Richmond, Sarah Jane; (1999) A study of in situ outer hair cells from the adult mammalian cochlea. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

This thesis investigates three characteristics of outer hair cells (OHCs) of the adult mammalian cochlea. The first investigation compared the basolateral membrane K+ channel expression of turn 4 (T4) and turn 1 (T1) OHCs, cells that respond to low and high frequency sound respectively. The second and third studies investigated two further aspects of T4 OHCs, the equivalent concentration of endogenous Ca2+ buffer and the characteristics of the mechanoelectric transduction (MET) current. OHCs in situ were recorded from using the whole-cell patch-clamp technique. For the first study, the kinetics, pharmacology and Ca2+ sensitivity of T4 and T1 OHC K+ channels were investigated. This work demonstrated that T4 OHCs express at least three types of K+ channels, termed Ikss, IkCa and lkT4. In contrast, T1 OHCs express different ion channels that exhibit faster onset kinetics, different pharmacology and an insensitivity to raised intracellular Ca2+ concentrations compared to those channels expressed in T4. These T1 channels have been termed IkT1 and Ik,n. The characteristics of these ion channels are discussed in relation to the particular sound frequency to which the OHC best responds. The equivalent concentration of endogenous Ca2+ buffer in T4 OHCs was investigated by using the time constant of current onset as a tool to compare the effects of various concentrations of BAPTA, introduced into the cell via the patch pipette, with those of the endogenous buffer, assayed using the perforated-patch technique. The concentration of endogenous Ca2+ buffer was found to be equivalent to the Ca2+ binding capacity of 2.1 mM BAPTA. This value converts to a Ca2+ binding ratio of 10,500. These results indicate that OHCs posess an enormous Ca2+ buffering capacity, have a low free [Ca2+]i and a huge pool of bound Ca2+ within their cytosol. Finally, the biophysical properties of the MET current of T4 OHCs were investigated. The few recordings obtained were variable but indicated that MET currents in situ are small (60 pA), limited in their onset kinetics only by the kinetics of the fluid-jet stimulus and run-down over a period of 20 minutes in the whole-cell recording configuration. These currents were found to be physiologically effective in activating the motile response of the OHC.

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