Spontaneous Otoacoustic Emissions in TectaY1870C/+ Mice Reflect Changes in Cochlear Amplification and How It Is Controlled by the Tectorial Membrane.

Spontaneous otoacoustic emissions (SOAEs) recorded from the ear canal in the absence of sound reflect cochlear amplification, an outer hair cell (OHC) process required for the extraordinary sensitivity and frequency selectivity of mammalian hearing. Although wild-type mice rarely emit, those with mutations that influence the tectorial membrane (TM) show an incidence of SOAEs similar to that in humans. In this report, we characterized mice with a missense mutation in Tecta, a gene required for the formation of the striated-sheet matrix within the core of the TM. Mice heterozygous for the Y1870C mutation (TectaY1870C/+ ) are prolific emitters, despite a moderate hearing loss. Additionally, Kimura's membrane, into which the OHC stereocilia insert, separates from the main body of the TM, except at apical cochlear locations. Multimodal SOAEs are also observed in TectaY1870C/+ mice where energy is present at frequencies that are integer multiples of a lower-frequency SOAE (the primary). Second-harmonic SOAEs, at twice the frequency of a lower-frequency primary, are the most frequently observed. These secondary SOAEs are found in spatial regions where stimulus-evoked OAEs are small or in the noise floor. Introduction of high-level suppressors just above the primary SOAE frequency reduce or eliminate both primary and second-harmonic SOAEs. In contrast, second-harmonic SOAEs are not affected by suppressors, either above or below the second-harmonic SOAE frequency, even when they are much larger in amplitude. Hence, second-harmonic SOAEs do not appear to be spatially separated from their primaries, a finding that has implications for cochlear mechanics and the consequences of changes to TM structure.

Optical Coherence Tomography to Measure Sound-Induced Motions Within the Mouse Organ of Corti In Vivo.

The measurement of mechanical vibrations within the living cochlea is critical to understanding the first nonlinear steps in auditory processing, hair cell stimulation, and cochlear amplification. However, it has proven to be a challenging endeavor. This chapter describes how optical coherence tomography (OCT) can be used to measure vibrations within the tissues of the organ of Corti. These experimental measurements can be performed within the unopened cochlea of living mice routinely and reliably.

Loss of the tectorial membrane protein CEACAM16 enhances spontaneous, stimulus-frequency, and transiently evoked otoacoustic emissions.

α-Tectorin (TECTA), ß-tectorin (TECTB), and carcinoembryonic antigen-related cell adhesion molecule 16 (CEACAM) are secreted glycoproteins that are present in the tectorial membrane (TM), an extracellular structure overlying the hearing organ of the inner ear, the organ of Corti. Previous studies have shown that TECTA and TECTB are both required for formation of the striated-sheet matrix within which collagen fibrils of the TM are imbedded and that CEACAM16 interacts with TECTA. To learn more about the structural and functional significance of CEACAM16, we created a Ceacam16-null mutant mouse. In the absence of CEACAM16, TECTB levels are reduced, a clearly defined striated-sheet matrix does not develop, and Hensen's stripe, a prominent feature in the basal two-thirds of the TM in WT mice, is absent. CEACAM16 is also shown to interact with TECTB, indicating that it may stabilize interactions between TECTA and TECTB. Although brain-stem evoked responses and distortion product otoacoustic emissions are, for most frequencies, normal in young mice lacking CEACAM16, stimulus-frequency and transiently evoked emissions are larger. We also observed spontaneous otoacoustic emissions (SOAEs) in 70% of the homozygous mice. This incidence is remarkable considering that <3% of WT controls have SOAEs. The predominance of SOAEs >15 kHz correlates with the loss of Hensen's stripe. Results from mice lacking CEACAM16 are consistent with the idea that the organ of Corti evolved to maximize the gain of the cochlear amplifier while preventing large oscillations. Changes in TM structure appear to influence the balance between energy generation and dissipation such that the system becomes unstable.

Traveling waves on the organ of corti of the chinchilla cochlea: spatial trajectories of inner hair cell depolarization inferred from responses of auditory-nerve fibers.

Spatial magnitude and phase profiles for inner hair cell (IHC) depolarization throughout the chinchilla cochlea were inferred from responses of auditory-nerve fibers (ANFs) to threshold- and moderate-level tones and tone complexes. Firing-rate profiles for frequencies ≤2 kHz are bimodal, with the major peak at the characteristic place and a secondary peak at 3-5 mm from the extreme base. Response-phase trajectories are synchronous with peak outward stapes displacement at the extreme cochlear base and accumulate 1.5 period lags at the characteristic places. High-frequency phase trajectories are very similar to the trajectories of basilar-membrane peak velocity toward scala tympani. Low-frequency phase trajectories undergo a polarity flip in a region, 6.5-9 mm from the cochlear base, where traveling-wave phase velocity attains a local minimum and a local maximum and where the onset latencies of near-threshold impulse responses computed from responses to near-threshold white noise exhibit a local minimum. That region is the same where frequency-threshold tuning curves of ANFs undergo a shape transition. Since depolarization of IHCs presumably indicates the mechanical stimulus to their stereocilia, the present results suggest that distinct low-frequency forward waves of organ of Corti vibration are launched simultaneously at the extreme base of the cochlea and at the 6.5-9 mm transition region, from where antiphasic reflections arise.

Cochlear compression: effects of low-frequency biasing on quadratic distortion product otoacoustic emission.

Distortion product otoacoustic emissions (DPOAEs) are generated from the nonlinear transduction n cochlear outer hair cells. The transducer function demonstrating a compressive nonlinearity can be estimated from low-frequency modulation of DPOAEs. Experimental results from the gerbils showed that the magnitude of quadratic difference tone (QDT, f2-f1) was either enhanced or suppressed depending on the phase of the low-frequency bias tone. Within one period of the bias tone, QDT magnitudes exhibited two similar modulation patterns, each resembling the absolute value of the second derivative of the transducer function. In the time domain, the center notches of the modulation patterns occurred around the zero crossings of the bias pressure, whereas peaks corresponded to the increase or decrease in bias pressure. Evaluated with respect to the bias pressure, modulated QDT magnitude displayed a double-modulation pattern marked by a separation of the center notches. Loading/unloading of the cochlear transducer or rise/fall in bias pressure shifted the center notch to positive or negative sound pressures, indicating a mechanical hysteresis. These results suggest that QDT arises from the compression that coexists with the active hysteresis in cochlear transduction. Modulation of QDT magnitude reflects the dynamic regulation of cochlear transducer gain and compression.

Tectorial membrane regeneration in acoustically damaged birds: an immunocytochemical technique.

A novel immunocytochemical method was used to determine whether the sound-damaged adult quail ear can repair its tectorial membrane (TM) and to compare the repair in quail to that in chicks. Birds were exposed to an octave band noise with a center frequency of 1.5 kHz at 116 dB SPL for 4 h. The chicks were grouped based on recovery duration (0 and 7 days), while the quail were divided into 0-, 7-, and 14-day recovered groups. At the end of the recovery period, the animals were sacrificed, and their basilar papillae labeled with a TM-specific monoclonal primary antibody solution followed by a diaminobenzidine process. Examinations under a stereoscope revealed that a patch lesion devoid of TM was located on all 0-day recovered papillae. Seven days later, a honeycomb-patterned layer was observed covering the lesion. In 14-day recovered quail ears, the honeycomb layer appeared similar to that seen at 7 days post-exposure. These observations indicated that both chicks and quail were able to repair their TM within 7 days following exposure to intense sound.

Shearing motion in the hearing organ measured by confocal laser heterodyne interferometry.

To investigate the presence of the postulated shearing motion in the micromechanics of the inner ear during sound stimulations we measured the vibratory response of the tectorial membrane and the reticular lamina in the third cochlear turn in an isolated temporal bone preparation using confocal laser heterodyne interferometry. The mechanical response of the tectorial membrane had the same frequency of maxima as the underlying reticular lamina, but was not as sharply tuned. When the two-dimensional motion was calculated from measurements made from several viewing angles it was found that the vibration of the reticular lamina had significant components both normal and tangential to its surface. The tectorial membrane motion, however, was primarily in a direction approximately perpendicular to the surface of the reticular lamina. The results indicate that shearing motion is produced predominantly by the radial motion of the reticular lamina.

Quantitative anatomical basis for a model of micromechanical frequency tuning in the Tokay gecko, Gekko gecko.

The basilar papilla of the Tokay gecko was studied with standard light- and scanning electron microscopy methods. Several parameters thought to be of particular importance for the mechanical response properties of the system were quantitatively measured, separately for the three different hair-cell areas that are typical for this lizard family. In the basal third, papillar structure was very uniform. The apical two-thirds are subdivided into two hair-cell areas running parallel to each other along the papilla and covered by very different types of tectorial material. Both of those areas showed prominent gradients in hair-cell bundle morphology, i.e., in the height of the stereovillar bundles and the number of stereovilli per bundle, as well as in hair cell density and the size of their respective tectorial covering. Based on the direction of the observed anatomical gradients, a 'reverse' tonotopic organization is suggested, with the highest frequencies represented at the apical end.

The effect of acoustic overexposure on the tonotopic organization of the nucleus magnocellularis.

We assessed the effect a sound-induced cochlear lesion had on the tonotopic organization of the nucleus magnocellularis (NM) immediately after acoustic overexposure and following a twelve day recovery period. The acoustic overexposure was a 0.9 kHz tone at 120 dB sound pressure level (SPL) for 48 h. Initially after the acoustic overexposure, the tonotopic organization of the NM was statistically different from that of age-matched controls. Specifically, it appeared that the center frequencies of units in the frequency region of the NM associated with the acoustic overexposure had higher center frequencies than their control counterparts. Following a twelve day recovery period, when threshold sensitivity and frequency selectivity were operating normally, the tonotopic organization of the NM was not statistically different from age-matched controls. We suggest that the sound-induced changes in the tonotopic organization of the NM reflect peripheral damage in the basilar papilla. It has been well documented that similar exposure paradigms produce a loss of short hair cells and a degeneration of the tectorial membrane in the region of the basilar membrane associated with the overexposure. We postulate that the loss of these structures alters the micromechanics and tuning of the basilar membrane which is reflected in the observed changes in NM tonotopy. Following the recovery period, when those structures destroyed by the overexposure had regenerated and basilar membrane micromechanics were operating normally, the tonotopic organization of the NM returned to normal.

Video-enhanced DIC images of the noise-damaged and regenerated chick tectorial membrane.

Exposure of the chick cochlea to acoustic overstimulation results in a loss of hair cells and a disruption of the tectorial membrane. With time, new hair cells are produced to replace those that are lost and, concurrently, a new tectorial membrane is regenerated. Previous studies of tectorial membrane regeneration examined tissues that were fixed and processed for scanning and transmission electron microscopy. This processing results in a considerable shrinkage of the membrane, and, therefore, it was unclear how the noise damage and subsequent regeneration affected the unfixed, in situ structure of the tectorial membrane. We have recently developed techniques for studying the unfixed tectorial membrane with video-enhanced differential-interference-contrast (DIC) light microscopy. Exposure to a 1500-Hz pure tone at 120 dB SPL for 24 h causes localized damage to the hair cells and tectorial membrane in the mid-proximal region of the basilar papilla. Examination of the unfixed membrane immediately after noise exposure shows that the damage to the tectorial membrane is actually caused by the acoustic trauma and is not an artifact of fixation. After 14 days of recovery, a thick, honeycomb of new matrix has grown from the supporting cells in the basilar papilla and has formed new connections with the stereocilia of surviving and regenerating hair cells. Moreover, this new honeycomb has fused with the remainder of the surrounding, undamaged tectorial membrane, thus reestablishing a continuity in the structure of the membrane across both the damaged and undamaged regions of the basilar papilla.