The peripheral vestibular system detects head position and movement through activation of vestibular hair cells (HCs) in vestibular end organs. HCs transmit this information to the CNS by way of primary vestibular afferent neurons. The CNS, in turn, modulates HCs and afferents via the efferent vestibular system (EVS) through activation of cholinergic signaling mechanisms. In mice, we previously demonstrated that activation of muscarinic acetylcholine receptors (mAChRs), during EVS stimulation, gives rise to a slow excitation that takes seconds to peak and tens of seconds to decay back to baseline. This slow excitation is mimicked by muscarine and ablated by the non-selective mAChR blockers scopolamine, atropine, and glycopyrrolate. While five distinct mAChRs (M1-M5) exist, the subtype(s) driving EVS-mediated slow excitation remain unidentified and details on how these mAChRs alter vestibular function is not well understood. The objective of this study is to characterize which mAChR subtypes drive the EVS-mediated slow excitation, and how their activation impacts vestibular physiology and behavior. In C57Bl/6J mice, M3mAChR antagonists were more potent at blocking slow excitation than M1mAChR antagonists, while M2/M4 blockers were ineffective. While unchanged in M2/M4mAChR double KO mice, EVS-mediated slow excitation in M3 mAChR-KO animals were reduced or absent in irregular afferents but appeared unchanged in regular afferents. In agreement, vestibular sensory-evoked potentials (VsEP), known to be predominantly generated from irregular afferents, were significantly less enhanced by mAChR activation in M3mAChR-KO mice compared to controls. Finally, M3mAChR-KO mice display distinct behavioral phenotypes in open field activity, and thermal profiles, and balance beam and forced swim test. M3mAChRs mediate efferent-mediated slow excitation in irregular afferents, while M1mAChRs may drive the same process in regular afferents.

Noisy galvanic vestibular stimulation (nGVS) delivers band-limited subsensory electrical noise at the mastoids to amplify vestibular signaling via stochastic resonance. This review synthesizes human studies involving healthy participants and proposes a practical stimulation protocol and reporting checklist to promote standardization of nGVS in vestibular research and clinical translation. Evidence supports the use of a bilateral bipolar mastoid montage; 1.75-10 cm2 electrodes; zero-mean Gaussian white noise with a 0-30 Hz passband (or 0-640 Hz when replicating prior work or testing cortical or perceptual hypotheses); participant-titrated subsensory intensities (typically 100-400 μA, with fixed protocols often 400-1,000 μA); and task-matched stimulation epochs of 10-30 seconds. Reports should specify the electrode polarity, passband and filter details, amplitude reference (peak, peak-to-peak, or root-mean-square), threshold procedures, and total stimulation dose. Under these conditions, nGVS has been shown to improve vestibulospinal outcomes (reduced postural sway and better gait stability), modulate vestibulo-ocular physiology, and lower vestibular motion detection thresholds. The effects scale with task difficulty and baseline vestibular function; tolerability was favorable at the subsensory level. Priorities for future research include head-to-head tests of montage and frequency, individualized dose-response and after-effect mapping, stratification by baseline vestibular status, and mechanistic biomarkers tied to behavior. Standardized parameters and reporting will enable robust meta-analyses and faster clinical translation.

The inner ear houses both hearing and balance sensory modalities. The hearing and balance organs consist of similar cell types, including sensory hair cells and associated supporting cells. Previously we showed that Notch1 is required for maintaining supporting cell survival during cochlear maturation. To understand the role of Notch during vestibular maturation, we deleted Notch1 from the vestibular organs of both male and female mice at birth. Histological analyses showed a reduction of supporting cells accompanied by an increase in type II hair cells, indicating a conversion of supporting cells to hair cells. Analysis of mature sensory organs indicate the converted hair cells survive, despite a severe reduction of supporting cells. Vestibular sensory evoked potentials (VsEPs), thought to be generated within the striola regions of the maculae, were absent, indicating that NOTCH1 is critical for striolar function. Specialized type I hair cells in the striola failed to develop the complex calyces typical of these cells. Notch1 mutants did not exhibit vestibular behaviors such as circling and head shaking but showed difficulties with tests of balance and swimming. These results indicate that, unlike the cochlea, supporting cells in balance organs retain the plasticity to convert to hair cells which can survive into adulthood. Despite hair cell survival, vestibular function is compromised likely due to the loss of supporting cells and altered innervation.

Calcitonin gene-related peptide (CGRP) has been implicated in migraine, and CGRP-based therapeutics have shown efficacy in the treatment of migraine headaches. CGRP is also present in efferent synapses of the inner ear, so we questioned whether increases in systemic CGRP can act directly on inner ear end organs. In this study, we determined systemic CGRP changes auditory (ABR) and vestibular (VsEP) endorgan potentials and produces migraine behaviors similar to phonophobia and postural control deficits.

We hypothesized that active outer hair cells drive cochlear fluid circulation. The hypothesis was tested by delivering the neurotoxin, kainic acid, to the intact round window of young gerbil cochleae while monitoring auditory responses in the cochlear nucleus. Sounds presented at a modest level significantly expedited kainic acid delivery. When outer-hair-cell motility was suppressed by salicylate, the facilitation effect was compromised. A low-frequency tone was more effective than broadband noise, especially for drug delivery to apical locations. Computational model simulations provided the physical basis for our observation, which incorporated solute diffusion, fluid advection, fluid–structure interaction, and outer-hair-cell motility. Active outer hair cells deformed the organ of Corti like a peristaltic tube to generate apically streaming flows along the tunnel of Corti and basally streaming flows along the scala tympani. Our measurements and simulations coherently suggest that active outer hair cells in the tail region of cochlear traveling waves drive cochlear fluid circulation.

Primary vestibular afferents transmit information from hair cells about head position and movement to the CNS, which is critical for maintaining balance, gaze stability and spatial navigation. The CNS, in turn, modulates hair cells and afferents via the efferent vestibular system (EVS) and its activation of several cholinergic signaling mechanisms. Electrical stimulation of EVS neurons gives rise to three kinetically– and mechanistically-distinct afferent responses including a slow excitation, a fast excitation, and a fast inhibition. EVS-mediated slow excitation is attributed to odd-numbered muscarinic acetylcholine receptors (mAChRs) on the afferent whose activation leads to the closure of a potassium conductance and increased afferent discharge. Likely effector candidates include low-threshold, voltage-gated potassium channels belonging to the KCNQ (Kv7.X) family, which are involved in neuronal excitability across the nervous system and are subject to mAChR modulation. Specifically, KCNQ2/3 heteromeric channels may be the molecular correlates for the M-current, a potassium current that is blocked following the activation of odd-numbered mAChRs. To this end, multiple members of the KCNQ channel family, including KCNQ2 and KCNQ3, are localized to several microdomains within vestibular afferent endings, where they influence afferent excitability and could be targeted by EVS neurons. Additionally, the relative expression of KCNQ subunits appears to vary across the sensory epithelia and among different afferent types. However, it is unclear which KCNQ channel subunits are targeted by mAChR activation and whether that also varies among different afferent classes. Here we show that EVS-mediated slow excitation is blocked and enhanced by the non-selective KCNQ channel blocker XE991 and opener retigabine, respectively. Using KCNQ subunit-selective drugs, we observed that a KCNQ2 blocker blocks the slow response in irregular afferents, while a KCNQ2/3 opener enhances slow responses in regular afferents. The KCNQ2 blockers did not appear to affect resting afferent discharge rates, while KCNQ2/3 or KCNQ2/4 openers decreased afferent excitability. Here, we show pharmacological evidence that KCNQ2/3 subunits are likely targeted by mAChR activation in mammalian vestibular afferents. Additionally, we show that KCNQ3 KO mice have altered resting discharge rate as well as EVS-mediated slow response. These data together suggest that KCNQ channels play a role in slow response and discharge rate of vestibular afferents, which can be modulated by EVS in mammals. ### Competing Interest Statement The authors have declared no competing interest.

Tone-pip-evoked otoacoustic emissions (PEOAEs) are transient-evoked otoacoustic emissions (OAEs) that are hypothesized to originate from reflection of energy near the best-frequency (BF) cochlear place of the stimulus frequency. However, individual PEOAEs have energy with a wide range of delays. We sought to determine whether some PEOAE energy is consistent with having been generated far from BF. PEOAEs from 35 and 47 dB SPL tone pips were obtained by removing pip-stimulus energy by subtracting the ear-canal sound pressure from scaled-down 59 dB SPL tone pips (which evoke relatively small OAEs). PEOAE delays were measured at each peak in the PEOAE absolute-value waveforms. While measuring PEOAEs and auditory-nerve compound action potentials (CAPs), amplification was blocked sequentially from apex to base by cochlear salicylate perfusion. The perfusion time when a CAP was reduced identified when the perfusion reached the tone-pip BF place. The perfusion times when each PEOAE peak was reduced identified where along the cochlea it received cochlear amplification. PEOAEs and CAPs were measured simultaneously using one pip frequency in each ear (1.4 to 4 kHz across 16 ears). Most PEOAE peaks received amplification primarily between the BF place and 1–2 octaves basal of the BF place. PEOAE peaks with short delays received amplification basal of BF place. PEOAE peaks with longer delays sometimes received amplification apical of BF place, consistent with previous stimulus-frequency-OAE results. PEOAEs provide information about cochlear amplification primarily within 1.5 octave of the tone-pip BF place, not about regions > 3 octaves basal of BF.

Objectives: Our recent empirical findings have shown that the auditory nerve compound action potential (CAP) evoked by a low-level tone burst originates from a narrow cochlear region tuned to the tone burst frequency. At moderate to high sound levels, the origins shift to the most sensitive audiometric regions rather than the extended high-frequency regions of the cochlear base. This means that measurements evoked from extended high-frequency sound stimuli can shift toward the apex with increasing level. Here we translate this study to understand the spatial origin of acoustically evoked responses from ears that receive cochlear implants, an emerging area of research and clinical practice that is not completely understood. An essential step is to first understand the influence of the cochlear implant in otherwise naive ears. Our objective was to understand how function of the high-frequency cochlear base, which can be excited by the intense low-frequency sounds that are frequently used for objective intra- and postoperative monitoring, can be influenced by the presence of the cochlear implant. Design: We acoustically evoked responses and made measurements with an electrode placed near the guinea pig round window. The cochlear implant was not utilized for either electrical stimulation or recording purposes. With the cochlear implant in situ, CAPs were acoustically evoked from 2 to 16 kHz tone bursts of various levels while utilizing the slow perfusion of a kainic acid solution from the cochlear apex to the cochlear aqueduct in the base, which sequentially reduced neural responses from finely spaced cochlear frequency regions. This cochlear perfusion technique reveals the spatial origin of evoked potential measurements and provides insight on what influence the presence of an implant has on acoustical hearing. Results: Threshold measurements at 3 to 11 kHz were elevated by implantation. In an individual ear, thresholds were elevated and lowered as cochlear implant was respectively inserted and removed, indicative of “conductive hearing loss” induced by the implant. The maximum threshold elevation occurred at most sensitive region of the naive guinea pig ear (33.66 dB at 8 kHz), making 11 kHz the most sensitive region to acoustic sounds for guinea pig ears with cochlear implants. Conversely, the acute implantation did not affect the low-frequency, 500 Hz thresholds and suprathreshold function, as shown by the auditory nerve overlapped waveform. As the sound pressure level of the tone bursts increased, mean data show that the spatial origin of CAPs along the cochlear length shifted toward the most sensitive cochlear region of implanted ears, not the extended high-frequency cochlear regions. However, data from individual ears showed that after implantation, measurements from moderate to high sound pressure levels originate in places that are unique to each ear. Conclusions: Alterations to function of the cochlear base from the in situ cochlear implant may influence objective measurements of implanted ears that are frequently made with intense low-frequency sound stimuli. Our results from guinea pigs advance the interpretation of measurements used to understand how and when residual acoustic hearing is lost in human ears receiving a cochlear implant.

The inner ear houses two sensory modalities: the hearing organ, located in the cochlea, and the balance organs, located throughout the vestibular regions of the ear. Both hearing and vestibular sensory regions are composed of similar cell types, including hair cells and associated supporting cells. Recently, we showed that Notch1 is required for maintaining supporting cell survival postnatally during cochlear maturation. However, it is not known whether Notch1 plays a similar role in the balance organs of the inner ear. To characterize the role of Notch during vestibular maturation, we conditionally deleted Notch1 from Sox2-expressing cells of the vestibular organs in the mouse at P0/P1. Histological analyses showed a dramatic loss of supporting cells accompanied by an increase in type II hair cells without cell death, indicating the supporting cells are converting to hair cells in the maturing vestibular regions. Analysis of 6-week old animals indicate that the converted hair cells survive, despite the reduction of supporting cells. Interestingly, measurements of vestibular sensory evoked potentials (VsEPs), known to be generated in the striolar regions of the vestibular afferents in the maculae, failed to show a response, indicating that NOTCH1 expression is critical for striolar function postnatally. Consistent with this, we find that the specialized type I hair cells in the striola fail to develop the complex calyces typical of these cells. These defects are likely due to the reduction in supporting cells, which have previously been shown to express factors critical for the striolar region. Similar to other mutants that lack proper striolar development, Notch1 mutants do not exhibit typical vestibular behaviors such as circling and head shaking, but do show difficulties in some vestibular tests, including the balance beam and forced swim test. These results indicate that, unlike the hearing organ in which the supporting cells undergo cell death, supporting cells in the balance regions retain the ability to convert to hair cells during maturation, which survive into adulthood despite the reduction in supporting cells.

Migraine is a severe and chronic neurological disorder that affects ~18% of people worldwide, the majority being female (3:1). It is characterized by recurrent, debilitating headaches and heightened sensory sensitivities. People with migraine may also experience a heightened motion sensitivity that leads to episodes of vertigo and imbalance, termed vestibular migraine (VM). Calcitonin gene-related peptide (CGRP) is implicated in migraine and is believed to act on brain meninges or in subcortical CNS structures, and CGRP-based antagonists have shown efficacy for migraine treatment. CGRP also signals at efferent synapses of the cochlea and vestibular endorgans, but it is unclear if exogenous CGRP can modulate inner ear function at the endorgan level and cause heightened behavioral responses consistent with VM. We tested if intraperitoneally (IP) delivered CGRP to wildtype mice can modulate endorgan potentials to sound [via auditory brainstem responses (ABRs)] and jerk stimuli [via vestibular sensory evoked potentials (VsEPs)]. We also assessed behavioral measures of phonophobia [acoustic startle reflex (ASR)] and static imbalance [postural sway-center of pressure (CoP)] after IP CGRP, and observed female mice exhibited heightened sensitivities to IP CGRP in all assays. Male mice showed similar auditory sensitivity and endorgan effects to CGRP, but systemic CGRP did not modify male postural sway as it did in females. In conclusion, we show that systemically delivered CGRP modulates activity of the auditory and vestibular endorgan responses and elicits behavioral effects that are consistent with symptoms in a disorder like VM.### Competing Interest StatementThe authors have declared no competing interest.

Stimulation of cholinergic efferent neurons innervating the inner ear has profound, well-characterized effects on vestibular and auditory physiology, after activating distinct ACh receptors (AChRs) on afferents and hair cells in peripheral endorgans. Efferent-mediated fast and slow excitation of vestibular afferents are mediated by α4β2*-containing nicotinic AChRs (nAChRs) and muscarinic AChRs (mAChRs), respectively. On the auditory side, efferent-mediated suppression of distortion product otoacoustic emissions (DPOAEs) is mediated by α9α10nAChRs. Previous characterization of these synaptic mechanisms utilized cholinergic drugs, that when systemically administered, also reach the CNS, which may limit their utility in probing efferent function without also considering central effects. Use of peripherally-acting cholinergic drugs with local application strategies may be useful, but this approach has remained relatively unexplored. Using multiple administration routes, we performed a combination of vestibular afferent and DPOAE recordings during efferent stimulation in mouse and turtle to determine whether charged mAChR or α9α10nAChR antagonists, with little CNS entry, can still engage efferent synaptic targets in the inner ear. The charged mAChR antagonists glycopyrrolate and methscopolamine blocked efferent-mediated slow excitation of mouse vestibular afferents following intraperitoneal, middle ear, or direct perilymphatic administration. Both mAChR antagonists were effective when delivered to the middle ear, contralateral to the side of afferent recordings, suggesting they gain vascular access after first entering the perilymphatic compartment. In contrast, charged α9α10nAChR antagonists blocked efferent-mediated suppression of DPOAEs only upon direct perilymphatic application, but failed to reach efferent synapses when systemically administered. These data show that efferent mechanisms are viable targets for further characterizing drug access in the inner ear.

Recording the linear vestibular sensory evoked potential (VsEP) relies on moving the head in a prescribed manner to synchronously activate neurons of the gravity receptor organs. One problematic issue in accomplishing this is the potential coactivation of cochlear neurons. Although the major stimulus parameters required to elicit the vestibular response have been characterized, some of the determinants of auditory coactivation have not been clearly addressed. In the present study, we show that the duration of the linear cranial jerk stimulus plays a critical role in avoiding coactivation of auditory responses during VsEP recordings. Acoustic masking procedures are essential when recording the VsEP, particularly when using stimulus durations of less than 1 ms.

Hearing depends on glutamatergic synaptic transmission mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). AMPARs are tetramers, where inclusion of the GluA2 subunit reduces overall channel conductance and Ca2+ permeability. Cochlear afferent synapses between inner hair cells (IHCs) and auditory nerve fibers (ANFs) contain the AMPAR subunits GluA2, 3, and 4. However, the tetrameric complement of cochlear AMPAR subunits is not known. It was recently shown in mice that chronic intracochlear delivery of IEM-1460, an antagonist selective for GluA2-lacking AMPARs [also known as Ca2+-permeable AMPARs (CP-AMPARs)], before, during, and after acoustic overexposure prevented both the trauma to ANF synapses and the ensuing reduction of cochlear nerve activity in response to sound. Surprisingly, baseline measurements of cochlear function before exposure were unaffected by chronic intracochlear delivery of IEM-1460. This suggested that cochlear afferent synapses contain GluA2-lacking CP-AMPARs alongside GluA2-containing Ca2+-impermeable AMPA receptors (CI-AMPARs), and that the former can be antagonized for protection while the latter remain conductive. Here, we investigated hearing function in the guinea pig during acute local or systemic delivery of CP-AMPAR antagonists. Acute intracochlear delivery of IEM-1460 or systemic delivery of IEM-1460 or IEM-1925 reduced the amplitude of the ANF compound action potential (CAP) significantly, for all tone levels and frequencies, by > 50% without affecting CAP thresholds or distortion product otoacoustic emissions (DPOAE). Following systemic dosing, IEM-1460 levels in cochlear perilymph were ~ 30% of blood levels, on average, consistent with pharmacokinetic properties predicting permeation of the compounds into the brain and ear. Both compounds were metabolically stable with half-lives >5 h in vitro, and elimination half-lives in vivo of 118 min (IEM-1460) and 68 min (IEM-1925). Heart rate monitoring and off-target binding assays suggest an enhanced safety profile for IEM-1925 over IEM-1460. Compound potency on CAP reduction (IC50 ~ 73 μM IEM-1460) was consistent with a mixture of GluA2-lacking and GluA2-containing AMPARs. These data strongly imply that cochlear afferent synapses of the guinea pig contain GluA2-lacking CP-AMPARs. We propose these CP-AMPARs may be acutely antagonized with systemic dosing, to protect from glutamate excitotoxicity, while transmission at GluA2-containing AMPARs persists to mediate hearing during the protection.

Electrical stimulation of the mammalian efferent vestibular system (EVS) predominantly excites primary vestibular afferents along two distinct time scales. Although roles for acetylcholine (ACh) have been demonstrated in other vertebrates, synaptic mechanisms underlying mammalian EVS actions are not well-characterized. To determine if activation of ACh receptors account for efferent-mediated afferent excitation in mammals, we recorded afferent activity from the superior vestibular nerve of anesthetized C57BL/6 mice while stimulating EVS neurons in the brainstem, before and after administration of cholinergic antagonists. Using a normalized coefficient of variation (CV*), we broadly classified vestibular afferents as regularly- (CV* < 0.1) or irregularly-discharging (CV* > 0.1) and characterized their responses to midline or ipsilateral EVS stimulation. Afferent responses to efferent stimulation were predominantly excitatory, grew in amplitude with increasing CV*, and consisted of fast and slow components that could be identified by differences in rise time and post-stimulus duration. Both efferent-mediated excitatory components were larger in irregular afferents with ipsilateral EVS stimulation. Our pharmacological data show, for the first time in mammals, that muscarinic AChR antagonists block efferent-mediated slow excitation whereas the nicotinic AChR antagonist DHβE selectively blocks efferent-mediated fast excitation, while leaving the efferent-mediated slow component intact. These data confirm that mammalian EVS actions are predominantly cholinergic.

Cochlear amplification of basilar membrane traveling waves is thought to occur between a tone's characteristic frequency (CF) place and within one octave basal of the CF. Evidence for this view comes only from the cochlear base. Stimulus-frequency otoacoustic emissions (SFOAEs) provide a noninvasive alternative to direct measurements of cochlear motion that can be measured across a wide range of CF regions. Coherent reflection theory indicates that SFOAEs arise mostly from the peak region of the traveling wave, but several studies using far-basal suppressor tones claimed that SFOAE components originate many octaves basal of CF. We measured SFOAEs while perfusing guinea pig cochleas from apex to base with salicylate or KCl solutions that reduced outer-hair-cell function and SFOAE amplification. Solution effects on inner hair cells reduced auditory nerve compound action potentials (CAPs) and provided reference times for when solutions reached the SFOAE-frequency CF region. As solution flowed from apex to base, SFOAE reductions generally occurred later than CAP reductions and showed that the effects of cochlear amplification usually peaked ∼1/2 octave basal of the CF region. For tones ≥2 kHz, cochlear amplification typically extended ∼1.5 octaves basal of CF, and the data are consistent with coherent reflection theory. SFOAE amplification did not extend to the basal end of the cochlea, even though reticular lamina motion is amplified in this region, which indicates that reticular lamina motion is not directly coupled to basilar membrane traveling waves. Previous reports of SFOAE-frequency residuals produced by suppressor frequencies far above the SFOAE frequency are most likely due to additional sources created by the suppressor. For some tones <2 kHz, SFOAE amplification extended two octaves apical of CF, which highlights that different vibratory motions produce SFOAEs and CAPs, and that the amplification region depends on the cochlear mode of motion considered. The concept that there is a single "cochlear amplification region" needs to be revised.

There is a strong association between endolymphatic hydrops and low-frequency hearing loss, but the origin of the hearing loss remains unknown. A reduction in the number of cochlear afferent synapses between inner hair cells and auditory nerve fibres may be the origin of the low-frequency hearing loss, but this hypothesis has not been directly tested in humans or animals. In humans, measurements of hearing loss and postmortem temporal-bone based measurements of endolymphatic hydrops are generally separated by large amounts of time. In animals, there has not been a good objective, physiologic, and minimally invasive measurement of low-frequency hearing. We overcame this obstacle with the combined use of a reliable surgical approach to ablate the endolymphatic sac in guinea pigs and create endolymphatic hydrops, the Auditory Nerve Overlapped Waveform to measure low-frequency hearing loss (≤ 1 kHz), and immunohistofluorescence-based confocal microscopy to count cochlear synapses. Results showed low- and mid-(1–4 kHz) frequency hearing loss at all postoperative days, 1, 4, and 30. There was no statistically significant loss of cochlear synapses, and there was no correlation between synapse loss and hearing function. We conclude that cochlear afferent synaptic loss is not the origin of the low-frequency hearing loss in the early days following endolymphatic sac ablation. Understanding what is, and is not, the origin of a hearing loss can help guide preventative and therapeutic development.

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere's disease, though the pathophysiologic mechanism(s) remain unclear. In order to adequately study the attributes of endolymphatic hydrops, such as the origins of low-frequency hearing loss, a reliable model is needed. The guinea pig is a good model because it hears in the low-frequency regions that are putatively affected by endolymphatic hydrops. Previous research has demonstrated that endolymphatic hydrops can be induced surgically via intradural or extradural approaches that involve drilling on the endolymphatic duct and sac. However, whether it was possible to create an endolymphatic hydrops model using an extradural approach that avoided dangerous drilling on the endolymphatic duct and sac was unknown. The objective of this study was to demonstrate a revised extradural approach to induce experimental endolymphatic hydrops at 30 days post-operatively by obliterating the endolymphatic sac and injuring the endolymphatic duct with a fine pick. The sample size consisted of seven guinea pigs. Functional measurements of hearing were made and temporal bones were subsequently harvested for histologic analysis. The approach had a success rate of 86% in achieving endolymphatic hydrops. The risk of cerebrospinal fluid leak was minimal. No perioperative deaths or injuries to the posterior semicircular canal occurred in the sample. The presented method demonstrates a safe and reliable way to induce endolymphatic hydrops at a relatively quick time point of 30 days. The clinical implications are that the presented method provides a reliable model to further explore the origins of low-frequency hearing loss that can be associated endolymphatic hydrops.

Little is known about the spatial origins of auditory nerve (AN) compound action potentials (CAPs) evoked by moderate to intense sounds. We studied the spatial origins of AN CAPs evoked by 2- to 16-kHz tone bursts at several sound levels by slowly injecting kainic acid solution into the cochlear apex of anesthetized guinea pigs. As the solution flowed from apex to base, it sequentially reduced CAP responses from low- to high-frequency cochlear regions. The times at which CAPs were reduced, combined with the cochlear location traversed by the solution at that time, showed the cochlear origin of the removed CAP component. For low-level tone bursts, the CAP origin along the cochlea was centered at the characteristic frequency (CF). As sound level increased, the CAP center shifted basally for low-frequency tone bursts but apically for high-frequency tone bursts. The apical shift was surprising because it is opposite the shift expected from AN tuning curve and basilar membrane motion asymmetries. For almost all high-level tone bursts, CAP spatial origins extended over 2 octaves along the cochlea. Surprisingly, CAPs evoked by high-level low-frequency (including 2 kHz) tone bursts showed little CAP contribution from CF regions ≤ 2 kHz. Our results can be mostly explained by spectral splatter from the tone-burst rise times, excitation in AN tuning-curve "tails," and asynchronous AN responses to high-level energy ≤ 2 kHz. This is the first time CAP origins have been identified by a spatially specific technique. Our results show the need for revising the interpretation of the cochlear origins of high-level CAPs-ABR wave 1. NEW & NOTEWORTHY Cochlear compound action potentials (CAPs) and auditory brain stem responses (ABRs) are routinely used in laboratories and clinics. They are typically interpreted as arising from the cochlear region tuned to the stimulus frequency. However, as sound level is increased, the cochlear origins of CAPs from tone bursts of all frequencies become very wide and their centers shift toward the most sensitive cochlear region. The standard interpretation of CAPs and ABRs from moderate to intense stimuli needs revision.