Mary Ann Cheatham

Prestin-Based Outer Hair Cell Motility Is Necessary for Mammalian Cochlear Amplification

It is a central tenet of cochlear neurobiology that mammalian ears rely on a local, mechanical amplification process for their high sensitivity and sharp frequency selectivity. While it is generally agreed that outer hair cells provide the amplification, two mechanisms have been proposed: stereociliary motility and somatic motility. The latter is driven by the motor protein prestin. Electrophysiological phenotyping of a prestin knockout mouse intimated that somatic motility is the amplifier. However, outer hair cells of knockout mice have significantly altered mechanical properties, making this mouse model unsatisfactory. Here, we study a mouse model without alteration to outer hair cell and organ of Corti mechanics or to mechanoelectric transduction, but with diminished prestin function. These animals have knockout-like behavior, demonstrating that prestin-based electromotility is required for cochlear amplification.

Compound action potential (AP) tuning curves

With a tone‐on‐tone masking procedure the compound action potential (AP), elicited by brief tone bursts of set frequency and intensity, was decreased by a constant fraction. The frequency–intensity pairs formed by the masker that yield this decrease generate the AP tuning curve. It is demonstrated that such tuning curves are very similar to both psychophysical tuning curves and single VIIIth‐nerve‐fiber tuning curves. Changes in the properties of these curves are described as functions of stimulus frequency and level, mode of masking (simultaneous and forward), and parameters of the masker.Subject Classification: [43]65.40, [43]65.42, [43]65.58.

Production of cochlear potentials by inner and outer hair cells

Cochlear microphonic and summating potential recordings were obtained from preparations where only either inner hair cells (first‐turn recording) or outer hair cells (fourth‐turn recording) could have contributed to the potentials. A comparison suggests that outer hair cells do produce the preponderance of receptor potentials.Subject Classification: [43]65.40, [43]65.42.

Consequences of neural asynchrony: a case of auditory neuropathy.

AbstractAbstract The neural representation of sensory events depends upon neural synchrony. Auditory neuropathy, a disorder of stimulus-timing-related neural synchrony, provides a model for studying the role of synchrony in auditory perception. This article presents electrophysiological and behavioral data from a rare case of auditory neuropathy in a woman with normal hearing thresholds, making it possible to separate audibility from neuropathy. The experimental results, which encompass a wide range of auditory perceptual abilities and neurophysiologic responses to sound, provide new information linking neural synchrony with auditory perception. Findings illustrate that optimal eighth nerve and auditory brainstem synchrony do not appear to be essential for understanding speech in quiet listening situations. However, synchrony is critical for understanding speech in the presence of noise.

Positive endocochlear potential: mechanism of production by marginal cells of stria vascularis

The positive endocochlear potential (EP+) and high K+ concentration of the endolymph in the scala media of the mammalian cochlea are unusual. They have long been assumed to be due to a putative K-pump in the luminal membrane of the marginal cells of the stria vascularis, which were believed to have a negative internal potential. We show that the cell potential is more positive than the EP+, and that the ion pump is conventional Na,K-ATPase, probably in the basolateral membrane. The latter was determined from experiments in which the ionic environment of the strial cells was controlled by perfusion of the perilymphatic space of the cochlea, in the absence of vascular circulation. While the usual EP+ was maintained by normal perfusate, replacement of Na+ by choline resulted in a negative EP, showing that Na,K-ATPase is necessary for the production of EP+. Elimination of K+ as well as Na+ from the perfusate did not change the value of the negative EP, showing that no K-ATPase is involved.

Cochlear function in Prestin knockout mice

Gross‐potential recordings in mice lacking the Prestin gene indicate that compound action potential (CAP) thresholds are shifted by ∼45 dB at 5 kHz and by ∼60 dB at 33 kHz. However, in order to conclude that outer hair cell (OHC) electromotility is associated with the cochlear amplifier, frequency selectivity must be evaluated and the integrity of the OHCs forward transducer ascertained. The present report demonstrates no frequency selectivity in CAP tuning curves recorded in homozygotes. In addition, CAP input–output functions indicate that responses in knockout mice approach those in controls at high levels where the amplifier has little influence. Although the cochlear microphonic in knockout mice remains ∼12 dB below that in wild‐type mice even at the highest levels, this deficit is thought to reflect hair cell losses in mice lacking prestin. A change in OHC forward transduction is not implied because knockout mice display non‐linear responses similar to those in controls. For example, homozygotes exhibit a bipolar summating potential (SP) with positive responses at high frequencies; negative responses at low frequencies. Measurement of intermodulation distortion also shows that the cubic difference tone, 2f1–f2, is ∼20 dB down from the primaries in both homozygotes and their controls. Because OHCs are the sole generators of the negative SP and because 2f1–f2 is also thought to originate in OHC transduction, these data support the idea that forward transduction is not degraded in OHCs lacking prestin. Finally, application of AM1‐43, which initially enters hair cells through their transducer channels, produces fluorescence in wild‐type and knockout mice indicating transducer channel activity in both inner and outer hair cells.

Prestin and the cochlear amplifier

In non‐mammalian, hair cell‐bearing sense organs amplification is associated with mechano‐electric transducer channels in the stereovilli (commonly called stereocilia). Because mammals possess differentiated outer hair cells (OHC), they also benefit from a novel electromotile process, powered by the motor protein, prestin. Here we consider new work pertaining to this protein and its potential role as the mammalian cochlear amplifier.

Analysis of the Oligomeric Structure of the Motor Protein Prestin

Prestin, a member of the solute carrier family 26, is expressed in the basolateral membrane of outer hair cells. This protein provides the molecular basis for outer hair cell somatic electromotility, which is crucial for the frequency selectivity and sensitivity of mammalian hearing. It has long been known that there are abundantly expressed ∼11-nm protein particles present in the basolateral membrane. These particles were hypothesized to be the motor proteins that drive electromotility. Because the calculated size of a prestin monomer is too small to form an ∼11-nm particle, the possibility of prestin oligomerization was examined. We investigated possible quaternary structures of prestin by lithium dodecyl sulfate-PAGE, perfluoro-octanoate-PAGE, a membrane-based yeast two-hybrid system, and chemical cross-linking experiments. Prestin, obtained from different host or native cells, is resistant to dissociation by lithium dodecyl sulfate and behaves as a stable oligomer on lithium dodecyl sulfate-PAGE. In the membrane-based yeast two-hybrid system, homo-oligomeric interactions between prestin-bait/prestin-prey suggest that prestin molecules can associate with each other. Chemical cross-linking experiments, perfluoro-octanoate-PAGE/Western blot, and affinity purification experiments all indicate that prestin exists as a higher order oligomer, such as a tetramer, in prestin-expressing yeast, mammalian cell lines and native outer hair cells. Our data from experiments using hydrophobic and hydrophilic reducing reagents suggest that the prestin dimer is connected by a disulfide bond embedded in the prestin hydrophobic core. This stable dimer may act as the building block for producing the higher order oligomers that form the ∼11-nm particles in the outer hair cells basolateral membrane.

Cochlear mechanics, nonlinearities, and cochlear potentials

An up‐to‐date description of the relationships among stimulus‐related cochlear potentials and various mechanical events in the inner ear is presented. Of special interest is to consider (1) how well the mechanical tuning characteristics (amplitude and phase) of the basilar membrane are represented in cochlear microphonics and summating potentials, (2) what these potentials reveal about the mode of excitation of the two groups of sensory cells of the cochlea, and (3) how nonlinearities are reflected in the recorded electrical responses.

Carcinoembryonic antigen-related cell adhesion molecule 16 interacts with α-tectorin and is mutated in autosomal dominant hearing loss (DFNA4)

We report on a secreted protein found in mammalian cochlear outer hair cells (OHC) that is a member of the carcinoembryonic antigen-related cell adhesion molecule (CEACAM) family of adhesion proteins. Ceacam16 mRNA is expressed in OHC, and its protein product localizes to the tips of the tallest stereocilia and the tectorial membrane (TM). This specific localization suggests a role in maintaining the integrity of the TM as well as in the connection between the OHC stereocilia and TM, a linkage essential for mechanical amplification. In agreement with this role, CEACAM16 colocalizes and coimmunoprecipitates with the TM protein α-tectorin. In addition, we show that mutation of CEACAM16 leads to autosomal dominant nonsyndromic deafness (ADNSHL) at the autosomal dominant hearing loss (DFNA4) locus. In aggregate, these data identify CEACAM16 as an α-tectorin–interacting protein that concentrates at the point of attachment of the TM to the stereocilia and, when mutated, results in ADNSHL at the DFNA4 locus.