotransmitter vesicles that are tethered at the synaptic ribbon might derive, and whether there are organised microtubule networks along which vesicles traffic from the membrane system to the synaptic ribbon.

Who will benefit from the research? 1. People with hearing impairment and balance disequilibrium 2. Clinical professionals 3. UK and other companies producing cochlear prostheses and auditory diagnostics 4. The wider public. How will they benefit from this research? Hearing loss and balance dysfunction are major disabling conditions. They are particularly prevalent in the elderly population. More than 50% of people over 60 years old have hearing loss sufficient to impair normal communication. Balance dysfunction is a major contributor to falls in the elderly, which cost the NHS more than £1billion, and it causes dizziness, the most common reason for visits to a GP by elderly people. By contributing to detailed understanding of how hair cells function this project will lay foundations for addressing the fundamental bases of dysfunctions that lead to deafness and balance disequilibrium. Thus in the longer term the project will contribute to determining means to alleviate these conditions and improving the quality of life for elderly people and the many younger sufferers, as well as relieving the economic and social burden that they impose, with significant benefits to the wider public. In the shorter term, the results of the project will be beneficial clinically in enabling doctors and health workers to inform their patients better about the nature of their disease. The close association of the Ear Institute with the Royal National Throat Nose and Ear hospital provides a ready conduit for bringing the results of the scientific research to the attention of the clinical community. Professor Forge already has several professional relationships with this group and also is a regular speaker to interested professional and patients groups about his research. The work is also of benefit to manufacturers of instrumentation for objective measurement of hearing. This technology is being developed for early detection of hearing problems and the identification of susceptible ears but requires knowledge of the structures most intimately involved in hair cell function, the focus of the present study, to allow their physical behaviour to be modelled and simulated. From this details of the signals that reveal hair cell activity in response to sound can be interpreted. The leading company in this field is UK based and was established by a researcher in what is now the Ear Institute, so ties are close. Likewise companies producing cochlear implants, devices inserted into a deaf ear that can partially restore hearing, are interested in the fundamental basis of hair cell function in order to improve the effectiveness, and extend the beneficial use, of implants. We are already working with one such company to explore preservation of residual hearing after implantation and have contacts with others. These relationship afford opportunities for direct knowledge transfer. The project will also provide an attractive research opportunity for excellent young scientists looking for multi-disciplinary areas of discovery and thereby retain talented young researchers in the UK. In addition, transferable skills - such as time- and project-management, presentation and collaboration - that can be applied in all employment sectors will be acquired, particularly through transferable skills training at UCL and Birkbeck. It is important that the results of the work are communicated to the general public. Dr Moores has a proven track-record of public communication of science. She was the 2006 winner of the prestigious DeMontfort medal for science communication (SET for Britain) and she also attended the BBSRC MediaTraining Day. Prof Forge has given interviews about hearing and deafness to BBC radio and is also regularly involved in presenting his work at events organised by hearing research charities, Deafness Research UK and the RNID. Similar means of communication will be developed during the course of this project.