Membrane prestin expression correlates with the magnitude of prestin-associated charge movement

Abstract

Full expression of electromotility, generation of non-linear capacitance (NLC), and high-acuity mammalian hearing require prestin function in the lateral wall of cochlear outer hair cells (OHCs). Estimates of the number of prestin molecules in the OHC membrane vary, and a consensus has not emerged about the correlation between prestin expression and prestin-associated charge movement in the OHC. Using an inducible prestin-expressing cell line, we demonstrate that the charge density, but not the voltage at peak capacitance, directly correlates with the amount of prestin in the plasma membrane. This correlation is evident in studies involving a controlled increase of prestin expression with time after induction and inducer dose-response. Conversely, membrane prestin levels and charge density gradually decline together following the reduction of prestin levels from a steady state by removal of the inducer. Thus, charge density directly correlates with the level of membrane prestin expression, whereas changing membrane levels of prestin have no effect on the voltage at peak capacitance in this inducible prestin-expressing cell line.

The cochlear outer hair cell speed paradox

“…The analysis is agnostic to the specific molecules responsible for piezoelectricity but places constraints on what is thermodynamically feasible. It is known that OHC electromotility requires expression of the transmembrane protein prestin , a member of the SLC26 family of anion transporters ( 3 , 31 , 32 ). There is strong evidence that Cl − is essential and is electrostatically bound in the central core region of the protein ( 33 – 35 ).…”

“…Power tuning curves in Fig. 3 D and E partially account for the OHC electrical corner frequency by driving the cell with a low-pass-filtered voltage, but no attempt was made to address the influence of MET kinetics ( 39 ), ion channel gating and expression ( 9 , 10 ), prestin expression ( 31 ), hair bundle electromotility ( 40 ), inhomogeneous expression and deformation, or mechanical forces associated with the traveling wave. The present OHC model is minimalistic, and reduces a complex cell with inhomogeneous expression and properties into a single lumped element, yet is sufficient to resolve the OHC speed paradox.…”

Diflunisal inhibits prestin by chloride-dependent mechanism

“…Stable HEKs 293 expressing prestin [ 22 ] were grown at 37°C with 5% CO 2 in DMEM (Cellgro) complemented with 10% Tet-System Approved FBS (Clontech) in the presence of penicillin, streptomycin, G418 and hygromycin (Invitrogen). Doxycycline (Clontech) was added to the growing media at a final concentration of 2 μg/ml to induce the expression of prestin-mGFP.…”

“…Doxycycline (Clontech) was added to the growing media at a final concentration of 2 μg/ml to induce the expression of prestin-mGFP. The stable cell line expressing prestin was constructed as described previously [ 22 ].…”

“…In order to understand the physiological consequences of prestin modulation, we aimed at identifying more direct effectors and inhibitors of prestin activity. Based on the effective inhibition of salicylate, we have investigated the effect of the salicylate-derivative diflunisal (DFL) on mouse OHCs and on HEKs expressing prestin[ 14 , 15 , 22 , 23 ]. DFL was discovered in the 1980’s to have improved lipophilicity, increased anti-inflammatory and analgesic properties over salicylate [ 24 ].…”