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The basilar membrane in the cochlea can be modeled as an array of fluid coupled segments driven by stapes vibration and by the undamping nonlinear force simulating cochlear amplification. If stimulated with two tones, the model generates additional tones due to nonlinear distortion. These distortion products (DPs) can be transmitted into the ear canal and produce distortion-product otoacoustic emissions (DPOAEs) known to be generated in the healthy ear of various vertebrates. This study presents a solution for DPs in a two-dimensional nonlinear cochlear model with cochlear roughness—small irregularities in the impedance along the basilar membrane, which may produce additional DPs due to coherent reflection. The solution allows for decomposition of various sources of DPs in the model. In addition to the already described nonlinear-distortion and coherent-reflection mechanisms of DP generation, this study identifies a long-latency DPOAE component due to perturbation of nonlinear force. DP wavelets that are coherently reflected due to impedance irregularities travel toward the stapes across the primary generation region of DPs and there evoke perturbation of the nonlinear undamping force. The ensuing DP wavelets have opposite phase to the wavelets arising from coherent reflection, which results in partial cancellation of the coherent-reflection DP wavelets.

The plasma-membrane marker FM1-43 was employed to reveal the relative significance of different types of endocytic and transcytic mechanisms in outer hair cells (OHCs) of the guinea-pig cochlea. A double-barrel local perfusion system was used to label independently the apical or synaptic pole of the isolated OHC, to study mechanisms of vesicle uptake at the poles and of vesicle trafficking along and across the cell. Treatment with an inhibitor of macropino- and phagocytosis, phenylarsine oxide, or of clathrin-mediated endocytic activity, concanavalin A, significantly reduced the dye uptake at both the apical and the synaptic poles, indicating the presence of both clathrin-independent and clathrin-mediated processes at both poles. However, measurement of uptake speed in the presence of the inhibitors suggested that clathrin-independent processes contribute more extensively to endocytosis at the basal pole than the apical pole. Treatment with an inhibitor of myosin VI, 2,4,6-triiodophenol, significantly delayed both the apicobasal and the basoapical fluorescence signals. However, treatment with an inhibitor of kinesin, monastrol, or of dynein, ciliobrevin D, significantly delayed the signals only in the basoapical direction. The myosin-VI inhibitor, but neither the kinesin nor dynein inhibitors, significantly delayed the signals to the subsurface cisternae. That is, myosin VI carries vesicles in both longitudinal directions as well as radially to the sub-surface cisternae, whereas kinesin and dynein participate primarily in basoapical trafficking. This fundamental information is essential for elucidating recycling mechanisms of specific proteins involved in establishing, controlling, and maintaining the electromechanical action of OHCs and, therefore, vital for understanding auditory perception.

Background Intense endocytic activity at the apex of outer hair cells (OHCs)—the electromechanical cells of the cochlea—has been demonstrated using the vital plasma-membrane marker FM1-43 and confocal laser-scanning microscopy. Vesicular traffic toward the cell nucleus to distinct locations of the endoplasmic reticulum has also been shown. Objective The current study characterizes the dynamics of endocytic activity, as well as apicobasal and basoapical trafficking, using a local perfusion technique that we recently developed and published to visualize bidirectional trafficking in isolated bipolar cells. Materials and methods The fluorescent plasma-membrane markers FM1-43 (10 µM) and FM4-64 (10 µM), together with a fluid-phase marker, Lucifer yellow (50 µM), were used to label endocytosed vesicles in isolated OHCs of the guinea pig cochlea. Targets of endocytosed vesicles were examined with a fluorescent marker of subsurface cisternae, DiOC6 (0.87 µM). Single- and two-photon confocal laser-scanning microscopy was used to visualize labeled vesicles. Results The plasma-membrane markers presented more intense vesicle internalization at the synaptic pole than at the apical pole of the OHC. Intracellular basoapical vesicle trafficking was faster than apicobasal trafficking. Vesicles endocytosed at the synaptic pole were transcytosed to the endoplasmic reticulum system. An intracellular Lucifer yellow signal was not detected. Conclusion The larger endocytic fluorescent signals in the synaptic pole and the faster basoapical trafficking imply that membrane internalization and vesicle trafficking are more efficient at the synaptic pole than at the apical pole of the OHC.

Sensorineural deafness is caused by the loss of peripheral neural input to the auditory nerve, which may result from peripheral neural degeneration and/or a loss of inner hair cells. Provided spiral ganglion cells and their central processes are patent, cochlear implants can be used to electrically stimulate the auditory nerve to facilitate hearing in the deaf or severely hard-of-hearing. Neural degeneration is a crucial impediment to the functional success of a cochlear implant. The present, first-of-its-kind two-dimensional finite-element model investigates how the depletion of neural tissues might alter the electrically induced transmembrane potential of spiral ganglion neurons. The study suggests that even as little as 10% of neural tissue degeneration could lead to a disproportionate change in the stimulation profile of the auditory nerve. This result implies that apart from encapsulation layer formation around the cochlear implant electrode, tissue degeneration could also be an essential reason for the apparent inconsistencies in the functionality of cochlear implants.

Stimulus-frequency otoacoustic emissions (SFOAEs) are generated by coherent reflection of forward traveling waves by perturbations along the basilar membrane. The strongest wavelets are backscattered near the place where the traveling wave reaches its maximal amplitude (tonotopic place). Therefore, the SFOAE group delay might be expected to be twice the group delay estimated in the cochlear filters. However, experimental data have yielded steady-state SFOAE components with near-zero latency. A cochlear model is used to show that short-latency SFOAE components can be generated due to nonlinear reflection of the compressor or suppressor tones used in SFOAE measurements. The simulations indicate that suppressors produce more pronounced short-latency components than compressors. The existence of nonlinear reflection components due to suppressors can also explain why SFOAEs can still be detected when suppressors are presented more than half an octave above the probe-tone frequency. Simulations of the SFOAE suppression tuning curves showed that phase changes in the SFOAE residual as the suppressor frequency increases are mostly determined by phase changes of the nonlinear reflection component.

Distortion-product otoacoustic emissions (DPOAEs) emerge from the cochlea when elicited with two tones of frequencies f1 and f2. DPOAEs mainly consist of two components, a nonlinear-distortion and a coherent-reflection component. Input-output (I/O) functions of DPOAE pressure at the cubic difference frequency, f DP = 2 f 1 − f 2, enable the computation of estimated distortion-product thresholds (EDPTs), offering a noninvasive approach to estimate auditory thresholds. However, wave interference between the DPOAE components and suboptimal stimulus-level pairs reduces the accuracy of EDPTs. Here, the amplitude P of the nonlinear-distortion component is extracted from short-pulse DPOAE time signals. DPOAE level maps representing the growth behavior of P in L 1 , L 2 space are recorded for 21 stimulus-level pairs and 14 frequencies with f 2 = 1 to 14 kHz ( f 2 / f 1 = 1.2) from 20 ears. Reproducing DPOAE growth behavior using a least-squares fit approach enables the derivation of ridge-based I/O functions from model level maps. Objective evaluation criteria assess the fit results and provide EDPTs, which correlate significantly with auditory thresholds (p

Background Fluorescence membrane markers are efficient tools for visualizing the dynamics of membrane recycling processes in living cells. The outer hair cell (OHC) − a bipolar epithelial cell in the cochlea − possesses endocytic activity at both its apical and basal poles. The best visual overview of transcytosis in the OHC is achieved when the cell is isolated, so that both the apical and the basal poles are in the same focal plane to allow confocal imaging. Until now, fluorescent markers were applied to the extracellular environment of isolated OHCs without distinguishing the apical and basal poles. The drawback of that configuration is that apicobasal and basoapical vesicle traffic labelled at the opposite poles cannot be visualized independently because the same fluorescent marker has access to both poles. New method A double-barrel, capillary perfusion system was developed to independently stain either one pole or both the apical and the basal poles of isolated OHCs using different types of fluorescence membrane markers. Results Producing laminar fluid flow, the double-barrel perfusor allows investigation of the dynamics of apicobasal and basoapical vesicle traffic independently and/or simultaneously in the same OHC. Comparison with existing method This method offers a unique option for investigating bidirectional vesicle traffic in bipolar epithelial cells, which is superior to other already established labelling techniques. Conclusions The double-barrel perfusion system, suitable for selectively staining a longitudinal section of the plasma membrane of an isolated bipolar epithelial cell, opens new possibilities for investigating cell labelling and intracellular vesicle traffic.

As a by-product of nonlinear amplification in the cochlea, the inner ear emits sound waves in response to two tones with different frequencies. These sound waves are measurable in the ear canal as distortion-product otoacoustic emissions (DPOAEs). DPOAEs putatively consist of two components emerging at different locations in the cochlea. Wave interference between the two components limits the accuracy of DPOAEs as a noninvasive measure of cochlear function. Using short stimulus pulses instead of continuous stimuli, the two DPOAE components can be separated in the time domain due to their different latencies. The present work utilizes a nonlinear hydrodynamic cochlea model to simulate short-pulse DPOAEs in the time domain. When adding irregularities to the mechanical parameters of the model, the simulated DPOAE signals show two distinguishable components and long-lasting beat tones, similar to band-pass filtered experimental data from normal-hearing human subjects. The model results suggest that the beat tones can occur solely due to interference of the coherent-reflection component with the fading nonlinear-distortion component.

Distortion-product otoacoustic emissions (DPOAEs) arise in the cochlea in response to two tones with frequencies f1 and f2 and mainly consist of two components, a nonlinear-distortion and a coherent-reflection component. Wave interference between these components limits the accuracy of DPOAEs when evaluating the function of the cochlea with conventional continuous stimulus tones. Here, DPOAE components are separated in the time domain from DPOAE signals elicited with short stimulus pulses. The extracted nonlinear-distortion components are used to derive estimated distortion-product thresholds (EDPTs) from semi-logarithmic input-output (I/O) functions for 20 normal-hearing and 21 hearing-impaired subjects. I/O functions were measured with frequency-specific stimulus levels at eight frequencies f2 = 1,…, 8 kHz (f2/f1 = 1.2). For comparison, DPOAEs were also elicited with continuous primary tones. Both acquisition paradigms yielded EDPTs, which significantly correlated with behavioral thresholds (p

The amplitudes of distortion-product otoacoustic emissions (DPOAEs) may abruptly decrease even though the stimulus level is relatively high. These notches observed in the DPOAE input/output functions or distortion-product grams have been hypothesized to be due to destructive interference between wavelets generated by distributed sources of the nonlinear-distortion component of DPOAEs. In this paper, simulations with a smooth cochlear model and its analytical solution support the hypothesis that destructive interference between individual wavelets may lead to the amplitude notches and explain the cause for onset and offset amplitude overshoots in the DPOAE signal measured for intensity pairs in the notches.

Cochlear implantation leads to many structural changes within the cochlea which can impair residual hearing. In patients with preserved low-frequency hearing, a delayed hearing loss can occur weeks-to-years post-implantation. We explore whether stiffening of the basilar membrane (BM) may be a contributory factor in an animal model. Our objective is to map changes in morphology and Young's modulus of basal and apical areas of the BM after cochlear implantation, using quantitative nanomechanical atomic force microscopy (QNM-AFM) after cochlear implant surgery. Cochlear implantation was undertaken in the guinea pig, and the BM was harvested at four time-points: 1 day, 14 days, 28 days and 84 days post-implantation for QNM-AFM analysis. Auditory brainstem response thresholds were determined prior to implantation and termination. BM tissue showed altered morphology and a progressive increase in Young's modulus, mainly in the apex, over time after implantation. BM tissue from the cochlear base demonstrated areas of extreme stiffness which are likely due to micro-calcification on the BM. In conclusion, stiffening of the BM after cochlear implantation occurs over time, even at sites far apical to a cochlear implant.

Distortion product otoacoustic emissions (DPOAEs) are evoked by two stimulus tones with frequency f 1 and f 2 of ratio f 2 / f 1 in the range between approximately 1.05 and 1.4. This study theoretically and experimentally analyzes the cubic 2 f 1 − f 2 DPOAE for different stimulus levels of one of the tones while the other is constant. Simulations for f 2 / f 1 of 1.2 and moderate stimulus levels (30–70 dB sound pressure level) indicate that cubic distortion products are generated along a relatively large length of the basilar membrane, the extent of which increases with stimulus level. However, apical from the place of maximum nonlinear force, the wavelets generated by these distributed sources mutually cancel. Therefore, although the spatial extent of the primary DPOAE sources broadens with increasing stimulus level (up to 1.5 oct), the basilar-membrane region contributing to the DPOAE signal is relatively narrow (0.6 oct) and level independent. The observed dependence of DPOAE amplitude on stimulus level can be well-approximated by a point source at the basilar-membrane place where the largest distortion product (maximum of the nonlinear force) is generated. Onset and offset of the DPOAE signal may contain amplitude overshoots (complexities), which are in most cases asymmetrical. Two-tone suppression was identified as the main cause of these onset and offset complexities. DPOAE measurements in two normal-hearing subjects support the level dependence of the steady-state DPOAE amplitude and the asymmetry in the onset and offset responses predicted by the theoretical analysis.

When referred to baseline measures, serial monitoring of pure-tone behavioral thresholds and distortion-product otoacoustic emissions (DPOAEs) can be used to detect the progression of cochlear damage. Semi-logarithmic DPOAE input-output (I/O) functions enable the computation of estimated distortion-product thresholds (EDPTs) by means of linear regression, a metric that provides a quantitative estimate of hearing loss due to cochlear-amplifier degradation. DPOAE wave interference and a suboptimal choice of stimulus levels limit the accuracy of EDPTs. This work identifies the test-retest reliability of EDPTs derived from short-pulse DPOAE level maps (EDPTLM), a method that circumvents limitations associated with both wave interference and suboptimal choice of stimulus levels. The test-retest reliability was compared to that of EDPTs derived from semi-logarithmic I/O functions (EDPTI/O) and that of behavioral thresholds acquired with pure-tone audiometry (PTA) and modified Bekesy tracking audiometry (TA) to provide a foundation for identifying and interpreting significant threshold shifts. The DPOAE-based auditory thresholds (EDPTLM and EDPTI/O) and behavioral thresholds (PTA and TA) were recorded seven times within three months at 14 frequencies with f2 = 1-14 kHz in 20 ears from ten subjects with normal hearing (4PTA0.5-4kHz < 20 dB HL). To obtain EDPTLM, short-pulse DPOAEs were recorded using 21 L1,L2 pairs. Reconstruction of DPOAE growth behavior as a function of L1 and L2 using nonlinear curve fitting enabled the derivation of EDPTLM for each frequency. Test-retest reliability was determined using three different approaches: 1) centered thresholds, 2) average threshold differences, and 3) average absolute threshold differences, between each possible test session (N = 21). Test-retest reliability based on centered thresholds and average threshold differences showed no statistically significant difference between EDPTLM, EDPTI/O, PTA, and TA for the pooled analysis incorporating all stimulus frequencies. Average absolute threshold differences presented small but significant differences in test-retest reliability with median values of 3.00 dB for PTA, 3.20 dB for TA, 3.34 dB for EDPTLM, and 3.51 dB for EDPTI/O. A considerable frequency dependence of test-retest reliability was found; namely, the highest test-retest reliability was for EDPTLM at f2 = 11 - 14 kHz. Otherwise, at lower frequencies, the highest test-retest reliability was for TA at f2 =1 - 2 kHz. Overall, the test-retest reliability of EDPTLM was better than that of EDPTI/O and was similar to that for behavioral thresholds. Hence, deriving EDPTLM from individual level maps is a promising and sensitive method for objectively monitoring the state of the cochlea. Furthermore, the detection of an equidirectional threshold change at a single frequency in both EDPTLM and TA might allow reducing the threshold shift as indication of a follow-up examination from the clinical standard of 10 dB down to 5 dB. This stricter indicator might be beneficial when monitoring cochlear damage, for example ototoxicity, in the presence of (remnant) cochlear amplification at baseline.

Distortion-product otoacoustic emissions (DPOAEs) arise as byproduct of nonlinear amplification in the cochlea in response to two stimulus tones of different frequencies. They are assumed to comprise mainly two components, a nonlinear-distortion component and a coherent-reflection component, which may cause wave interference in recordings with conventional, continuous stimulus tones, hampering the accuracy of DPOAEs when evaluating cochlear function. Here, DPOAEs are recorded with short stimulus pulses enabling the separation of the two source components in the time domain. Comparison with conventional stimulation endorses the importance of reliable source separation when using DPOAEs in clinical audiometry or research.

Aim: To obtain direct evidence for the cochlear travelling wave in humans by performing electrocochleography from within the cochlea in subjects implanted with an auditory prosthesis. Background: Sound induces a travelling wave that propagates along the basilar membrane, exhibiting cochleotopic tuning with a frequency-dependent phase delay. To date, evoked potentials and psychophysical experiments have supported the presence of the travelling wave in humans, but direct measurements have not been made. Methods: Electrical potentials in response to rarefaction and condensation acoustic tone bursts were recorded from multiple sites along the human cochlea, directly from a cochlear implant electrode during, and immediately after, its insertion. These recordings were made from individuals with residual hearing. Results: Electrocochleography was recorded from 11 intracochlear electrodes in 7 ears from 6 subjects, with detectable responses on all electrodes in 5 ears. Cochleotopic tuning and frequency-dependent phase delay of the cochlear microphonic were demonstrated. The response latencies were slightly shorter than those anticipated which we attribute to the subjects' hearing loss. Conclusions: Direct evidence for the travelling wave was observed. Electrocochleography from cochlear implant electrodes provides site-specific information on hair cell and neural function of the cochlea with potential diagnostic value.

Distorsionsprodukte otoakustischer Emissionen (DPOAE) und transient evozierte otoakustische Emissionen (TEOAE) sind im Gehorgang messbare Schallwellen, die als Nebenerscheinung der cochlearen Verstarkung entstehen und ein objektives Verfahren zur Diagnostik von Innenohrfunktionsstorungen darstellen. Konventionelle DPOAE- und TEOAE-Verfahren ermoglichen die Detektion eines Horverlusts jedoch nur mit unbefriedigender Genauigkeit. Durch Berucksichtigung des Entstehungsmechanismus der DPOAE soll deren Genauigkeit in der Innenohrdiagnostik gesteigert werden. DPOAE bestehen aus zwei an unterschiedlichen Orten der Cochlea entstehenden Komponenten, welche Artefakte aufgrund von Interferenz erzeugen konnen und mithilfe von Kurzpulsstimulation im Zeitbereich getrennt werden. Optimale Anregungspegel werden zur Akquise von DPOAE mit moglichst maximaler Amplitude verwendet. DPOAE-Daten und Bekesy-Audiogramme wurden von 41 Probanden in einem klinisch relevanten Frequenzbereich von 1,5 bis 6 kHz erfasst. Kurzpuls-Primartone ermoglichen eine artefaktfreie Erfassung von DPOAE. Die mittels semilogarithmischen Wachstumsfunktionen geschatzten Distorsionsproduktschwellen korrelieren signifikant mit den subjektiv erfassten Horschwellen und ermoglichen die Detektion eines Horverlusts von ≥20 dB HL mit 95 % Sensitivitat bei nur 5 % falsch-positiven Diagnosen. Diese Genauigkeit wurde mit einer Messzeit von etwa 1–2 min pro Frequenz erreicht. Durch die Komponententrennung der Kurzpuls-DPOAE und Anwendung von optimalen Anregungsparametern kann die Genauigkeit in der Diagnostik von Schadigungen des cochlearen Verstarkers uber die von konventionellen DPOAE- und TEOAE-Verfahren hinaus gesteigert werden.

The basis of the extraordinary sensitivity and frequency selectivity of the cochlea is a chloride‐sensitive protein called prestin which can produce an electromechanical response and which resides in the basolateral plasma membrane of outer hair cells (OHCs). The compound 9‐anthracenecarboxylic acid (9‐AC), an inhibitor of chloride channels, has been found to reduce the electromechanical response of the cochlea and the OHC mechanical impedance. To elucidate these 9‐AC effects, the functional electromechanical status of prestin was assayed by measuring the nonlinear capacitance of OHCs from the guinea‐pig cochlea and of prestin‐transfected human embryonic kidney 293 (HEK 293) cells. Extracellular application of 9‐AC caused reversible, dose‐dependent and chloride‐sensitive reduction in OHC nonlinear charge transfer, Q max. Prestin‐transfected cells also showed reversible reduction in Q max. For OHCs, intracellular 9‐AC application as well as reduced intracellular pH had no detectable effect on the reduction in Q max by extracellularly applied 9‐AC. In the prestin‐transfected cells, cytosolic application of 9‐AC approximately halved the blocking efficacy of extracellularly applied 9‐AC. OHC inside‐out patches presented the whole‐cell blocking characteristics. Disruption of the cytoskeleton by preventing actin polymerization with latrunculin A or by decoupling of spectrin from actin with diamide did not affect the 9‐AC‐evoked reduction in Q max. We conclude that 9‐AC acts on the electromechanical transducer principally by interaction with prestin rather than acting via the cytoskeleton, chloride channels or pH. The 9‐AC block presents characteristics in common with salicylate, but is almost an order of magnitude faster. 9‐AC provides a new tool for elucidating the molecular dynamics of prestin function.

Vetesnik and Gummer [J. Acoust. Soc. Am. 131, 3914–3934 (2012)] obtained a stationary solution for distortion products (DPs) in a two-dimensionalcochlear model in which the impedance of the basilar membrane varied smoothly along its length. Their solution was used to study the dependence of the nonlinear-distortion source of distortion-product otoacoustic emission (DPOAE) on stimulus level [Vencovský et al., J. Acoust. Soc. Am. 145, 2909–2931 (2019); Vencovský et al., J. Acoust. Soc. Am. 146, EL92–EL98 (2019)]. Here, the solution is extended to “impedance irregularities” in the mechanics of the cochlea causing the generation of the coherent-reflection source of DPOAEs near the DP characteristic place. The coherent-reflection source of DPOAEs can be relatively pronounced in normal-hearing humans and the interference between the nonlinear-distortion and coherent-reflection components complicates the use of DPOAEs in routine clinical screening of the auditory periphery. The solution for the model with mechanical perturbation can be used to study the relationship between nonlinear-distortion and coherent-reflection components of DPOAEs.

Distortion-product otoacoustic emissions (DPOAEs) emerge when presenting two primary tones with different frequencies f1 and f2 to the cochlea and are commonly used in diagnosis and research to evaluate the functional state of the cochlea. Optimal primary-tone stimulus levels accounting for the different level dependencies of the traveling-wave amplitudes of the two primary tones near the f2-tonotopic place on the basilar membrane are often used to maximize DPOAE amplitudes. However, parameters defining the optimal levels can be affected by wave interference between the nonlinear-distortion and coherent-reflection components of the DPOAE. Here, the components were separated in the time domain using a pulsed stimulus paradigm and optimal levels determined. Based on the amplitude dependence of the nonlinear-distortion components on primary-tone stimulus levels, level parameters yielding maximum DPOAE amplitudes were derived for six normal-hearing adults and compared to data recorded with continuous two-tone stimulation. The level parameters resulting from analysis of the nonlinear-distortion components show dependence on stimulus frequency and small standard deviations. DPOAE input/output functions derived for optimal levels exhibit larger slopes, wider dynamic range and less variability across subjects than those derived for conventional stimulus and analysis conditions, potentially increasing their reliability and sensitivity for assessing cochlea function.

Distortion-product otoacoustic emissions (DPOAEs) are presumed to consist mainly of two components, a nonlinear-distortion component and a coherent-reflection component. Wave interference between these two components reduces the accuracy of DPOAEs when used to evaluate cochlear function. Here, short tone pulses are utilized to record DPOAE signals in normal-hearing subjects. DPOAE components are extracted from recordings at discrete frequencies using two different techniques in the time domain. The extracted DPOAE components are compared to recordings obtained with conventional, continuous primary tones.

Distortion product otoacoustic emissions (DPOAEs) acquired in normal-hearing subjects show considerable variation in amplitude with varying frequency. This is known as DPOAE fine structure. It is widely accepted that fine structure results from wave interference from two DPOAE sources, a non-linear generation component and a coherent reflection component. Here a method is presented that decomposes short-pulse DPOAE recordings into pulse basis functions and enables the quantification of both source components in the time domain, independent of their relative phase and at low cost of measurement time. Input-output functions utilizing the extracted primary-source component are analyzed.

A concept for a partially implantable hearing device, for which the power supply and signal transmission are provided by an optical transmission path, is evaluated. The actuator is designed to fit into the round-window niche and to couple directly to the round-window membrane. Implantable hearing aids can be a suitable solution in the case of severe hearing loss, where conventional hearing aids often fail. However, the surgical effort for an implantation is comparatively high. Therefore, the objective of our work was to provide a hearing system which combines reliable coupling to the auditory system with an easy implantation technique. The actuator was designed as a piezoelectric thin-film cantilever. The optical transmission path was realised using an infrared light-emitting diode combined with an active receiver circuit. For a voltage of 1 V, the unloaded actuator presents displacement amplitudes of 1 μm up to a stimulus frequency of 25 kHz and forces up to 0.2 mN. Proportionally larger forces can be achieved by stacking single actuators. The overall transmission loss from the electrical input of the light-emitting diode driver to the mechanical output of the unloaded actuator was less than 25 dB at 1 kHz and maximum output.

The motor protein, prestin, situated in the basolateral plasma membrane of cochlear outer hair cells (OHCs), underlies the generation of somatic, voltage-driven mechanical force, the basis for the exquisite sensitivity, frequency selectivity and dynamic range of mammalian hearing. The molecular and structural basis of the ontogenetic development of this electromechanical force has remained elusive. The present study demonstrates that this force is significantly reduced when the immature subcellular distribution of prestin found along the entire plasma membrane persists into maturity, as has been described in previous studies under hypothyroidism. This observation suggests that cochlear amplification is critically dependent on the surface expression and distribution of prestin. Searching for proteins involved in organizing the subcellular localization of prestin to the basolateral plasma membrane, we identified cochlear expression of a novel truncated prestin splice isoform named prestin 9b (Slc26A5d) that contains a putative PDZ domain-binding motif. Using prestin 9b as the bait in a yeast two-hybrid assay, we identified a calcium/calmodulin-dependent serine protein kinase (CASK) as an interaction partner of prestin. Co-immunoprecipitation assays showed that CASK and prestin 9b can interact with full-length prestin. CASK was co-localized with prestin in a membrane domain where prestin-expressing OHC membrane abuts prestin-free OHC membrane, but was absent from this area for thyroid hormone deficiency. These findings suggest that CASK and the truncated prestin splice isoform contribute to confinement of prestin to the basolateral region of the plasma membrane. By means of such an interaction, the basal junction region between the OHC and its Deiter's cell may contribute to efficient generation of somatic electromechanical force.

Fine structure in the frequency response of distortion product otoacoustic emissions (DPOAEs) can severely limit the usefulness of DPOAEs in estimating auditory thresholds. Here, fine structure is removed by extracting the primary-source DPOAE component using the onset-decomposition technique (Vetešník et al., 2009) and auditory threshold estimates are compared to those obtained from DPOAEs in response to conventional, continuous two-tone stimulation. Auditory thresholds are predicted using the estimated distortion product thresholds (EDPTs), obtained from linear regression of input-output (I/O) functions of DPOAE pressure amplitude versus second-tone stimulus level (Boege and Janssen, 2002). The accuracy of the auditory-threshold predictions is derived by comparison with measured auditory thresholds. The parameters of the two primary stimulus tones of frequency f(1) and f(2) and levels of L(1) and L(2) are chosen as: f(2)/f(1) = 1.2 with 1.5 ≤ f(2) ≤ 2.5 kHz, and L(1) = 0.4L(2) + 39 dB SPL, with 25 ≤ L(2) ≤ 65 dB SPL. Data are from 12 normal-hearing subjects with profound DPOAE fine structure. 255 DPOAE I/O functions were measured for each of the two DPOAE paradigms. An EDPT value was accepted as reliable if: 1) the squared correlation coefficient, r(2) ≥ 0.8, 2) the regression slope, s(I/O) ≥ 0.2 μPa/dB, and 3) the standard deviation of the EDPT, σ(EDPT) ≤ 10 dB. The proportion of rejected I/O functions was 8% for onset-decomposition DPOAEs, and 25% for continuous-tone DPOAEs. Removal of data points from the saturation region of the DPOAE I/O function by an automated algorithm reduced the rejection rate, to zero for onset-decomposition DPOAEs, but to only 13% for continuous-tone DPOAEs. In the absence of saturated DPOAE responses, auditory thresholds were predicted with standard deviation of only 4 dB for onset-decomposition DPOAEs, but 12 dB for continuous-tone DPOAEs. In summary, by extracting the primary-source component of the DPOAE by the method of onset-decomposition it is possible to predict human auditory threshold with hitherto unattainable accuracy.

The basis of the extraordinary sensitivity and frequency selectivity of the mammalian cochlea is an electromechanical feedback system that amplifies the deflection of the hair cell stereocilia. Electromechanical force is generated by the soma of the outer hair cell and acts against viscous forces. These forces are expected to be particularly large in the narrow subtectorial space between the reticular lamina and the overlying tectorial membrane. Fundamental aspects of this amplifying process are not fully understood. Three main questions are addressed in this chapter. First, given capacitive and inertial characteristics of cellular and acellular cochlear components, how is the electromechanical force coupled to the stereocilia locally, radially, and longitudinally, with correct phase to produce gain rather than attenuation? Second, how is temporal fidelity achieved in the presence of high gain? Third, what is the evidence for power amplification rather than just amplitude amplification? This chapter presents modern experimental approaches that are addressing these issues. Presented in a conceptual framework of experiment and theory, three approaches are highlighted: (1) combined pressure and voltage measurements near the organ of Corti, (2) vibration measurements at the reticular lamina and tectorial membrane, and (3) mechanical and electrokinetic properties of the tectorial membrane. The experimental evidence supports the thesis that average power gain in the region of maximum amplitude response and coupling through the tectorial membrane are both crucial for shaping the frequency response of the cochlear amplifier and, therefore, of stereocilia deflection.

The exquisite spectral and temporal properties of the cochlea are achieved by functional integration of specialized subsystems. Being highly integrated, these components had remained technically inaccessible in vivo for fundamental research and clinical diagnosis without damaging the whole system. In the last three decades, however, great steps forward in our understanding of cochlear function have been made possible by technical developments in biophysics, molecular biology, genetics, and imaging. The following chapters provide comprehensive descriptions of the fundamental principles and experimental results associated with various subsystems and their integration. This chapter highlights the major concepts and findings described in each chapter and links them across chapters, demonstrating the major advances in cochlear research.

The active middle-ear implant Vibrant Soundbridge© (VSB) is used to treat mild-to-severe sensorineural hearing losses. The standard surgical approach for incus vibroplasty is a mastoidectomy and a posterior tympanotomy, crimping the Floating Mass Transducer (FMT) to the long process of the incus (LPI) (standard crimped application). However, tight crimping increases the risk of necrosis of the LPI, resulting in reduction of energy transfer and loss of amplification. The aim of this study was to develop a new coupling device for the LPI, that does not require crimping, and to test its vibrational transfer properties in temporal-bone preparations. An extended antrotomy and a posterior tympanotomy were performed in ten fresh human temporal bones. As a control for normal middle-ear function, the tympanic membrane was stimulated acoustically and the vibration of the stapes footplate was measured by laser Doppler vibrometry (LDV). FMT-induced vibration responses of the stapes were then measured for the standard crimped application at the LPI and for the newly designed elastic long process coupler (LP coupler). For the LP coupler, velocity-amplitude responses in temporal-bone preparations showed increased mean amplitudes at around 1 kHz (∼10 dB) and a reduction between 1.8 and 6 kHz (13 dB on average for 2 ≤ f ≤ 5 kHz). In conclusion, attachment of the FMT to the LPI with the LP coupler leads to generally good mechanical and functional coupling in temporal-bone preparations with a notable disadvantage between 1.8 and 6 kHz. Due to its elastic clip attachment it is expected that the LP coupler will reduce the risk of necrosis of the incus long process, which has to been shown in further studies. Clinical results of the LP coupler are pending.

Coupling of somatic electromechanical force from the outer hair cells (OHCs) into the organ of Corti is investigated by measuring transverse vibration patterns of the organ of Cori and tectorial membrane (TM) in response to intracochlear electrical stimulation. Measurement places at the organ of Corti extend from the inner sulcus cells to Hensen’s cells and at the lower (and upper) surface of the TM from the inner sulcus to the OHC region. These locations are in the neighborhood of where electromechanical force is coupled into (1) the mechanoelectrical transducers of the stereocilia and (2) fluids of the organ of Corti. Experiments are conducted in the first, second, and third cochlear turns of an in vitro preparation of the adult guinea pig cochlea. Vibration measurements are made at functionally relevant stimulus frequencies (0.48–68 kHz) and response amplitudes (

Neuere Veroffentlichungen zeigen, dass Messungen der Distorsionsprodukte otoakustischer Emissionen (DPOAE) treffsicherere Diagnosen liefern, wenn erstens die DPOAE-Feinstruktur unterdruckt wird und zweitens die akustische Kalibration verbessert wird. Die Reduzierung der Feinstruktur wirkt sich v. a. im Frequenzbereich bis 4 kHz bei Personen mit intaktem cochlearen Verstarker aus und kann die Standardabweichung einer auf DPOAE-Wachstumsfunktionen basierenden Schwellenschatzung von ublicherweise 11 auf 6 dB reduzieren. Eine verbesserte Kalibration des Schallfelds wirkt sich v. a. bei Frequenzen oberhalb 4 kHz aus. Auf laserinterferometrischen Messungen von DPOAE-Wachstumsfunktionen basierende Schwellenschatzungen mit Schallkalibration nahe dem Trommelfell lieferten eine Standardabweichung von 8,6 dB bei Menschen und 6,5 dB bei Meerschweinchen. Eine auf DPOAE-Wachstumsfunktionen basierende Schwellenschatzung hat gegenuber konventionellen DPOAE-Masen wie DPOAE-Amplitude oder -Rauschabstand den Vorteil, dass mit ihrer Steigung ein unabhangiges Mas zur Verfugung steht, das – wenn auch mit begrenzter Spezifitat – Informationen uber das Mittelohr liefert. Die Kombination mit Verfahren wie der Messung der akustischen Reflektanz oder der Laservibrometrie am Umbo konnte in Zukunft eine zuverlassige Abgrenzung von Mittelohranteilen in DPOAE-Messungen liefern.

A finite-element analysis is used to explore the impact of elastic material properties, boundary conditions, and geometry, including coiling, on the spatial characteristics of the compliance of the unloaded basilar membrane (BM). It is assumed that the arcuate zone is isotropic and the pectinate zone orthotropic, and that the radial component of the effective Young's modulus in the pectinate zone decreases exponentially with distance from base to apex. The results concur with tonotopic characteristics of compliance and neural data. Moreover, whereas the maximum compliance in a radial profile is located close to the boundary between the two zones in the basal region, it shifts to the midpoint of the pectinate zone for the apical BM; the width of the profile also expands. This shift begins near the 1 kHz characteristic place for guinea pig and the 2.4 kHz place for gerbil. Shift and expansion are not observed for linear rather than exponential decrease of the radial component of Young's modulus. This spatial change of the compliance profile leads to the prediction that mechanical excitation in the apical region of the organ of Corti is different to that in the basal region.

Distortion product otoacoustic emissions (DPOAEs) measured as vibration of the human eardrum have been successfully used to estimate hearing threshold. The estimates have proved more accurate than similar methods using sound-pressure DPOAEs. Nevertheless, the estimation accuracy of the new technique might have been influenced by endogenous noise, such as heart beat, breathing and swallowing. Here, we investigate in an animal model to what extent the accuracy of the threshold estimation technique using velocity-DPOAEs might be improved by reducing noise sources. Velocity-DPOAE I/O functions were measured in normal and hearing-impaired anaesthetized guinea pigs. Hearing loss was either conductive or induced by furosemide injection. The estimated distortion product threshold (EDPT) obtained by extrapolation of the I/O function to the abscissa was found to linearly correlate with the compound action potential threshold at the f(2) frequency, provided that furosemide data were excluded. The standard deviation of the linear regression fit was 6 dB as opposed to 8 dB in humans, suggesting that this accuracy should be achievable in humans with appropriate improvement of signal-to-noise ratio. For the furosemide animals, the CAP threshold relative to the regression line provided an estimate of the functional loss of the inner hair cell system. For mechanical losses in the middle ear and/or cochlear amplifier, DPOAEs measured as velocity of the umbo promise an accuracy of hearing threshold estimation comparable to classical audiometry.

Bisher konnten mit dem Laser-Doppler-Vibrometer (LDV) durchgefuhrte Schwingungsmessungen am Trommelfell keine Informationen uber die Kochlea liefern, da die Empfindlichkeit der Gerate nicht ausreichte. Mittels eines von uns neu entwickelten, im Pikometerbereich funktionierenden LDV haben wir bei 20 Probanden Wachstumsfunktionen der Distorsionsprodukte otoakustischer Emissionen (DPOAE) am Umbo gemessen und aus ihnen die Horschwelle geschatzt. Parallel wurden DPOAE-Wachstumsfunktionen mit einer konventionellen, akustischen Sonde gemessen. Die Standardabweichung der aus LDV-Daten geschatzten Horschwellen lag mit 8,6 dB deutlich unter der Standardabweichung der Horschwellen (16,7 dB), die aus akustisch ermittelten DPOAE errechnet wurde. Die geringere Standardabweichung fuhren wir darauf zuruck, dass die laservibrometrisch gemessenen DPOAE weniger anfallig fur Kalibrationsfehler des Schallfelds sind. Mit dem LDV lassen sich DPOAE artefaktfrei messen. Dies erlaubt eine prazise Schatzung der Horschwelle. Schwingungsmessungen am Trommelfell haben damit ein erhebliches differentialdiagnostisches Potential fur Storungen der Mittel- und Innenohrmechanik.

It has previously not been possible to measure eardrum vibration of human subjects in the region of auditory threshold. It is proposed that such measurements should provide information about the status of the mechanical amplifier in the cochlea. It is this amplifier that is responsible for our extraordinary hearing sensitivity. Here, we present results from a laser Doppler vibrometer that we designed to noninvasively probe cochlear mechanics near auditory threshold. This device enables picometer-sized vibration measurements of the human eardrum in vivo . With this sensitivity, we found the eardrum frequency response to be linear down to at least a 20-dB sound pressure level (SPL). Nonlinear cochlear amplification was evaluated with the cubic distortion product of the otoacoustic emissions (DPOAEs) in response to sound stimulation with two tones. DPOAEs originate from mechanical nonlinearity in the cochlea. For stimulus frequencies, f 1 and f 2 , with f 2 / f 1 = 1.2 and f 2 = 4–9.5 kHz, and intensities L 1 and L 2 , with L 1 = 0.4 L 2 + 39 dB and L 2 = 20–65 dB SPL, the DPOAE displacement amplitudes were no more than 8 pm across subjects ( n = 20), with hearing loss up to 16 dB. DPOAE vibration was nonlinearly dependent on vibration at f 2 . The dependence allowed the hearing threshold to be estimated objectively with high accuracy; the standard deviation of the threshold estimate was only 8.6 dB SPL. This device promises to be a powerful tool for differentially characterizing the mechanical condition of the cochlea and middle ear with high accuracy.

INTRODUCTION Active middle-ear implants with floating-mass transducer (FMT) technology are used to treat mild-to-severe sensorineural hearing losses. The standard surgical approach for incus vibroplasty is a mastoidectomy and a posterior tympanotomy, crimping the FMT to the long incus process. An alternative fixation side with less surgical trauma might be the short incus process and incus body.The aim of this study was to develop and test a short incus process coupling device for its functional properties in temporal bone preparations and clinical practice. MATERIALS AND METHODS An extended antrotomy and a posterior tympanotomy were performed in 10 fresh human temporal bones. As a control for normal middle-ear function, the tympanic membrane was stimulated acoustically, and the vibration of the stapes footplate was measured using laser Doppler vibrometry. FMT-induced vibration responses of the stapes were then measured for standard attachment at the long process and for 2 types of couplers designed for attachment at the short process of the incus (SP1 and SP2 coupler). Additionally, the functional outcome in 2 patients provided with an SP2 coupler was assessed postoperatively at 2 weeks, 3 months, and then 11 months, using pure-tone audiometry, auditory thresholds for frequency-modulated (warble) tones, vibroplasty thresholds, and speech audiometry in quiet and noise. RESULTS For the SP2 coupler, velocity-amplitude responses in temporal-bone preparations showed generally similar mean amplitudes as compared with the standard coupling of the FMT to the long process but with clearly increased mean amplitudes between 0.7 and 1.5 kHz and with reduced interindividual variation between 0.5 and 3 kHz. The clinical data of 2 patients with mild-to-severe sensory hearing loss showed good vibroplasty thresholds and convincing results for speech audiometry in quiet (Freiburger monosyllables at 65 dB SPL, 23 ± 31% unaided versus 83 ± 4% aided) and noise (Hochmair-Schulz-Moser-test at 65 dB SPL at 10 dB SNR, 32 ± 45% unaided and 42 ± 29% aided). CONCLUSION The attachment of the FMT to the short incus process with the SP2 coupler leads to good mechanical and functional coupling in an experimental setup and clinical practice.

Outer hair cells (OHCs), the sensory-motor cells responsible for the extraordinary frequency selectivity and dynamic range of the cochlea, rapidly endocytose membrane and protein at their apical surface. Endocytosis and transcytosis in isolated OHCs from the mature guinea-pig cochlea were investigated using the amphipathic membrane probe FM1-43. We observed membrane transport from the apical surface to both the basolateral wall and the subnuclear pole. By double-labelling with DiOC6, a stain for endoplasmic reticulum, and aspiration of the plasma membrane, we showed that the basolateral target was the subsurface cisternae. The fluorescent signal was about three times weaker at the basal than at the apical pole. The speed of vesicle transport to the subnuclear pole was approximately 0.4 microm/s. Changing extracellular Ca2+ concentration from 25 microM to 2 mM accelerated rapid endocytosis. Extracellular application of BAPTA-AM (25 microM), an intracellular Ca2+ chelator, and TFP (20 microM), a specific inhibitor of calmodulin, reduced endocytic activity, as did depolarization of the whole cell. The presence of extracellular Cd2+ (200 microM), a Ca2+-channel blocker, had no effect on the voltage dependence of endocytosis at the apical pole, and inhibited the voltage dependence at the subnuclear pole. These results suggest that rapid endocytosis is a Ca2+/calmodulin-dependent process, with extracellular Ca2+ entering through voltage-gated Ca2+ channels at the basal pole. The two distinct destinations of endocytosed membrane are consistent with the functional polarization of the OHC, with the basolateral wall being dedicated to electromechanical transduction and the subnuclear pole being dedicated to electrochemical transduction processes.

The stereocilia of the cochlear inner hair cells (IHCs) transduce vibrations into the sensory receptor current. Until now, mechanisms for deflecting these stereocilia have not been identified experimentally. Here, we identify a mechanism by using the electromechanical properties of the soma of the outer hair cell to produce an intracochlear, mechanical force stimulus. It is known that the soma of this cell generates mechanical force in response to a change of its transmembrane potential. In the present experiments, the force was induced by intracochlear electrical stimulation at frequencies that covered the entire functionally relevant range of 50 kHz. Vibration responses were measured in the transverse direction with a laser Doppler vibrometer. For frequencies up to approximately 3 kHz in the first three turns of the guinea-pig cochlea, the apical surface of the IHC and the opposing surface of the tectorial membrane were found to vibrate with similar amplitudes but opposite phases. At high frequencies, there was little relative motion between these surfaces in the transverse direction. The counterphasic motion up to approximately 3 kHz results in a pulsatile motion of the fluid surrounding the stereocilia of the IHCs. Based on physical principles of fluid flow between narrowly spaced elastic plates, we show that radial fluid motion is amplified relative to transverse membrane motion and that the radial motion is capable of bending the stereocilia. In conclusion, for frequencies up to at least 3 kHz, there appears to be direct fluid coupling between outer hair cells and IHCs.

There is general agreement that distortion product (DP) otoacoustic emissions elicited by stimuli up to 80–90 dB SPL originate from the saturating nonlinearity of the cochlear amplifier at the basilar membrane site, S, where the responses to the two primary tones overlap. There are, however, different interpretations of how the inner ear transmits the effects of this process to the stapes. The supporters of transmission line models assert that the phenomenon depends upon two main mechanisms: (1) the generation of forward and backward traveling waves (TWs) by DP oscillations at S; (2) the backward propagation of wave components reflected by ‘micromechanical impedance perturbations’ at the sites where the DP TWs peak. However, quantitative predictions based on this view are still lacking. In contrast, here we show, using a nonlinear hydrodynamic model, that the emissions are propagated almost instantaneously through the fluid.

Electromechanical force derived from the soma of the outer hair cell has long been postulated as the basis of the exquisite sensitivity of the cochlea. The problem with this postulate is that the electrical source and mechanical load for the electromechanical outer hair cell might be severely attenuated and phase-shifted by the electrical impedance of the cell and the mechanical impedance of the organ of Corti, respectively. Until now, it has not been possible to experimentally derive the high-frequency electrically induced force at the reticular lamina when the cells are embedded within the organ of Corti. In the study reported here, we succeeded in determining the frequency spectrum of the force up to 50 kHz. This was achieved by measuring both the electrically induced velocity and the mechanical impedance at different radial positions on the reticular lamina without tectorial membrane and with clamped basilar membrane. Velocity was measured with a laser interferometer and impedance, with a magnetically driven atomic force cantilever. The electromechanical force, normalized to the electric current density, exhibited a broad amplitude maximum at 7–20 kHz with a quality factor, Q 3dB , of 0.6 - 0.8. The displacement response was independent of frequency up to 10–20 kHz. The force response compensates for the viscoelastic impedance of the organ of Corti, extending the amplitude response of the organ to high frequencies. It is proposed that the electrical phase response of the cell is compensated with Zwislocki's original mechanism of a parallel resonance in the tectorial membrane–stereocilia complex.

An innovative method is presented to measure the mechanical driving point impedance of biological structures up to at least 40kHz. The technique employs an atomic force cantilever with a ferromagnetic coating and an external magnetic field to apply a calibrated force to the cantilever. Measurement of the resulting cantilever velocity using a laser Doppler vibrometer yields the impedance. A key feature of the method is that it permits measurements for biological tissue in physiological solutions. The method was applied to measure the point impedance of the organ of Corti in situ, to elucidate the biophysical basis of cochlear amplification. The basilar membrane was mechanically clamped at its tympanic surface and the measurements conducted at different radial positions on the reticular lamina. The tectorial membrane was removed. The impedance was described by a generalized Voigt-Kelvin viscoelastic model, in which the stiffness was real-valued and independent of frequency, but the viscosity was complex-valued with positive real part, which was dependent on frequency and negative imaginary part, which was independent of frequency. There was no evidence for an inertial component. The magnitude of the impedance was greatest at the tunnel of Corti, and decreased monotonically in each of the radial directions. In the absence of inertia, the mechanical load on the outer hair cells causes their electromotile displacement responses to be reduced by only 10-fold over the entire range of auditory frequencies.