ABR/DPOAE publications

Hypothesis: Intratympanic (IT) application of dexamethasone will reduce ototoxicity associated with systemic cisplatin therapy. Background: Cisplatin is a common chemotherapeutic drug often dose-limited by ototoxicity attributed to the formation of reactive oxygen and nitrogen species damaging critical inner ear structures. Steroids have been shown to reduce formation of reactive oxygen species and thus may reduce ototoxicity. In the present pilot study, we test this hypothesis by IT administration of dexamethasone in a novel murine model of cisplatin ototoxicity. Methods: Click- and pure-tone-evoked auditory brainstem responses (ABRs) in young CBA/J mice were measured. The first phase consisted of a dosing study to identify the optimal cisplatin dose for ototoxicity. In the next phase, ABR thresholds were measured in cisplatin-treated mice after 5 days of IT injection of 24 mg/ml of dexamethasone in 1 ear and normal saline in the opposite ear to serve as controls. Results: Intraperitoneal injection of 14 mg/kg of cisplatin induces significant hearing loss (click-evoked ABR threshold elevation = 12 +/- 7 dB, mu +/- standard error of the mean) with acceptable mortality (20%). The ears that received IT dexamethasone in cisplatin-treated mice had minimal ABR threshold shifts with the click, 8 and 16 kHz of stimuli. There was no significant difference between IT dexamethasone and IT saline ears at 32 kHz. Conclusion: IT dexamethasone protected the mouse ear against cisplatin-induced ototoxicity in a frequency-dependent manner. The present results suggest that IT dexamethasone may be a safe, simple, and effective intervention that minimizes cisplatin ototoxicity without interfering with the chemotherapeutic actions of cisplatin.

Objectives/Hypothesis: To test whether noise‐exposure, which activates a cochlear immune response with cytokine expression and infiltration of circulating leukocytes could augment the response to antigen (Ag). Study Design: Randomized, prospective, mice. Methods: We sensitized mice to an Ag, injected it intrathecally, and subsequently exposed the mice to noise (8–16 kHz, 90, 100, or 118 dB for 2 hours). Control mice received either noise exposure alone (100 or 118 dB), Ag challenge alone, intrathecal surgery and phosphate‐buffered saline injection or no treatment. Four hours or 7 days later the mice were killed and cochlear sections were evaluated immunohistochemically for CD45, ICAM‐1, and phospho‐nuclear transcription factor‐κB expression. Results: Intrathecal Ag injection caused no hearing loss, but did result in a small immune response. Loud noise (118 dB) caused severe hearing loss and slight inflammation. The number of CD45‐positive cells was significantly greater in the Ag plus‐118 dB noise group relative to the Ag‐alone group or 118 dB noise‐exposure group. ICAM expression was seen in the lower part of the spiral ligament and small vessels within the normal cochlea. The amount of expression increased after Ag injection and acoustic trauma. Activated nuclear transcription factor‐κB occurred in the nuclei of hair cells, supporting cells, spiral ligament fibrocytes, and neurons 4 hours after noise exposure. Conclusions: It seems that noise exposure can activate a cochlear immune response, which in the presence of Ag, allows for greater recruitment of inflammatory cells than occurred in response to Ag alone.

The influence of noise exposure on the parameters of a convolution model of the compound action potential (CAP) was examined. CAPs were recorded in normal-hearing gerbils and in gerbils exposed to a 117dBSPL 8kHz band of noise for various durations. The CAPs were fitted with an analytic CAP to obtain the parameters representing the number of nerve fibers (N), the probability density function [P(t)] from a population of nerve fibers, and the single-unit waveform [U(t)]. The results showed that the analytic CAP fitted the physiologic CAPs well with correlations of approximately 0.90. A subsequent analysis using hierarchical linear modeling quantified the change in the parameters as a function of both signal level and hearing threshold. The results showed that noise exposure caused some of the parameter-level functions to simply shift along the signal level axis in proportion to the amount of hearing loss, whereas others shifted along the signal level axis and steepened. Significant changes occurred in the U(t) parameters, but they were not related to hearing threshold. These results suggest that noise exposure alters the physiology underlying the CAP, some of which can be explained by a simple lack of gain, whereas others may not.

Schairer et al. [(2003). “Effects of peripheral nonlinearity on psychometric functions for forward-masked tones,” J. Acoust. Soc. Am. 133, 1560–1573] demonstrated that cochlear nonlinearity is reflected in psychometric-function (PF) slopes for 4 kHz forward-masked tones. The goals of the current study were to use PF slopes to compare the degree of compression between signal frequencies of 0.25 and 4 kHz in listeners with normal hearing (LNH), and between LNH and listeners with cochlear hearing loss (LHL). Forward-masked thresholds were estimated in LNH and LHL using on- and off-frequency maskers and 0.25 and 4 kHz signals in three experiments. PFs were reconstructed from adaptive-procedure data for each subject in each condition. Trends in PF slopes across conditions suggest comparable compression at 0.25 and 4 kHz, and potentially a wider bandwidth of compression in relative frequency at 0.25 kHz. This is consistent with other recent behavioral studies that revise earlier estimates of less compression at lower frequencies. The preliminary results in LHL demonstrate that PF slopes are abnormally steep at frequencies with HL, but are similar to those for LNH at frequencies with NH. Overall, the results are consistent with the notion that PF slopes reflect degree of cochlear nonlinearity and can be used as an additional measure of compression across frequency.

Objectives/Hypothesis: The cochlear amplifier is required for the exquisite sensitivity of mammalian hearing. Outer hair cells underlie the cochlear amplifier and they are unique in that they maintain an intracellular turgor pressure. Changing the turgor pressure of an isolated outer hair cells through osmotic challenge modulates its ability to produce electromotile force. We sought to determine the effect of osmotic challenge on cochlear function. Study Design: In vivo animal study. Methods: Hypotonic and hypertonic artificial perilymph was perfused through the scala tympani of anesthetized guinea pigs. Cochlear function was assessed by measuring the compound action potential, distortion product otoacoustic emissions, the cochlear microphonic, and the endocochlear potential. Results: Hypotonic perilymph decreased and hypertonic perilymph increased compound action potential and distortion product otoacoustic emission thresholds in a dose‐dependent and reversible manner. The cochlear microphonic quadratic distortion product magnitude increased after hypotonic perfusion and decreased with hypertonic perfusion. There were no changes in the stimulus intensity growth curve of the low‐frequency cochlear microphonic. The endocochlear potential was not affected by perilymph osmolality. Conclusions: These data demonstrate that perilymph osmolality can modulate cochlear function and are consistent with what would be expected if outer hair cells turgor pressure changes the gain of the cochlear amplifier in vivo.

The relationship between outer hair cell (OHC) loss and cochlear sensitivity is still unclear, because in many animal models there exist surviving but dysfunctional OHCs and also injured/dead inner hair cells (IHC). Styrene is an ototoxic agent, which targets and destroys OHCs starting from the third row to the second and first rows depending on the exposure level. The remaining cells may be less affected. In this experiment, rats were exposed to styrene by gavage at different doses (200-800 mg/kg/day) for varying periods (5 days/week for 3-12 weeks). An interesting finding was that the cochlear sensitivity was not affected in a few rats with all OHCs in the third row being destroyed by styrene. A further loss of OHCs was usually accompanied with a linear input/output (I/O) function of cochlear compound action potentials (CAP), indicating the loss of cochlear amplification. However, normal CAP amplitudes at the highest stimulation level of 90 dB SPL were often observed when all OHCs were destroyed, indicating normal function of the remaining IHCs. The OHC-loss/hearing-loss relation appeared to be a sigmoid-type function. Initially, styrene-induced OHC losses (<33%) did not result in a significant threshold shift. Then CAP threshold shift increased dramatically with OHC loss from 33% to 66%. Then, CAP threshold changed less with OHC loss. The data suggest a tri-modal relationship between OHC loss and cochlear amplification. That is, under the condition that all surviving OHCs are ideally functioning, the cochlear amplifier is not affected until 33% of OHCs are absent, then the gain of the amplifier decreases proportionally with the OHC loss, and at last the amplifier may fail completely when more than 67% of OHCs are lost.

Distortion-product otoacoustic emissions (DPOAEs) were measured in a New World primate, the common marmoset (Callithrix jacchus). We determined the optimal primary-tone frequency ratio (f(2)/f(1)) to generate DPOAEs of maximal amplitude between 3 and 24 kHz. The optimal f(2)/f(1), determined by varying f(2)/f(1) from 1.02 to 1.40 using equilevel primary tones, decreased with increasing f(2) frequency between 3 and 17 kHz, and increased at 24 kHz. The optimal f(2)/f(1) ratio increased with increasing primary-tone levels from 50 to 74 dB SPL. When all stimulus parameters were considered, the mean optimal f(2)/f(1) was 1.224-1.226. Additionally, we determined the effect of reducing L(2) below L(1). Decreasing L(2) below L(1) by 0, 5, and 10 dB (f(2)/f(1)=1.21) minimally affected DPOAE strength. DPOAE levels were stronger in females than males and stronger in the right ear than the left, just as in humans. This study is the first to measure OAEs in the marmoset, and the results indicate that the effect of varying the frequency ratio and primary-tone level difference on marmoset DPOAEs is similar to the reported effects in humans and Old World primates.

As part of an ongoing study of age-related changes in auditory processing, sensitivity to small changes in frequency were assessed using the cortical auditory evoked potential, P1-N1-P2, in younger and older adults with normal hearing. Behavioral measures have shown age-related differences in intensity and frequency discrimination that are larger at lower than higher frequencies. However, substantial individual differences and equivocal results among studies have been reported. This variability may reflect differences in tasks and procedures, as well as subject variables, such as hearing sensitivity and level of attention. To minimize these subject variables, the P1-N1-P2 response was investigated using a passive listening paradigm. Subjects were 10 younger and 10 older adults. The P1-N1-P2 was elicited by a 150-ms change in frequency in otherwise continuous 500-Hz and 3000-Hz pure tones presented at 70 dB SPL. P1-N1-P2 threshold was defined as the smallest change in frequency needed to evoke a P1-N1-P2 response. Furthermore, a frequency-dependent aging effect was observed for P1-N1-P2 thresholds, such that older subjects were significantly less sensitive to the frequency change than younger subjects, with significantly larger age-related differences at 500 Hz than at 3000 Hz. Age-related changes in response latencies and amplitude of the P1-N1-P2 response were also evident at 500 and 3000 Hz. These results are consistent with age-related changes in the central auditory system and suggest that changes in frequency discrimination abilities of older adults may be, in part, related to changes in preattentive levels of auditory processing.

The purpose of this study was to determine how closely the auditory brainstem response (ABR) can estimate sensorineural threshold shifts in rats exposed to loud sound. Behavioral and ABR thresholds were obtained for tones or noise before and after exposure to loud sound. The results showed that the ABR threshold shift obtained with tone pips estimated the initial pure-tone threshold shifts to within ±5dB 11% of the time and the permanent pure-tone threshold shifts 55% of the time, both with large errors. Determining behavioral thresholds for the same tone pips used for the ABR did not improve the agreement between the measures. In contrast, the ABR obtained with octave noise estimated the initial threshold shifts for that noise to within ±5dB 25% of the time and the permanent threshold shifts 89% of the time, with much smaller errors. Thus, it appears that the noise-evoked ABR is more accurate in estimating threshold shift than the tone-evoked ABR.

Synucleins are widely expressed synaptic proteins within the central nervous system that have been implicated in such neurodegenerative disorders as Parkinson's disease. In this study, an initial characterization of all three synucleins, alpha-, beta-, and gamma-synuclein, within the cochlea was undertaken. Reverse transcriptase-polymerase chain reaction (PCR) demonstrated all three synuclein mRNA species within microdissected cochlear tissue. Quantitative PCR suggests that beta-synuclein is the most abundantly expressed form, followed by gamma- and then alpha-synuclein. Western blot analysis similarly demonstrates all three synuclein proteins within microdissected cochlear tissue. Immunofluorescence localizes the three synucleins predominantly to the efferent neuronal system at the efferent outer hair cell synapse, with some additional localization within the efferent tunnel-crossing fibers (alpha- and gamma-synuclein), spiral ganglion (beta-synuclein), inner spiral bundle (gamma-synuclein), and stria vascularis (alpha- > beta-synuclein). Developmentally, gamma-synuclein can be seen in the region of the outer hair cells by E19, while alpha- and beta-synuclein do not clearly appear there until approximately P10. Additional studies in a null-mutant gamma-synuclein mouse show no histological changes in the organ of Corti with normal hair cell and spiral ganglion cell counts, and normal ABR and DPOAE thresholds in wild-type vs mutant littermates. Together, these results localize synucleins to the efferent cholinergic neuronal auditory system, pointing to a role in normal auditory function, and raising the potential implications for their role in auditory neurodegenerative disorders. However, gamma-synuclein alone is not required for the development and maintenance of normal hearing through P21. Whether overlapping roles of the other synucleins help compensate for the loss of gamma-synuclein remains to be determined.

Background: The Fischer 344/NHsd rat undergoes age-related, progressive, high-frequency hearing loss beginning at age 12 months. The loss has been linked to defects/death in the outer hair cells related to oxidative stress originating in the mitochondria. Acetyl L-carnitine (ALCAR) is known to enhance mitochondrial bioenergetics and membrane efficiency. Therefore, ALCAR was targeted as a possible pharmacologic intervention to prevent, or even restore, hearing loss from aging. Methods: Three different paradigms were used to deliver ALCAR to aging Fischer 344/NHsd rats. Rats in each condition had their hearing evaluated by auditory brainstem responses before, during, and after treatment. First, 24-month-old rats were given ALCAR (100 mg/kg dissolved 25 mg/ml in saline) by IP injection daily for one month. Second, 18-month-old rats were given ALCAR (100 mg/kg) by oral gavage for 90 days. Third, 15-month-old rats were given ALCAR (100 mg/kg) by oral gavage for 90 days. Control rats in each condition received saline by i.p. injection or gavage. Results: Hearing thresholds of the three sets of ALCAR-treated animals were never significantly different from their matched controls before, during, or after the treatments at any of the five test stimuli (5, 10, 20, and 40 kHz tone bursts and a click). Conclusion: The current study does not provide evidence that age-related hearing loss in the Fischer 344/NHsd rat can be altered with systemic administration of ALCAR.

Studies of the F344 rat have shown a variety of age-related auditory anatomy and physiology changes. The current study was undertaken to clarify the ARHL in the F344 rat, by examining the auditory pathway of the F344/NHsd substrain that is distributed by Harlan Laboratories for research in the United States. The F344/NHsd rat begins to lose its hearing at about 12 months, and by 24 months, there are 50-60 dB auditory brainstem response threshold shifts at 20 and 40 kHz and 20 dB losses at 5-10 kHz. Distortion product otoacoustic emissions (DPOAE) amplitudes at 1.8-12 kHz stimuli were depressed in the older (18-24 months) rats. Amplitude input-output functions of the compound action potential (CAP) were also depressed across frequency. The endocochlear potential (EP) was 90-100 mV in the 3 month old rats. All but one of the 24 month old rats' EPs were in the +75-85 mV range. Tympanometry revealed no differences in middle ear function between the young and older rats. Collectively, these findings suggest damage to the outer hair cells, but anatomical examination of the outer hair cells revealed a relative lack of cell loss compared to the magnitude of the hearing and DPOAE loss.

Cisplatin, a widely used anticancer drug, preferentially damages outer hair cells (OHCs) of the inner ear. In this study, an antibody microarray was used to identify early changes in protein expression in the rat cochlea induced by cisplatin. Only small changes in hearing thresholds (4-34 dB elevation) were detected two days after cisplatin treatment (12 mg/kg). OHC function, measured by otoacoustic emissions, was slightly depressed (10 dB), and little or no receptor cell loss was observed. However, cisplatin induced large changes in the expression of 19 proteins involved in apoptosis, cell survival, or progression through the cell cycle. Fifteen of the proteins are novel to the study of the inner ear. Immunoblotting confirmed increases in the levels of the pro-survival activating transcription factor 2 (ATF2), of pro-apoptotic serine-threonine protein kinase, receptor interacting protein, and a 70/75 kDa nitrotyrosine bearing doublet of unknown function. Anti-nitrotyrosine antibodies localized these oxidatively damaged proteins to the stereocilia of OHCs, the Golgi-centrosome region of Hensen's cells, nuclei of outer pillar cells, and tunnel crossing fibers innervating OHCs. The results of this proteomic analysis reflect the commencement of ototoxic and cell survival responses before the observation of a significant functional or anatomical loss.

The transformation of auditory information from the cochlea to the cortex is a highly nonlinear process. Studies using tone stimuli have revealed that changes in even the most basic parameters of the auditory stimulus can alter neural response properties; for example, a change in stimulus intensity can cause a shift in a neuron's preferred frequency. However, it is not yet clear how such nonlinearities contribute to the processing of spectrotemporal features in complex sounds. Here, we use spectrotemporal receptive fields (STRFs) to characterize the effects of stimulus intensity on feature selectivity in the mammalian inferior colliculus (IC). At low intensities, we find that STRFs are relatively simple, typically consisting of a single excitatory region, indicating that the neural response is simply a reflection of the stimulus amplitude at the preferred frequency. In contrast, we find that STRFs at high intensities typically consist of a combination of an excitatory region and one or more inhibitory regions, often in a spectrotemporally inseparable arrangement, indicating selectivity for complex auditory features. We show that a linear–nonlinear model with the appropriate STRF can predict neural responses to stimuli with a fixed intensity, and we demonstrate that a simple extension of the model with an intensity-dependent STRF can predict responses to stimuli with varying intensity. These results illustrate the complexity of auditory feature selectivity in the IC, but also provide encouraging evidence that the prediction of nonlinear responses to complex stimuli is a tractable problem.

The results reported here for CBA/CaJ mice describe the effects of regular dosing with a common antiretroviral drug combination on outer hair cell (OHC) function using measures of 2f1-f2 distortion product otoacoustic emissions (DPOAEs) and auditory brainstem responses (ABRs). Specifically, experimental mice were treated daily over a 3-mo period with the nucleoside reverse transcriptase inhibitors (NRTIs), zidovudine (ZDV) and lamivudine (3TC), dissolved in their drinking water, while their control counterparts received untreated water. DPOAE levels and ABR detection thresholds prior to and after 12 wk of NRTI treatment did not differ between experimental and control groups. To assess whether NRTI treatment potentiates the adverse effects of noise over-exposure on OHC function, both experimental and control mice were exposed 1 wk later, while still on the drug regimen, to a 10-kHz octave-band noise (OBN) at 105-dB SPL for 1h. A major outcome of the sound over-exposure episode was that the NRTI-pretreated mice showed significantly greater permanent OBN-induced reductions in DPOAE levels at 2 wk postexposure than were observed for the untreated control animals. These findings support the notion that a synergistic relationship exists between certain NRTIs and intense sounds in that such preexposure drug treatments produced greater noise-induced decreases in DPOAE activity than did noise exposure alone. This drug/noise interaction is consistent with the known harmful effects of NRTIs on cellular mitochondrial activity.

The hormone estradiol affects the auditory system both by itself and by its interaction with neuroprotective factors. In this study, we examined the role of estrogen receptors (ERs) in response to auditory trauma. We found a ligand-dependent protective role for ERbeta in the auditory system by investigating mice deficient in ERalpha (ERKO mice), ERbeta (BERKO mice), and aromatase (ARKO mice). Basal auditory brainstem response (ABR) thresholds were similar in all animals. An acoustic trauma causing a temporary hearing loss raised ABR thresholds in male and female BERKO and ARKO mice compared with WT and ERKO mice. The ERalpha-selective agonist, propyl(1H) pyrazole-1,3,5-triyl-trisphenol (PPT), partially protected ARKO mice from trauma, while the ERbeta-selective agonist, 2,3-bis (4-hydroxyphenyl)-propionitrile (DPN), protected WT and ARKO mice. Immunohistochemistry and western blotting confirmed the expression of ERbeta in cochlea of WT males and females. Levels of brain-derived neurotrophic factor (BDNF), a neuroprotective peptide that can be induced by estrogen, was lower in BERKO and ARKO mice compared with WT. DPN treatment increased BDNF expression in ARKO mice. These data indicate ERbeta-mediated neuroprotection involving BDNF in the auditory system of males and females.

Our sense of gravitation and linear acceleration is mediated by stimulation of vestibular hair cells through displacement of otoconia in the utricle and saccule (the gravity receptor organ). We recently showed that otoconin-90 (Oc90) deletion led to formation of giant otoconia. In the present study, we determined the extent to which the giant otoconia affected balance and gravity receptor sensory input and compared the findings with other otoconia mutants. We employed a wide spectrum of balance behavioral tests, including reaching and air-righting reflexes, gait, swimming, beam-crossing, rotorod latencies, and a direct measure of gravity receptor input, vestibular evoked potentials (VsEPs). All tests on homozygous adult mutants consistently ranked the order of imbalance as (from worst to best) Nox3(het)<otopetrin 1(tlt)<Oc90 null<Oc90 wild type and C57Bl/6 mice using systematic statistical comparisons of the frequency of occurrence or the severity of abnormal functions. This order coincides with the degree of otoconia deficiencies and is consistent with VsEP measures. Notably, all mice (except Nox3(het)) showed remarkable learned adaptation to peripheral vestibular deficits by staying on the rotating rod significantly longer in each successive trial, and the rate and extent of such learned improvements ranked the same order as their initial balance ability. Despite the vestibular morbidity, Oc90 null mice had normal hearing, as measured by auditory brainstem responses (ABRs) and distortion products of otoacoustic emissions (DPOAEs). The study demonstrates that the remnant otoconia mass in Oc90 nulls does stimulate the gravity receptor organs, which was likely responsible for the improved balance performance relative to strains with absent otoconia. Furthermore, the combination of direct electrophysiological measures and a series of behavioral tests can be used to interpret the imbalance severity arising from altered inputs from the gravity receptor end organ.

It is well established that the responses of binaural auditory neurons can adapt and change dramatically depending on the nature of a preceding sound. Examples of how the effects of ensuing stimuli play a functional role in auditory processing include motion sensitivity and precedence-like effects. To date, these types of effects have been documented at the level of the midbrain and above. Little is known about sensitivity to ensuing stimuli below in the superior olivary nuclei where binaural response properties are first established. Here we report on single cell responses in the gerbil lateral superior olive, the initial site where sensitivity to interaural level differences is established. In contrast to our expectations we found a robust sensitivity to ensuing stimuli. The majority of the cells we tested (86%), showed substantial suppression and/or enhancement to a designated target stimulus, depending on the nature of a preceding stimulus. Hence, sensitivity to ensuing stimuli is already established at the first synaptic station of binaural processing.

Background Several heavy metals have been shown to have toxic effects on the peripheral and central auditory system. Cadmium (Cd2+) is an environmental contaminant showing a variety of adverse effects. Given the current rate of release into the environment, the amount of Cd2+ present in the human body and the incidence of Cd2+-related diseases are expected to increase. Objective The overall aim of this study was to gain further insights into the mechanism of Cd2+-induced ototoxicity. Methods Cell viability, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), cytochrome c (cyt c), phosphorylated extracellular signal-regulated protein kinase (p-ERK), caspases, morphologic change, and functional changes in HEI-OC1 cells, rat cochlear explants, and mouse cochlea after Cd2+ exposure were measured by flow cytometry, immunohistochemical staining, Western blot analysis, and auditory brainstem response (ABR) recording. Mechanisms underlying Cd2+ototoxicity were studied using inhibitors of different signaling pathways, caspases, and antioxidants. Results Cd2+ exposure caused cell death, ROS generation, MMP loss, cyt c release, activation of caspases, ERK activation, apoptosis, and finally auditory threshold shift. Cd2+ toxicity interfered with inhibitors of cellular signaling pathways, such as ERK and c-jun N-terminal kinase, and with caspase inhibitors, especially inhibitors of caspase-9 and caspase-3. The antioxidants N-acetyl-l-cysteine and ebselen showed a significant protective effect on the Cd2+ toxicity. Conlcusions Cd2+ is ototoxic with a complex underlying mechanism. However, ROS generation may be the cause of the toxicity, and application of antioxidants can prevent the toxic effect.

Otoacoustic emissions, sounds generated by the inner ear, are widely used for diagnosing hearing disorders and studying cochlear mechanics. However, it remains unclear how emissions travel from their generation sites to the cochlear base. The prevailing view is that emissions reach the cochlear base via a backward-traveling wave, a slow-propagating transverse wave, along the cochlear partition. A different view is that emissions propagate to the cochlear base via the cochlear fluids as a compressional wave, a fast longitudinal wave. These theories were experimentally tested in this study by measuring basilar membrane (BM) vibrations at the cubic distortion product (DP) frequency from two longitudinal locations with a laser interferometer. Generation sites of DPs were varied by changing frequencies of primary tones while keeping the frequency ratio constant. Here, we show that BM vibration amplitude and phase at the DP frequency are very similar to responses evoked by external tones. Importantly, the BM vibration phase at a basal location leads that at a more apical location, indicating a traveling wave that propagates in the forward direction. These data are in conflict with the backward- traveling-wave theory but are consistent with the idea that the emission comes out of the cochlea predominantly through compressional waves in the cochlear fluids.