Outer hair cells (OHCs) of the mammalian cochlea provide forces within the cochlea partician that alter the mechanics and lead to enhanced auditory sensitivity and frequency selectivity. This occurs by an assembly of ‘motor’ molecules in the basolateral membrane, identified and named as Prestin (Zheng et al. 2000), which cause changes in OHC length by conformational changes in response to acoustic frequencies upon membrane polarisation. The recent identification of this motor protein and its relation to a family (SLC26A) of anion exchange transporters (Oliver et al. 2001) has led us to explore the role of Prestin as a bicarbonate-chloride exchanger. As OHCs are known to uptake sugars, the role of GLUT 5 as a fructose transporter was investigated as it was previously identified in the basolateral membrane by antibody labelling (Geleoc et al. 1999).

Guinea-pigs were killed humanely and hair cells isolated from the cochlea were perfused in a flow chamber with perilymph buffered to pH 7.35 with Hepes. External chloride was reduced by gluconate replacement in the perfusion media and on doing so the cells initially shortened in length by approximately 6 %, reversible upon exposure for less than 100 s, as measured by imaging. The use of 1 mM DIDS and 10 mM salicylate in the external media blocked cell shortening in this situation. Conversely, when the external solution was buffered with 25 mM bicarbonate equilibrated with CO₂, no length change occurred when external chloride was reduced. These data suggest that the chloride-bicarbonate exchanger as described in OHC pH measurements (Ikeda et al. 1991) shares many properties of the OHC motor protein and may indeed be Prestin (SLC26A5). Electrophysiology determined a small chloride current (1-2 nS) which reversed around 0 mV with bilaterally symmetric chloride ions. Assuming an OHC motor density of 10⁷ per cell, the conductance per motor would be near those ascribed to ion permeation through other transporters. We hypothesize that Prestin is a low conductance chloride channel.

OHCs are known to take up fructose (Geleoc et al. 1999). A C-terminal GFP-tagged Prestin construct was expressed in HEK-293 cells. Cells were exposed to isotonic replacement of sugar in the external medium to investigate sugar uptake. Cellular fluorescent profiles were imaged and analysed. The isotonic replacement of external glucose with 30 mM fructose led to a reversible increase in the diameter of spherical cells in Prestin (mean ± S.D., n = 18) transfected cells by 2.9 ± 1.7 %. The cell area increased by 5.9 ± 3.6 %. There was no area increase in cells expressing GFP alone or when extracellular solution as control was applied. The data are consistent with an influx of water following fructose exposure and suggest that HEK-293 cells are able to take up fructose when transfected with Prestin. We hypothesize that although other subunits may co-assemble with these proteins, their α helical structure in the most economic morphology allows neutral substrate transport.

We are grateful to B. Fakler for the C-terminal GFP-tagged Prestin construct and R. Trembath for the C-terminal GFP-tagged Pendrin construct. This work was supported by The Wellcome Trust.