Auditory outer hair cells (OHCs) are characterized by an outwardly rectifying and non-inactivating potassium current termed Ik,n, that activates at unusually hyperpolarised voltages (half activation around -80 mV). Recently, KCNQ4 has been shown to be the molecular correlate of Ik,n in outer hair cells. KCNQ4 is thought to be involved in the cycling of potassium ions from the OHCs into the perilymph. Thus KCNQ4 dysfunction leads to progressive OHC degeneration in a hereditary hearing loss both in man (DFNA2) and in a mouse model, possibly due to potassium over-load in hair cells. Surprisingly, the biophysical properties of Ik,n cannot be reproduced in heterologous systems expressing KCNQ4 suggesting hair cell-specific modulation of KCNQ4 by a still unknown mechanism. Recently, the modulation of heterologously expressed KCNQ4 by oxidative modification has been demonstrated, but the physiological relevance is not known. We thus examined the sensitivity of Ik,n to redox-modification and to a cysteine-modifying reagent in whole cell patch clamp experiments from acutely isolated rat OHCs. Oxidizing conditions (5 mM GSSG) induced an approx. 10 mV shift of voltage-dependent activation to hyperpolarized voltages and prevented a current run-down as observed in control experiments. Interestingly, oxidation did not increase Ik,n currents as previously shown for heterologously expressed KCNQ4. In contrast, reducing conditions (2 mM GSH) induced a shift of approx. 10 mV towards depolarized voltages and decreased Ik,n current amplitudes. Since redox-sensitivity of Ik,n suggested the involvement of cysteine residues in voltage-dependent gating, we additionally tested for effects of a cysteine-alkylating compound (NEM). Whilst NEM dramatically increased KCNQ4 currents expressed in CHO cells as previously shown, Ik,n was completely insensitive to NEM (5 mM). Interestingly, when cysteines were completely reduced using DTT (2 mM) prior to NEM application, NEM induced a moderate shift of voltage-dependent activation of approx -10 mV relative to control experiments.

Taken together, our data suggest a substantial impact of redox-dependent modification on the KCNQ conductance in OHCs. Hence, it seems possible that oxidative stress may affect OHC survival via the modulation of hair cell potassium currents. However, oxidative modification does not provide an explanation for the striking difference between native Ik,n and recombinant KCNQ4.