Muallem, D.; (2005) An anion transporter theory of the outer hair cell motor protein. Doctoral thesis , University of London.

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Abstract

This thesis addresses the mechanism of electromotility in outer hair cells (OHCs) of the mammalian cochlea. Prestin, a protein densely packed in the lateral membrane is assumed to drive electromotility. A current hypothesis is that prestin is an incomplete transporter, shuttling chloride across the membrane without allowing it to dissociate at the extracellular surface. In this thesis kinetic models are formulated to show that this hypothesis cannot reproduce the previously published experimental data from electrical recordings. However an alternative model of prestin as an anion exchanger (modelled here as a chloride/sulphate exchanger) is formulated, which can reproduce many of the experimental observations. In this model the experimentally observed charge movements across the cell membrane are produced by the translocation of a chloride ion combined with some intrinsic charged residues. To further test the predictions of the model, patch clamp recordings were performed on dissociated OHCs, in the excised patch and whole-cell configurations. The OHC non-linear capacitance (NLC) depended on the concentration of intracellular chloride (Clj). When Clj was removed from internal and external solutions, a residual NLC (-15- 30%) was found, which was consistent with the predictions of the model for contaminant levels of Clj ( 10uM). Additionally the effect on the NLC of reducing Clj depended on the species of anion used to replace Clj. The largest effect was produced by replacement with sulphate, whilst the smallest effect was produced by replacement with glutamate. These findings support the model. Finally two potential causes for previous controversy in the literature were identified. 1) The NLC depended on the recording configuration when Clj was reduced below 1-1 OmM. 2) The dependence of the NLC on Clj was affected when Tris+ replaced Na+ as the major cation in solutions.

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