Anthony W. Gummer

Evidence for active, nonlinear, negative feedback in the vibration response of the apical region of the in-vivo guinea-pig cochlea

The transverse vibration response of the organ of Corti near the apical end of the guinea-pig cochlea was measured in vivo. For cochleae in good physiological condition, as ascertained with threshold compound action potentials and the endocochlear potential, increasing amounts of attenuation and phase lag were found as the intensity was decreased below 80 dB SPL. These nonlinear phenomena disappeared post mortem. The data suggest that an active, nonlinear damping mechanism exists at low intensities at the apex of the cochlea. The phase nonlinearity, evident at all frequencies except at the best frequency (BF), was limited to a total phase change of 0.25 cycles, implying negative feedback of electromechanical force from the outer hair cells into a compliant organ of Corti. The amplitude nonlinearity was largest above BF, possibly due to interaction with a second vibration mode. The high-frequency flank of the amplitude response curve was shifted to lower frequencies by as much as 0.6 octave (oct) for a 50-dB reduction of sound intensity; the reduction of BF was 0.3 oct, but there was no change of relative bandwidth (Q(10 dB)). Detailed frequency responses measured at 60 dB SPL were consistent with non-dispersive, travelling-wave motion: travel time to the place of BF (400 Hz at 60 dB SPL) was 2.9 ms, Q(10 dB) was 1.0; standing-wave motion occurred above 600 Hz. Based on comparison with neural and mechanical data from the base of the cochlea, amplitudes at the apex appear to be sufficient to yield behavioural thresholds. It is concluded that active negative feedback may be a hallmark of the entire cochlea at low stimulus frequencies and that, in contrast to the base, the apex does not require active amplification.

Direct measurement of basilar membrane stiffness in the guinea pig

Measurements of basilar membrane stiffness were made in the 0.8–2.3‐mm region of the guinea pig cochlea, within 1‐h post mortem. A tungsten wire probe glued to a piezoelectric crystal was used to statically displace the scala tympani side of the basilar membrane. The extirpated temporal bone was fixed to a barium titanate crystal and vibrated at 40 Hz with a displacement of 10 nm (rms). The filtered signal from the piezoelectric crystal was analyzed with a lock‐in amplifier and the stiffness calculated as a function of static displacement. The resulting curve was flat for a displacement of 1–3 μm, whereupon it increased, exhibiting two distinct sections, until piercing occurred at 50–60 μm. For measurements made at the membrane center the plateau value of the stiffness decreased exponentially with distance from the stapes with a space constant of 1.7±0.8 mm, in agreement with theory. The radial flexural rigidity and the effective membrane thickness were estimated from the first 10 μm of these curves; the ...

Distortion product otoacoustic emissions measured as vibration on the eardrum of human subjects

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, f1 and f2, with f2/f1 = 1.2 and f2 = 4–9.5 kHz, and intensities L1 and L2, with L1 = 0.4L2 + 39 dB and L2 = 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 f2. 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.

Distortion product otoacoustic emissions in human hypercholesterolemia.

Epidemiological and experimental studies suggest that hypercholesterolemia promotes the development of sensorineural hearing loss; however, the underlying cellular pathomechanism remains obscure. In the present study, 20 healthy subjects and 20 patients with familial hypercholesterolemia were compared with respect to their hearing function. None of the 40 persons reported any history of hearing disorder. In accordance with this subjective impression, mean hearing thresholds were within the normal, age-dependent ranges in both groups. In contrast, the single-generator distortion product otoacoustic emissions (sgDPOAE) were reduced at and above 4 kHz. Input-output functions of DPOAE could be subdivided into three groups: (i) normal, with unity slope at low intensities and slope less than unity (0.24+/-0.07 dB/dB at higher intensities; (ii) pathologic, described by a single straight line; (iii) ill-defined, with data usually indistinguishable from the background noise level. The ill-defined DPOAE behavior was only found in patients with hypercholesterolemia; namely, for 25% of patients at f(2)=1.5 kHz and for 50% at f(2)=4 kHz. Patients belonging to the pathologic and ill-defined DPOAE groups had significantly (P<0.05) higher total serum cholesterol and LDL-cholesterol levels compared with subjects from the normal DPOAE group. While hearing thresholds of patients with ill-defined growth functions were not statistically different from those of normal subjects, speech scores were significantly reduced in these cases. The data imply that nonlinear mechanical processes in the cochlea are compromised in hypercholesterolemic patients.

Pronounced infracuticular endocytosis in mammalian outer hair cells

Endocytosis in cochlear hair cells was investigated by staining with the vital fluorescent dye FM 1-43, that partitions reversibly into membranes and is trapped in vesicles during endocytosis. The temporal development and spatial distribution of FM 1-43 induced fluorescence was investigated using confocal laser-scanning microscopy. FM 1-43 rapidly and intensely stained cochlear hair cells, leaving the supporting cells unstained. For short application (0.2-30 s), only the infracuticular region of outer hair cells (OHCs) was labeled, whereas for long application (30-60 s), the OHCs were also labeled in the infranuclear zone and along a central strand extending from the infracuticular zone down to the nucleus, as well as along the entire cell membrane. Except for the cell membrane, the infracuticular zone, directly below the cuticular plate, showed the most rapid and intense staining, and in most cases staining was spherically shaped with a diameter of 3-7 microm. Localization and size of this infracuticular staining coincided with Hensens body, a specialized variant of the endoplasmic reticulum. In contrast to the OHCs, apical fluorescence of inner hair cells presented a homogeneous distribution. When OHCs were incubated in FM 1-43 for longer than 1 min, many points of contact between the central strand, the infracuticular zone and the lateral cell membrane were observed. Since Hensens bodies are a specialty of OHCs and the fluorescent staining pattern of these cells was unique, it is proposed that Hensens body is involved in the turnover of OHC-specific proteins, such as those involved in the molecular machinery of the motor action of the plasma membrane.

Extraction of sources of distortion product otoacoustic emissions by onset-decomposition.

The cubic component of the distortion product otoacoustic emission (DPOAE) in response to two tones of frequency f(1) and f(2) is generated by so-called primary- and secondary-source mechanisms in the cochlea. Interference between the resulting two source components can limit the usefulness of DPOAEs in assessing cochlear function. Although techniques are available for separating the source components, depending on the application, they can be either time-consuming or ineffective without a priori knowledge of optimal parameters. Here, we investigated, in humans, the possibility of separating the source components in the time-domain by sampling the onset and offset of the DPOAE-time signal at appropriate instants. Therefore, a DPOAE paradigm was developed in which the f(2) tone was periodically switched on during the continuous presence of the f(1) tone. F(2) was increased in 20-Hz steps from 1.5 to 2.5 kHz and the ratio f(2)/f(1) held constant at 1.2; measurements were made at six primary tone levels, ranging from L(2)=25 to 65 dB SPL. To investigate the possibility of separating the two sources by appropriate sampling, we developed an algorithm called onset-decomposition. The algorithm is based on the shape properties of DP-grams constructed from DPOAE responses at different time instants in the onset of the DPOAE signal. Thus, at each such time instant, the source components were extracted by time-windowing of the corresponding DP-gram. The time courses of the amplitude onsets of these separated primary- and secondary-source components provided evidence that the primary-source component attained its steady-state before the secondary-source component started to significantly influence the DPOAE by interference with the primary-source component. Consequently, in the final paradigm, the primary-source component is extracted by sampling the DPOAE signal at a single pre-defined time instant after the onset of the f(2) stimulus tone, before the secondary component begins to interfere. Based on the near-absence of interference maxima and minima in the DP-grams, the appropriate sampling instant was 8-10 ms for all frequencies and intensities in the stimulus set. Extracting the primary-source by onset sampling has the advantage that when individual source components for a given f(2) are to be investigated, there is no need to measure a DP-gram. In conclusion, it is shown that the technique can reliably and quickly separate the source components, making it an attractive paradigm for applications in basic research and clinical diagnosis.

Nonlinearity of mechanoelectrical transduction of outer hair cells as the source of nonlinear basilar-membrane motion and loudness recruitment.

The sound-induced travelling wave in the mammalian cochlea is believed to be enhanced and sharpened by a positive-feedback mechanism, causing the passive, linear growth function of the basilar membrane (BM) to become nonlinear. Based on direct measurements of the receptor potential of isolated outer hair cells, it is shown here how nonlinear BM motion might be due predominantly to the nonlinear growth function of the receptor potential. Since intensity coding in the inner ear is supposed to depend on an interaction of nonlinear BM motion with afferent fibres of different synaptic thresholds, intensity coding is expected to be directly dependent on the mechanoelectrical transduction of outer hair cells (OHC). According to the present experimental data and the feedback concept of outer hair cell action, disruption of the mechanoelectrical transduction of OHC leads to both a reduction of gain and linearizing of the response; that is, to both hearing loss and loudness recruitment.

Conditions for highly efficient and reproducible round-window stimulation in humans.

Round-window stimulation is a new clinical approach for the application of active middle-ear implants. To investigate factors influencing the efficiency of round-window stimulation, experiments in 6 human temporal bones were performed with different actuator geometries and coupling conditions. The experiments show that the amplitude ratio between stapes and round-window actuator vibration is most efficient when using a 1.0-mm diameter rod with a 30° inclined tip geometry and an attached silicone pad. In this case, the amplitude ratio is 0.34 for frequencies up to 1.5 kHz and 0.27 for frequencies up to 20 kHz, with a standard deviation of only 4–6 dB at most frequencies. The analysis of data presented here and in a companion paper suggests that control of proper round-window membrane pretension as well as the inclined tip geometry are the major requirements for maximal performance.

Experimental determination of the mechanical impedance of atomic force microscopy cantilevers in fluids up to 70 kHz

The high-frequency dynamical response of cantilevers used in atomic force microscopy (AFM) to an external force is important for applications such as dynamic AFM modes and dynamic mechanical measurements on biological tissues. In this paper we present a novel method for experimentally determining the mechanical driving-point impedance of an AFM cantilever up to frequencies of at least 70 kHz. A frequency-independent, point force was achieved by applying an electric field between the cantilever and a sharpened steel electrode positioned near (10 μm) the tip of the cantilever. The velocity of the cantilever in response to the force was measured with a laser interferometer. The method was tested in air and in tetrachloromethane (CCl4), a nonpolar, nondissociating fluid. The results agree with theoretical predictions. In addition, the mechanical impedance of a cantilever near a flat surface was measured.

Abolition of the receptor potential response of isolated mammalian outer hair cells by hair-bundle treatment with elastase : a test of the tip-link hypothesis

To test the hypothesis that the tip-links of hair-cell stereocilia are essential for mechanoelectrical transduction, tip-links of isolated outer hair cells (OHCs) of the guinea-pig cochlea were eliminated with a proteolytic enzyme, elastase, and the influence on the receptor potential measured with the whole-cell patch-clamp technique. Within 45 s of immersion of the hair bundle in 20 IU/ml elastase, the receptor potential in response to direct deflection of the hair bundle was irreversibly abolished. The electrical input impedance of the cell remained unchanged, implying that the channels of the basolateral membrane were not affected by elastase. The effect of elastase on the receptor potential was comparable to changes seen after mechanically induced hair-bundle damage. As a further control, a putative transduction-channel blocker, dihydrostreptomycin (68 microM), which does not affect tip-links, was applied to the hair bundle. Although the receptor potential was also blocked by dihydrostreptomycin, the effect was reversible. The results suggest that tip-links are required for mechanoelectrical transduction of mammalian OHCs.