M. Charles Liberman

Adding Insult to Injury: Cochlear Nerve Degeneration after “Temporary” Noise-Induced Hearing Loss

Overexposure to intense sound can cause temporary or permanent hearing loss. Postexposure recovery of threshold sensitivity has been assumed to indicate reversal of damage to delicate mechano-sensory and neural structures of the inner ear and no persistent or delayed consequences for auditory function. Here, we show, using cochlear functional assays and confocal imaging of the inner ear in mouse, that acoustic overexposures causing moderate, but completely reversible, threshold elevation leave cochlear sensory cells intact, but cause acute loss of afferent nerve terminals and delayed degeneration of the cochlear nerve. Results suggest that noise-induced damage to the ear has progressive consequences that are considerably more widespread than are revealed by conventional threshold testing. This primary neurodegeneration should add to difficulties hearing in noisy environments, and could contribute to tinnitus, hyperacusis, and other perceptual anomalies commonly associated with inner ear damage.

Prestin is required for electromotility of the outer hair cell and for the cochlear amplifier.

Hearing sensitivity in mammals is enhanced by more than 40 dB (that is, 100-fold) by mechanical amplification thought to be generated by one class of cochlear sensory cells, the outer hair cells. In addition to the mechano-electrical transduction required for auditory sensation, mammalian outer hair cells also perform electromechanical transduction, whereby transmembrane voltage drives cellular length changes at audio frequencies in vitro. This electromotility is thought to arise through voltage-gated conformational changes in a membrane protein, and prestin has been proposed as this molecular motor. Here we show that targeted deletion of prestin in mice results in loss of outer hair cell electromotility in vitro and a 40–60 dB loss of cochlear sensitivity in vivo, without disruption of mechano-electrical transduction in outer hair cells. In heterozygotes, electromotility is halved and there is a twofold (about 6 dB) increase in cochlear thresholds. These results suggest that prestin is indeed the motor protein, that there is a simple and direct coupling between electromotility and cochlear amplification, and that there is no need to invoke additional active processes to explain cochlear sensitivity in the mammalian ear.

Single-neuron labeling and chronic cochlear pathology. III. Stereocilia damage and alterations of threshold tuning curves

Tuning curves were obtained from 100 to 150 auditory-nerve fibers spanning the range of characteristic frequencies (CFs) in each of eight cases of permanent noise-induced and three cases of permanent kanamycin-induced threshold shift. In each ear, from one to six neurons were intracellularly labeled with horseradish peroxidase. Locating the labeled terminals in plastic-embedded surface preparations of the cochlea enabled us to accurately correlate particular tuning-curve abnormalities with the condition of the sensory cells generating them. The correlations between structural and functional changes suggest that a normal tuning-curve tip requires that the stereocilia on both the IHCs and OHCs (especially those from the first row) be normal. Selective damage to the OHCs is associated with elevation of the tips and hypersensitivity of the tuning-curve tails. This tuning-curve pattern also originates from cochlear regions at the basal border of hair cell lesions where the local hair cells (and their stereocilia) appear completely normal at the light-microscopic level. Total destruction of the OHCs in a region in which the IHCs appear normal (as can happen in cases of kanamycin poisoning) is associated with bowl-shaped tuning curves which appear to lack a tip. Combined damage to the IHCs and OHCs (as typically happens in cases of acoustic trauma) is invariably associated with elevation of both tips and tails on the tuning curve. A framework for the interpretation of the results is suggested in which the activity of the OHCs is transmitted via the tectorial membrane to the tall row of stereocilia on the IHCs.

The cochlear frequency map for the cat: Labeling auditory‐nerve fibers of known characteristic frequency

Iontophoresis of horseradish peroxidase was used to label single auditory nerve fibers after determination of threshold tuning curves and rates of spontaneous discharge. The relation between characteristic frequency (CF) and cochlear longitudinal location is reconstructed from 52 labeled neurons in 16 cochleas. The length of the organ of Corti allotted to an octave of stimulus frequency increases steadily from low to high frequencies. Thus there is not a simple linear-distance-to-log frequency conversion. When comparing cochleas of different total length, the best predictor of CF at a given location is the distance from base or apex expressed as a percentage of the total length. The cochlear frequency map derived from these single-neuron labeling experiments is compared to maps derived by a number of different physiological and psychophysical techniques, and the significance of the similarities and differences is discussed.

Acceleration of Age-Related Hearing Loss by Early Noise Exposure: Evidence of a Misspent Youth

Age-related and noise-induced hearing losses in humans are multifactorial, with contributions from, and potential interactions among, numerous variables that can shape final outcome. A recent retrospective clinical study suggests an age–noise interaction that exacerbates age-related hearing loss in previously noise-damaged ears (Gates et al., 2000). Here, we address the issue in an animal model by comparing noise-induced and age-related hearing loss (NIHL; AHL) in groups of CBA/CaJ mice exposed identically (8–16 kHz noise band at 100 dB sound pressure level for 2 h) but at different ages (4–124 weeks) and held with unexposed cohorts for different postexposure times (2–96 weeks). When evaluated 2 weeks after exposure, maximum threshold shifts in young-exposed animals (4–8 weeks) were 40–50 dB; older-exposed animals (≥16 weeks) showed essentially no shift at the same postexposure time. However, when held for long postexposure times, animals with previous exposure demonstrated AHL and histopathology fundamentally unlike unexposed, aging animals or old-exposed animals held for 2 weeks only. Specifically, they showed substantial, ongoing deterioration of cochlear neural responses, without additional change in preneural responses, and corresponding histologic evidence of primary neural degeneration throughout the cochlea. This was true particularly for young-exposed animals; however, delayed neuropathy was observed in all noise-exposed animals held 96 weeks after exposure, even those that showed no NIHL 2 weeks after exposure. Data suggest that pathologic but sublethal changes initiated by early noise exposure render the inner ears significantly more vulnerable to aging.

Dynamics of Noise-Induced Cellular Injury and Repair in the Mouse Cochlea

To assess the dynamics of noise-induced tissue injury and repair, groups of CBA/CaJ mice were exposed to an octave-band noise for 2 hours at levels of 94, 100, 106, 112, or 116 dB SPL and evaluated at survival times of 0, 12, 24 hours or 1, 2, or 8 weeks. Functional change, assessed via auditory brainstem response (ABR), ranged from a reversible threshold shift (at 94 dB) to a profound permanent loss (at 116 dB). Light microscopic histopathology was assessed in serial thick plastic sections and involved quantitative evaluation of most major cell types within the cochlear duct, including hair cells (and their stereocilia), supporting cells, ganglion cells, spiral ligament fibrocytes, spiral limbus fibrocytes, and the stria vascularis. Morphometry allowed patterns of damage to be systematically assessed as functions of (1) cochlear location, (2) exposure level, and (3) postexposure survival. Insights into mechanisms of acute and chronic noise-induced cellular damage are discussed.

Noise-induced cochlear neuropathy is selective for fibers with low spontaneous rates.

Acoustic overexposure can cause a permanent loss of auditory nerve fibers without destroying cochlear sensory cells, despite complete recovery of cochlear thresholds (Kujawa and Liberman 2009), as measured by gross neural potentials such as the auditory brainstem response (ABR). To address this nominal paradox, we recorded responses from single auditory nerve fibers in guinea pigs exposed to this type of neuropathic noise (4- to 8-kHz octave band at 106 dB SPL for 2 h). Two weeks postexposure, ABR thresholds had recovered to normal, while suprathreshold ABR amplitudes were reduced. Both thresholds and amplitudes of distortion-product otoacoustic emissions fully recovered, suggesting recovery of hair cell function. Loss of up to 30% of auditory-nerve synapses on inner hair cells was confirmed by confocal analysis of the cochlear sensory epithelium immunostained for pre- and postsynaptic markers. In single fiber recordings, at 2 wk postexposure, frequency tuning, dynamic range, postonset adaptation, first-spike latency and its variance, and other basic properties of auditory nerve response were all completely normal in the remaining fibers. The only physiological abnormality was a change in population statistics suggesting a selective loss of fibers with low- and medium-spontaneous rates. Selective loss of these high-threshold fibers would explain how ABR thresholds can recover despite such significant noise-induced neuropathy. A selective loss of high-threshold fibers may contribute to the problems of hearing in noisy environments that characterize the aging auditory system.

Role of α9 Nicotinic ACh Receptor Subunits in the Development and Function of Cochlear Efferent Innervation

Cochlear outer hair cells (OHCs) express alpha9 nACh receptors and are contacted by descending, predominately cholinergic, efferent fibers originating in the CNS. Mice carrying a null mutation for the nACh alpha9 gene were produced to investigate its role(s) in auditory processing and development of hair cell innervation. In alpha9 knockout mice, most OHCs were innervated by one large terminal instead of multiple smaller terminals as in wild types, suggesting a role for the nACh alpha9 subunit in development of mature synaptic connections. Alpha9 knockout mice also failed to show suppression of cochlear responses (compound action potentials, distortion product otoacoustic emissions) during efferent fiber activation, demonstrating the key role alpha9 receptors play in mediating the only known effects of the olivocochlear system.

Spiral Ligament Pathology: A Major Aspect of Age-Related Cochlear Degeneration in C57BL/6 Mice

Data from systematic, light microscopic examination of cochlear histopathology in an age-graded series of C57BL/6 mice (1.5-15 months) were compared with threshold elevations (measured by auditory brain stem response) to elucidate the functionally important structural changes underlying age-related hearing loss in this inbred strain. In addition to quantifying the degree and extent of hair cell and neuronal loss, all structures of the cochlear duct were qualitatively evaluated and any degenerative changes were quantified. Hair cell and neuronal loss patterns suggested two degenerative processes. In the basal half of the cochlea, inner and outer hair cell loss proceeded from base to apex with increasing age, and loss of cochlear neurons was consistent with degeneration occurring secondary to inner hair cell loss. In the apical half of the cochlea with advancing age, there was selective loss of outer hair cells which increased from the middle to the extreme apex. A similar gradient of ganglion cell loss was noted, characterized by widespread somatic aggregation and demyelination. In addition to these changes in hair cells and their innervation, there was widespread degeneration of fibrocytes in the spiral ligament, especially among the type IV cell class. The cell loss in the ligament preceded the loss of hair cells and/or neurons in both space and time suggesting that fibrocyte pathology may be a primary cause of the hearing loss and ultimate sensory cell degeneration in this mouse strain.

Auditory-nerve activity in cats exposed to ototoxic drugs and high-intensity sounds.

The response characteristics of auditory-nerve fibers in normal cats are compared with those in cats exposed to kanamycin and high-intensity sounds. The pathophysiology is characterized by an elevation of the tuning-curve “tips,” which is sometimes associated with hypersensitivity of the “tails.” Plots of unit thresholds are correlated with patterns of sensory-cell losses in the cochlea. There can be significant shifts in unit threshold without significant loss of hair cells; however, significant hair cell loss is always accompanied by highly abnormal unit thresholds. The presence of inner hair cells seems to be essential for the long-term survival of spiral ganglion cells. An incidental observation is that in the “normal” animal there is almost always a prominent “notch” at 3–4 kHz in the plots of threshold at characteristic frequency, which may have been produced by environmental noise.