Currents In Outer Hair Cells Research Articles

Synaptic Contributions to Cochlear Outer Hair Cell Ca2+ Dynamics.

For normal cochlear function, outer hair cells (OHCs) require a precise control of intracellular Ca2+ levels. In the absence of regulatory elements such as proteinaceous buffers or extrusion pumps, OHCs degenerate, leading to profound hearing impairment. Influx of Ca2+ occurs both at the stereocilia tips and the basolateral membrane. In this latter compartment, two different origins for Ca2+ influx have been poorly explored: voltage-gated L-type Ca2+ channels (VGCCs) at synapses with Type II afferent neurons, and α9α10 cholinergic nicotinic receptors at synapses with medio-olivochlear complex (MOC) neurons. Using functional imaging in mouse OHCs, we dissected Ca2+ influx individually through each of these sources, either by applying step depolarizations to activate VGCC, or stimulating MOC axons. Ca2+ ions originated in MOC synapses, but not by VGCC activation, was confined by Ca2+-ATPases most likely present in nearby synaptic cisterns. Although Ca2+ currents in OHCs are small, VGCC Ca2+ signals were comparable in size to those elicited by α9α10 receptors, and were potentiated by ryanodine receptors (RyRs). In contrast, no evidence of potentiation by RyRs was found for MOC Ca2+ signals over a wide range of presynaptic stimulation strengths. Our study shows that despite the fact that these two Ca2+ entry sites are closely positioned, they differ in their regulation by intracellular cisterns and/or organelles, suggesting the existence of well-tuned mechanisms to separate the two different OHC synaptic functions.SIGNIFICANCE STATEMENT Outer hair cells (OHCs) are sensory cells in the inner ear operating under very special constraints. Acoustic stimulation leads to fast changes both in membrane potential and in the intracellular concentration of metabolites such as Ca2+ Tight mechanisms for Ca2+ control in OHCs have been reported. Interestingly, Ca2+ is crucial for two important synaptic processes: inhibition by efferent cholinergic neurons, and glutamate release onto Type II afferent fibers. In the current study we functionally imaged Ca2+ at these two different synapses, showing close positioning within the basolateral compartment of OHCs. In addition, we show differential regulation of these two Ca2+ sources by synaptic cisterns and/or organelles, which could result crucial for functional segregation during normal hearing.

Cochlear microphonic latency predicts outer hair cell function in animal models and clinical populations

As recently reported, electrocochleography recorded in cochlear implant recipients showed reduced amplitude and shorter latency in patients with more severe high-frequency hearing loss compared with those with some residual hearing. As the response is generated primarily by receptor currents in outer hair cells, these variations in amplitude and latency may indicate outer hair cell function after cochlear implantation. We propose that an absence of latency shift when the cochlear microphonic is measured on two adjacent electrodes indicates an absence or dysfunction of outer hair cells between these electrodes. We test this preclinically in noise deafened guinea pigs (2 h of a 124 dB HL, 16–24 kHz narrow-band noise), and clinically, in electrocochleographic recordings made in cochlear implant recipients immediately after implantation. We found that normal hearing guinea pigs showed a progressive increase in latency from basal to apical electrodes. In contrast, guinea pigs with significantly elevated high-frequency hearing thresholds showed no change in cochlear microphonic latency measured on basal electrodes (located approximately at the 16–24 kHz location in the cochlea).. In the clinical cohort, a significant negative correlation existed between cochlear microphonic latency shifts and hearing thresholds at 1-, 2-, & 4 kHz when tested on electrodes located at the relevant cochlear tonotopic place. This reduction in latency shift was such that patients with no measurable hearing also had no detectable latency shift (place assessed by CT scan, r’s of -.70 to -.83). These findings suggest that electrocochleography can be used as a diagnostic tool to detect cochlear regions with functioning hair cells, which may be important for defining cross-over point for electro-acoustic stimulation.

Using Cochlear Microphonic Potentials to Localize Peripheral Hearing Loss.

The cochlear microphonic (CM) is created primarily by the receptor currents of outer hair cells (OHCs) and may therefore be useful for identifying cochlear regions with impaired OHCs. However, the CM measured across the frequency range with round-window or ear-canal electrodes lacks place-specificity as it is dominated by cellular sources located most proximal to the recording site (e.g., at the cochlear base). To overcome this limitation, we extract the “residual” CM (rCM), defined as the complex difference between the CM measured with and without an additional tone (saturating tone, ST). If the ST saturates receptor currents near the peak of its excitation pattern, then the rCM should reflect the activity of OHCs in that region. To test this idea, we measured round-window CMs in chinchillas in response to low-level probe tones presented alone or with an ST ranging from 1 to 2.6 times the probe frequency. CMs were measured both before and after inducing a local impairment in cochlear function (a 4-kHz notch-type acoustic trauma). Following the acoustic trauma, little change was observed in the probe-alone CM. In contrast, rCMs were reduced in a frequency-specific manner. When shifts in rCM levels were plotted vs. the ST frequency, they matched well the frequency range of shifts in neural thresholds. These results suggest that rCMs originate near the cochlear place tuned to the ST frequency and thus can be used to assess OHC function in that region. Our interpretation of the data is supported by predictions of a simple phenomenological model of CM generation and two-tone interactions. The model indicates that the sensitivity of rCM to acoustic trauma is governed by changes in cochlear response at the ST tonotopic place rather than at the probe place. The model also suggests that a combination of CM and rCM measurements could be used to assess both the site and etiology of sensory hearing loss in clinical applications.

A connexin30 mutation rescues hearing and reveals roles for gap junctions in cochlear amplification and micromechanics

Accelerated age-related hearing loss disrupts high-frequency hearing in inbred CD-1 mice. The p.Ala88Val (A88V) mutation in the gene coding for the gap-junction protein connexin30 (Cx30) protects the cochlear basal turn of adult CD-1Cx30A88V/A88V mice from degeneration and rescues hearing. Here we report that the passive compliance of the cochlear partition and active frequency tuning of the basilar membrane are enhanced in the cochleae of CD-1Cx30A88V/A88V compared to CBA/J mice with sensitive high-frequency hearing, suggesting that gap junctions contribute to passive cochlear mechanics and energy distribution in the active cochlea. Surprisingly, the endocochlear potential that drives mechanoelectrical transduction currents in outer hair cells and hence cochlear amplification is greatly reduced in CD-1Cx30A88V/A88V mice. Yet, the saturating amplitudes of cochlear microphonic potentials in CD-1Cx30A88V/A88V and CBA/J mice are comparable. Although not conclusive, these results are compatible with the proposal that transmembrane potentials, determined mainly by extracellular potentials, drive somatic electromotility of outer hair cells.

Potassium channel activator attenuates salicylate-induced cochlear hearing loss potentially ameliorating tinnitus.

High dose sodium salicylate causes moderate, reversible hearing loss and tinnitus. Salicylate-induced hearing loss is believed to arise from a reduction in the electromotile response of outer hair cells (OHCs) and/or reduction of KCNQ4 potassium currents in OHCs, which decreases the driving force for the transduction current. Therefore, enhancing OHC potassium currents could potentially prevent salicylate-induced temporary hearing loss. In this study, we tested whether opening voltage-gated potassium channels using ICA-105665, a novel small molecule that opens KCNQ2/3 and KCNQ3/5 channels, can reduce salicylate-induced hearing loss. We found that systemic application of ICA-105665 at 10 mg/kg prevented the salicylate-induced amplitude reduction and threshold shift in the compound action potentials recorded at the round window of the cochlea. ICA-105665 also prevented the salicylate-induced reduction of distortion-product otoacoustic emission. These results suggest that ICA-105665 partially compensates for salicylate-induced cochlear hearing loss by enhancing KCNQ2/3 and KCNQ3/5 potassium currents and the motility of OHCs.

Characteristics of K(+) Outward Currents in the Cochlear Outer Hair Cells of Circling Mice within the First Postnatal Week.

K+ outward currents in the outer hair cells (OHCs) of circling mice (homozygous (cir/cir) mice), an animal model for human deafness (DFNB6 type), were investigated using a whole cell patch clamp technique. Littermate heterozygous (+/cir) mice of the same age (postnatal day (P) 0 -P6) were used as controls. Similar slow rising K+ currents were observed in both genotypes, but their biophysical and pharmacological properties were quite different. The values of Vhalf for activation were significantly different in the heterozygous (+/cir) and homozygous (cir/cir) mice (-8.1±2.2 mV, heterozygous (+/cir) mice (n=7) and -17.2±4.2 mV, homozygous (cir/cir) mice (n=5)). The inactivation curve was expressed by a single first order Boltzmann equation in the homozygous (cir/cir) mice, while it was expressed by a sum of two first order Boltzmann equations in the heterozygous (+/cir) mice. The K+ current of homozygous (cir/cir) mice was more sensitive to TEA in the 1 to 10 mM range, while the 4-AP sensitivities were not different between the two genotypes. Removal of external Ca2+ did not affect the K+ currents in either genotype, indicating that the higher sensitivity of K+ current to TEA in the homozygous (cir/cir) mice was not due to an early expression of Ca2+ activated K+ channels. Our results suggest that the K+ outward current of developing homozygous (cir/cir) mice OHCs is different in both biophysical and pharmacological aspects than that of heterozygous (+/cir) mice.

Predicting the location of missing outer hair cells using the electrical signal recorded at the round window.

The electrical signal recorded at the round window was used to estimate the location of missing outer hair cells. The cochlear response was recorded to a low frequency tone embedded in high-pass filtered noise conditions. Cochlear damage was created by either overexposure to frequency-specific tones or laser light. In animals with continuous damage along the partition, the amplitude of the cochlear response increased as the high-pass cutoff frequency increased, eventually reaching a plateau. The cochlear distance at the onset of the plateau correlated with the anatomical onset of outer hair cell loss. A mathematical model replicated the physiologic data but was limited to cases with continuous hair cell loss in the middle and basal turns. The neural contribution to the cochlear response was determined by recording the response before and after application of Ouabain. Application of Ouabain eliminated or reduced auditory neural activity from approximately two turns of the cochlea. The amplitude of the cochlear response was reduced for moderate signal levels with a limited effect at higher levels, indicating that the cochlear response was dominated by outer hair cell currents at high signal levels and neural potentials at low to moderate signal levels.

Short-Term Synaptic Plasticity Regulates the Level of Olivocochlear Inhibition to Auditory Hair Cells

In the mammalian inner ear, the gain control of auditory inputs is exerted by medial olivocochlear (MOC) neurons that innervate cochlear outer hair cells (OHCs). OHCs mechanically amplify the incoming sound waves by virtue of their electromotile properties while the MOC system reduces the gain of auditory inputs by inhibiting OHC function. How this process is orchestrated at the synaptic level remains unknown. In the present study, MOC firing was evoked by electrical stimulation in an isolated mouse cochlear preparation, while OHCs postsynaptic responses were monitored by whole-cell recordings. These recordings confirmed that electrically evoked IPSCs (eIPSCs) are mediated solely by α9α10 nAChRs functionally coupled to calcium-activated SK2 channels. Synaptic release occurred with low probability when MOC-OHC synapses were stimulated at 1 Hz. However, as the stimulation frequency was raised, the reliability of release increased due to presynaptic facilitation. In addition, the relatively slow decay of eIPSCs gave rise to temporal summation at stimulation frequencies >10 Hz. The combined effect of facilitation and summation resulted in a frequency-dependent increase in the average amplitude of inhibitory currents in OHCs. Thus, we have demonstrated that short-term plasticity is responsible for shaping MOC inhibition and, therefore, encodes the transfer function from efferent firing frequency to the gain of the cochlear amplifier.

Pushing the Envelope of Sound

Normal hearing depends on the amplification of sound in the cochlea. In this issue of Neuron, Fettiplace and colleagues show that prestin-based amplification is a viable mechanism as it is not limited by the outer hair cell's membrane time constant as previously surmised.

BK Channels Mediate Cholinergic Inhibition of High Frequency Cochlear Hair Cells

BackgroundOuter hair cells are the specialized sensory cells that empower the mammalian hearing organ, the cochlea, with its remarkable sensitivity and frequency selectivity. Sound-evoked receptor potentials in outer hair cells are shaped by both voltage-gated K+ channels that control the membrane potential and also ligand-gated K+ channels involved in the cholinergic efferent modulation of the membrane potential. The objectives of this study were to investigate the tonotopic contribution of BK channels to voltage- and ligand-gated currents in mature outer hair cells from the rat cochlea.Methodology/PrincipalFindings In this work we used patch clamp electrophysiology and immunofluorescence in tonotopically defined segments of the rat cochlea to determine the contribution of BK channels to voltage- and ligand-gated currents in outer hair cells. Although voltage and ligand-gated currents have been investigated previously in hair cells from the rat cochlea, little is known about their tonotopic distribution or potential contribution to efferent inhibition. We found that apical (low frequency) outer hair cells had no BK channel immunoreactivity and little or no BK current. In marked contrast, basal (high frequency) outer hair cells had abundant BK channel immunoreactivity and BK currents contributed significantly to both voltage-gated and ACh-evoked K+ currents.Conclusions/SignificanceOur findings suggest that basal (high frequency) outer hair cells may employ an alternative mechanism of efferent inhibition mediated by BK channels instead of SK2 channels. Thus, efferent synapses may use different mechanisms of action both developmentally and tonotopically to support high frequency audition. High frequency audition has required various functional specializations of the mammalian cochlea, and as shown in our work, may include the utilization of BK channels at efferent synapses. This mechanism of efferent inhibition may be related to the unique acetylcholine receptors that have evolved in mammalian hair cells compared to those of other vertebrates.

Lead (Pb 2+) modulation of potassium currents of guinea pig outer hair cells

Outer hair cells (OHC) are mechanosensitive sensory cells of the inner ear cochlea and are involved in modulating the activity of inner hair cells in the transduction of an acoustic stimulus. Potassium (K +) currents play an important role in the sensory transduction process. K + currents were recorded from acutely dissociated OHC obtained from the guinea pig organ of Corti. The whole-cell patch clamp technique was employed. We identified a channel that exhibited outward current of the delayed rectifier type (Kv). Kv channels mediating inward currents carried by potassium ions were also identified and took on the appearance of a previously described inwardly rectifying current. Lead (Pb 2+) acetate at concentrations of 0.1, 1.0, 10, and 100 μM was bath applied. Time to activation for outward-going current was not affected by Pb 2+. The time course of Pb 2+ effects was seen as a dose-dependent reduction of K + current over time, with very little or no recovery after washout. Pb 2+ inhibited the outward Kv relative current with values of 0.10, 0.14, 0.18, and 0.30 at 0.1, 1.0, 10, and 100 μM, respectively. Pb 2+ did not modulate time to activation, peak current, or inactivation of inward I K. The effects of Pb 2+ on the potassium currents of OHC are not remarkable and therefore OHC are probably not a major cause of purported peripheral hearing loss observed in Pb 2+-exposed animals and humans.

Cav1.3 (alpha1D) Ca2+ currents in neonatal outer hair cells of mice.

Outer hair cells (OHC) serve as electromechanical amplifiers that guarantee the unique sensitivity and frequency selectivity of the mammalian cochlea. It is unknown whether the afferent fibres connected to adult OHCs are functional. If so, voltage-activated Ca2+ channels would be required for afferent synaptic transmission. In neonatal OHCs, Ca2+ channels seem to play a role in maturation since OHCs from Cav1.3-deficient (Cav1.3-/-) mice degenerate shortly after the onset of hearing. We therefore studied whole-cell Ca2+ currents in outer hair cells aged between postnatal day 1 (P1) and P8. OHCs showed a rapidly activating inward current that was 1.8 times larger with 10 mM Ba2+ as charge carrier (IBa) than with equimolar Ca2+ (ICa). IBa started activating at -50 mV with Vmax = -1.9 +/- 6.9 mV, V0.5 = -15.0 +/- 7.1 mV and k = 8.2 +/- 1.1 mV (n = 34). The peak IBa showed negligible inactivation (3.6 % after 300 ms) whereas the ICa (10 mM Ca2+) was inactivated by 50.7 %. OHC IBa was reduced by 33.5 +/- 10.3 % (n = 14) with 10 microM nifedipine and increased to 178.5 +/- 57.8 % (n = 14) by 5 microM Bay K 8644. A dose-response curve for nifedipine revealed an IC50 of 2.3 microM, a Hill coefficient of 2.7 and a maximum block of 36 %. Average IBa density in OHCs was 24.4 +/- 10.8 pA pF-1 (n = 105) which is only 38 % of the value in inner hair cells. Single cell RT-PCR revealed expression of Cav1.3 in OHCs. In OHCs from Cav1.3-/- mice, Ba2+ current density was reduced to 0.6 +/- 0.5 pA pF-1 (n = 9) indicating that > 97 % of the Ca2+ channel current in OHCs flows through Cav1.3.

Fast adaptation of mechanoelectrical transducer channels in mammalian cochlear hair cells.

Outer hair cells are centrally involved in the amplification and frequency tuning of the mammalian cochlea, but evidence about their transducing properties in animals with fully developed hearing is lacking. Here we describe measurements of mechanoelectrical transducer currents in outer hair cells of rats between postnatal days 5 and 18, before and after the onset of hearing. Deflection of hair bundles using a new rapid piezoelectric stimulator evoked transducer currents with ultra-fast activation and adaptation kinetics. Fast adaptation resembled the same process in turtle hair cells, where it is regulated by changes in stereociliary calcium. It is argued that sub-millisecond transducer adaptation can operate in outer hair cells under the ionic, driving force and temperature conditions that prevail in the intact mammalian cochlea.

Mercury (Hg 2+) suppression of potassium currents of outer hair cells

The heavy metal mercury (Hg 2+) is an insidious environmental pollutant that causes toxic effects on sensory systems. It is well known that the group IIB divalent cation Hg 2+ is an inhibitor of the group I monovalent potassium (K +) cation pore-forming channel in several biological preparations. Here, we used the whole cell patch clamp technique on freshly isolated outer hair cells (OHCs) of the guinea pig cochlea to record outward K + currents and inward K + currents treated with mercuric chloride (HgCl 2). HgCl 2 affected K + currents in a voltage- and dose-dependent manner. The effects of HgCl 2 at 1.0–100 μM are more pronounced on onset peak current than on steady-state end current. HgCl 2 depolarized also the resting membrane potential. Although the effect of HgCl 2 at 1.0 μM was partially washed out over several minutes, the effects at 10 and 100 μM were irreversible to washout. Since K + channels of OHCs are targets for HgCl 2 ototoxicity, this may lead to auditory transduction problems, including a loss in hearing sensitivity. A better understanding of fundamental mechanisms underlying K + channelopathies in OHCs due to HgCl 2 poisoning may lead to better preventive or therapeutic agents.

Inhibition of K+ currents of outer hair cells in guinea pig cochlea by fluoxetine

The effects of fluoxetine (Prozac®), a widely used antidepressant drug, on K+ channel in outer hair cells isolated from guinea pig cochlea were studied using the whole-cell patch clamp technique. Fluoxetine potently inhibited leak K+ currents with an IC50 of 0.78 μM. The inhibition was reversible and voltage-independent. At 45- to 103-fold higher concentrations than the plasma levels, fluoxetine reversibly blocked voltage-activated K+ currents. Kinetics of the current in the presence of fluoxetine resembled the control current, and the inhibition was not use-dependent. Neither the activation curve nor the reversal potential was affected by fluoxetine. This inhibition was voltage-dependent with an electric distance (δ value) of the binding site of at least 26% of the membrane field from the cytoplasmic side. Use-independent inhibition suggests that fluoxetine blocks the channel before its opening or instantly blocks the open channel. This is the first study of the action of this compound on K+ channel of outer hair cells of the mammalian inner ear. We conclude that the block of the leak K+ currents can occur at therapeutic levels of fluoxetine. Since the voltage-activated K+ currents are not potently blocked by fluoxetine, this action might not be related to its antidepressant action or adverse effects.

Tetraethylammonium ions block the nicotinic cholinergic receptors of cochlear outer hair cells

Cochlear outer hair cells (OHCs) express nicotinic acetylcholine receptors (nAChRs) whose calcium permeability allow the activation of co-localized Ca 2+-sensitive K + channels (SK-type). The large organic cation tetraethylammonium (TEA) is known to block at millimolar concentration voltage-gated and Ca 2+-activated K + currents in OHCs. In the present study, we show that extracellular TEA blocked much more efficiently, at micromolar concentrations, ACh-evoked K + currents in isolated guinea pig OHCs. The dose–inhibition curve indicated an IC 50 of 60 μM, a value two orders of magnitude lower than the one reported on SK or BK channels. The site of the blocking action was on the extracellular side of the plasma membrane since 10 mM intracellular TEA did not prevent or change the characteristics of the ACh-evoked K + current. The block of this K + current in OHCs was mainly explained by a direct action of TEA at the nAChRs. Indeed, we demonstrated that extracellular TEA inhibited directly the ionotropic cation current flowing through the nAChRs (IC 50=30 μM). This study demonstrated for the first time that extracellular TEA is an effective blocker of the OHCs’ nAChRs.

Regulation of ionic currents by protein kinase A and intracellular calcium in outer hair cells isolated from the guinea-pig cochlea.

Two prominent potassium currents, termed IK and IK,n, and a cation current are found in outer hair cells (OHCs) of the guinea-pig cochlea. We report here whole-cell recordings which indicate that the currents are regulated by intracellular factors. 8-bromo-cAMP (500 microM), a membrane-permeable cAMP analogue, activated potassium currents in OHCs in both apical and basal turns of the cochlea. In OHCs from the cochlear apex, the drug effect was largest at potentials positive to -40 mV, indicating IK as the target. In short cells from the cochlear base, both IK and IK,n were affected. The effects of 8-bromo-cAMP could be blocked by the presence of 1 microM H-89 (a protein kinase A inhibitor) in the patch pipette solution. Extracellular application of 10 nM okadaic acid, a protein phosphatase inhibitor, also activated both potassium currents. Currents were also modulated by intracellular calcium. IK was activated in long cells by photorelease of calcium from the caged compound nitr5. Cation current activation required calcium release by photolysis of DM-nitrophen, a compound releasing more calcium. The results show that OHC potassium channels are regulated by background phosphorylation through protein kinase A and dephosphorylation by protein phosphatase. Cellular calcium also activates IK and the cation channel, but with different sensitivities.

A potassium current in guinea-pig outer hair cells activated by ion channel blocker DCDPC.

Fenamate compounds have been reported to inhibit ion channels in a number of tissues, including a non-selective cation channel in the mammalian outer hair cell (OHC). We have further investigated the effects of 3'-5-dichlorodiphenylamine-2-carboxylic acid (DCDPC) on OHC currents using the whole-cell configuration of the patch clamp technique. Extracellular application of 10 microM DCDPC rapidly and reversibly activated an inward current at hyperpolarized potentials. The DCDPC-activated current appeared in the shorter OHCs from the basal turns of the cochlea. The reversal potential of the inward current was dependent on the external K+ ion concentration. An outwardly rectifying K+ current, found predominantly in OHCs from apical turns, was reversibly inhibited by DCDPC. These results suggest that DCDPC has a significant effect on OHC physiology at all tonotopic locations along the basilar membrane and so may have implications for cochlear function during fenamate intake.

Expression of potassium channels in gerbil outer hair cells during development does not require neural induction

Mammalian outer hair cells (OHCs) contain Ca and K channels in their synaptic pole. We questioned if the ontogeny of potassium currents of OHCs depends on the neural induction of early afferent contact. By recording whole-cell currents of OHCs grown in organotypic cultures deprived of afferent innervation, we show that a Ca-activated K channel is expressed in these cells, suggesting that the ontogeny of the K channel is an intrinsic process.

Action of salicylate on membrane capacitance of outer hair cells from the guinea‐pig cochlea.

1. The effect of salicylate on membrane capacitance and intracellular pH has been measured in isolated outer hair cells (OHCs) during whole cell recording. Cell membrane capacitance was measured using a lock-in amplifier technique. 2. Salicylate applied in the bath reduced the fast charge movement, equivalent to a voltage-dependent membrane capacitance, present in OHCs. Simultaneous measurement of membrane capacitance and voltage-driven cell length changes showed that salicylate reduced both together. 3. A small effect of salicylate on outward currents at 0 mV was observed. Sodium salicylate (5 mM) reduced the currents by 19% and another weak acid, sodium butyrate (10 mM), reduced outward currents in OHCs by 15%. 4. The ratiometric dye 2,7-bis(2-carboxymethyl)-5,6-carboxyfluorescein (BCECF) was used to measure pHi changes in OHCs during weak acid exposure. Membrane capacitance and pHi were measured simultaneously in OHCs exposed first to 10 mM sodium butyrate and then to 5 mM sodium salicylate. Although both compounds produced a similar reduction in pHi, butyrate decreased the resting capacitance from a mean resting capacitance of 35 pF (at -30 mV) by 5.4 +/- 2.1 pF, whereas salicylate decreased it by 15.7 +/- 2.3 pF (n = 4). 5. Exposure of OHCs to 10 mM sodium benzoate, an amphiphilic anion, reduced resting membrane capacitance at -30 mV by 9.2 +/- 3.2 pF (n = 3). Outward currents, measured at 0 mV, were reduced by 0.25 +/- 0.05 nA during benzoate application, comparable with the effect of salicylate. 6. Capacitance was measured during slow bath application of salicylate. The resulting dose-capacitance curve had a Hill coefficient of 3.40 +/- 0.85 (n = 4) and a half-maximal dose of 3.95 +/- 0.34 mM. The dose-capacitance curve was not significantly voltage dependent. 7. Salicylate had no detectable effect on the resting capacitance of Deiters' cells, a non-sensory cell type of the organ of Corti. 8. It is concluded that many of the described effects of salicylate on hearing may arise from the partitioning of the salicylate molecule into the membrane of the OHC and consequent inhibition of OHC motility.