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Efferent modulation of vestibular afferent excitability is linked to muscarinic signaling cascades that close low-voltage-gated potassium channels (i.e., KCNQ). Here, we show that muscarinic signaling cascades also depolarize the activation range of hyperpolarization-activated cyclic-nucleotide gated (HCN) channels. We compared the voltage activation range and kinetics of HCN channels and induced firing patterns before and after administering the muscarinic acetylcholine receptor (mAChR) agonist oxotremorine-M (Oxo-M) in dissociated vestibular ganglion neurons (VGNs) from rats of either sex using perforated whole-cell patch-clamp methods. Oxo-M depolarized HCN channels' half-activation voltage ( V 1/2 ) and sped up the rate of activation near resting potential twofold. HCN channels in large-diameter and/or transient firing VGN (putative cell bodies of irregular firing neuron from central epithelial zones) had relatively depolarized V 1/2 in control solution and were less sensitive to mAChR activation than those found in small-diameter VGN with sustained firing patterns (putatively belonging to regular firing afferents). The impact of mAChR on HCN channels is not a direct consequence of closing KCNQ channels since pretreating the cells with Linopirdine, a KCNQ channel blocker, did not prevent HCN channel depolarization by Oxo-M. Efferent signaling promoted ion channel configurations that were favorable to highly regular spiking in some VGN, but not others. This is consistent with previous observations that low-voltage gated potassium currents in VGN are conducted by mAChR agonist-sensitive and -insensitive channels. Connecting efferent signaling to HCN channels is significant because of the channel's impact on spike-timing regularity and nonchemical transmission between Type I hair cells and vestibular afferents. SIGNIFICANCE STATEMENT Vestibular afferents express a diverse complement of ion channels. In vitro studies identified low-voltage activated potassium channels and hyperpolarization-activated cyclic-nucleotide gated (HCN) channels as crucial for shaping the timing and sensitivity of afferent responses. Moreover, a network of acetylcholine-releasing efferent neurons controls afferent excitability by closing a subgroup of low-voltage activated potassium channels on the afferent neuron. This work shows that these efferent signaling cascades also enhance the activation of HCN channels by depolarizing their voltage activation range. The size of this effect varies depending on the endogenous properties of the HCN channel and on cell type (as determined by discharge patterns and cell size). Simultaneously controlling two ion-channel groups gives the vestibular efferent system exquisite control over afferent neuron activity., (Copyright © 2023 the authors.)

Aminoglycoside antibiotics, such as gentamicin, are known to have vestibulotoxic effects, including ataxia and disequilibrium. To date, however, the underlying cellular and molecular mechanisms are still unclear. In this study, we determined the role of gentamicin in regulating the sustained delayed rectifier K + current (I DR ) and membrane excitability in vestibular ganglion (VG) neurons in mice. Our results showed that the application of gentamicin to VG neurons decreased the I DR in a concentration-dependent manner, while the transient outward A-type K + current (I A ) remained unaffected. The decrease in I DR induced by gentamicin was independent of G-protein activity and led to a hyperpolarizing shift of the inactivation V half . The analysis of phospho-c-Jun N-terminal kinase (p-JNK) revealed that gentamicin significantly stimulated JNK, while p-ERK and p-p38 remained unaffected. Blocking Kv1 channels with α-dendrotoxin or pretreating VG neurons with the JNK inhibitor II abrogated the gentamicin-induced decrease in I DR . Antagonism of JNK signaling attenuated the gentamicin-induced stimulation of PKA activity, whereas PKA inhibition prevented the I DR response induced by gentamicin. Moreover, gentamicin significantly increased the number of action potentials fired in both phasic and tonic firing type neurons; pretreating VG neurons with the JNK inhibitor II and the blockade of the I DR abolished this effect. Taken together, our results demonstrate that gentamicin decreases the I DR through a G-protein-independent but JNK and PKA-mediated signaling pathways. This gentamicin-induced I DR response mediates VG neuronal hyperexcitability and might contribute to its pharmacological vestibular effects., (Copyright © 2021 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)

Background: Bilateral vestibular hypofunction is associated with chronic disequilibrium, postural instability, and unsteady gait owing to failure of vestibular reflexes that stabilize the eyes, head, and body. A vestibular implant may be effective in alleviating symptoms., Methods: Persons who had had ototoxic (7 participants) or idiopathic (1 participant) bilateral vestibular hypofunction for 2 to 23 years underwent unilateral implantation of a prosthesis that electrically stimulates the three semicircular canal branches of the vestibular nerve. Clinical outcomes included the score on the Bruininks-Oseretsky Test of Motor Proficiency balance subtest (range, 0 to 36, with higher scores indicating better balance), time to failure on the modified Romberg test (range, 0 to 30 seconds), score on the Dynamic Gait Index (range, 0 to 24, with higher scores indicating better gait performance), time needed to complete the Timed Up and Go test, gait speed, pure-tone auditory detection thresholds, speech discrimination scores, and quality of life. We compared participants' results at baseline (before implantation) with those at 6 months (8 participants) and at 1 year (6 participants) with the device set in its usual treatment mode (varying stimulus pulse rate and amplitude to represent rotational head motion) and in a placebo mode (holding pulse rate and amplitude constant)., Results: The median scores at baseline and at 6 months on the Bruininks-Oseretsky test were 17.5 and 21.0, respectively (median within-participant difference, 5.5 points; 95% confidence interval [CI], 0 to 10.0); the median times on the modified Romberg test were 3.6 seconds and 8.3 seconds (difference, 5.1; 95% CI, 1.5 to 27.6); the median scores on the Dynamic Gait Index were 12.5 and 22.5 (difference, 10.5 points; 95% CI, 1.5 to 12.0); the median times on the Timed Up and Go test were 11.0 seconds and 8.7 seconds (difference, 2.3; 95% CI, -1.7 to 5.0); and the median speeds on the gait-speed test were 1.03 m per second and 1.10 m per second (difference, 0.13; 95% CI, -0.25 to 0.30). Placebo-mode testing confirmed that improvements were due to treatment-mode stimulation. Among the 6 participants who were also assessed at 1 year, the median within-participant changes from baseline to 1 year were generally consistent with results at 6 months. Implantation caused ipsilateral hearing loss, with the air-conducted pure-tone average detection threshold at 6 months increasing by 3 to 16 dB in 5 participants and by 74 to 104 dB in 3 participants. Changes in participant-reported disability and quality of life paralleled changes in posture and gait., Conclusions: Six months and 1 year after unilateral implantation of a vestibular prosthesis for bilateral vestibular hypofunction, measures of posture, gait, and quality of life were generally in the direction of improvement from baseline, but hearing was reduced in the ear with the implant in all but 1 participant. (Funded by the National Institutes of Health and others; ClinicalTrials.gov number, NCT02725463.)., (Copyright © 2021 Massachusetts Medical Society.)

Vestibular afferent neurons convey information from hair cells in the peripheral vestibular end organs to central nuclei. Primary vestibular afferent neurons can fire action potentials at high rates and afferent firing patterns vary with the position of nerve terminal endings in vestibular neuroepithelia. Terminals contact hair cells as small bouton or large calyx endings. To investigate the role of Na + currents ( I Na ) in firing mechanisms, we investigated biophysical properties of I Na in calyx-bearing afferents. Whole cell patch-clamp recordings were made from calyx terminals in thin slices of gerbil crista at different postnatal ages: immature [postnatal day (P)5-P8, young (P13-P15), and mature (P30-P45)]. A large transient Na + current ( I NaT ) was completely blocked by 300 nM tetrodotoxin (TTX) in mature calyces. In addition, I NaT was accompanied by much smaller persistent Na + currents ( I NaP ) and distinctive resurgent Na + currents ( I NaR ), which were also blocked by TTX. ATX-II, a toxin that slows Na + channel inactivation, enhanced I NaP in immature and mature calyces. 4,9-Anhydro-TTX (4,9-ah-TTX), which selectively blocks Nav1.6 channels, abolished the enhanced I Na in mature, but not immature, calyces. Therefore, Nav1.6 channels mediate a component of I NaT and I NaP in mature calyces, but are minimally expressed at early postnatal days. I NaR was expressed in less than one-third of calyces at P6-P8, but expression increased with development, and in mature cristae I NaR was frequently found in peripheral calyces. I NaR served to increase the availability of Na + channels following brief membrane depolarizations. In current clamp, the rate and regularity of action potential firing decreased in mature peripheral calyces following 4,9-ah-TTX application. Therefore, Nav1.6 channels are upregulated during development, contribute to I NaT , I NaP, and I NaR , and may regulate excitability by enabling higher mean discharge rates in a subpopulation of mature calyx afferents. NEW & NOTEWORTHY Action potential firing patterns differ between groups of afferent neurons innervating vestibular epithelia. We investigated the biophysical properties of Na + currents in specialized vestibular calyx afferent terminals during postnatal development. Mature calyces express Na + currents with transient, persistent, and resurgent components. Nav1.6 channels contribute to resurgent Na + currents and may enhance firing in peripheral calyx afferents. Understanding Na + channels that contribute to vestibular nerve responses has implications for developing new treatments for vestibular dysfunction.

Exposure to high levels of bilirubin in hyperbilirubinemia patients and animal models can result in sensorineural deafness. However, the mechanisms underlying bilirubin-induced damage to the inner ear, including the cochlear and vestibular organs, remain unknown. The present analyses of cochlear and vestibular organotypic cultures obtained from postnatal day 3 rats exposed to bilirubin at varying concentrations (0, 10, 50, 100, or 250 μM) for 24 h revealed that auditory nerve fibers (ANFs) and vestibular nerve endings were destroyed even at low doses (10 and 50 μM). Additionally, as the bilirubin dose increased, spiral ganglion neurons (SGNs) and vestibular ganglion neurons (VGNs) exhibited gradual shrinkage in conjunction with nuclei condensation or fragmentation in a dose-dependent manner. The loss of cochlear and vestibular hair cells (HCs) was only evident in explants treated with the highest concentration of bilirubin (250 μM), and bilirubin-induced major apoptosis most likely occurred via the extrinsic apoptotic pathway. Thus, the present results indicate that inner ear neurons and fibers were more sensitive to, and exhibited more severe damage following, bilirubin-induced neurotoxicity than sensory HCs, which illustrates the underlying causes of auditory neuropathy and vestibulopathy in hyperbilirubinemia patients., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Transient receptor potential vanilloid (TRPV) 4 is a nonselective cation channel expressed in sensory neurons such as those in the dorsal root and trigeminal ganglia, kidney, and inner ear. TRPV4 is activated by mechanical stress, heat, low osmotic pressure, low pH, and phorbol derivatives such as 4α-phorbol 12,13-didecanoate (4α-PDD). We investigated the expression of TRPV4 in rat vestibular ganglion (VG) neurons. The TRPV4 gene was successfully amplified from VG neuron mRNA using reverse-transcription polymerase chain reaction. Furthermore, immunoblotting showed positive expression of TRPV4 protein in VG neurons. Immunohistochemistry indicated that TRPV4 was localized predominantly on the plasma membrane of VG neurons. Calcium (Ca2+) imaging of VG neurons showed that 4α-PDD and/or hypotonic stimuli caused an increase in intracellular Ca2+ concentration ([Ca2+]i) that was almost completely inhibited by ruthenium red, a selective antagonist of TRPV channels. Interestingly, a [Ca2+]i increase was evoked by both hypotonic stimuli and 4α-PDD in approximately 38% of VG neurons. These data indicate that TRPV4 is functionally expressed in VG neurons as an ion channel and that TRPV4 likely participates in VG neurons for vestibular neurotransmission as an osmoreceptor and/or mechanoreceptor., (© 2016 S. Karger AG, Basel.)

Optical visualization of neural network activity is limited by imaging system-dependent technical tradeoffs. To overcome these constraints, we have developed a powerful low-cost and flexible imaging system with high spectral variability and unique spatio-temporal precision for simultaneous optical recording and manipulation of neural activity of large cell groups. The system comprises eight high-power light-emitting diodes, a camera with a large metal-oxide-semiconductor sensor and a high numerical aperture water-dipping objective. It allows fast and precise control of excitation and simultaneous low noise imaging at high resolution. Adjustable apertures generated two independent areas of variable size and position for simultaneous optical activation and image capture. The experimental applicability of this system was explored in semi-isolated preparations of larval axolotl (Ambystoma mexicanum) with intact inner ear organs and central nervous circuits. Cyclic galvanic stimulation of semicircular canals together with glutamate- and γ-aminobutyric acid (GABA)-uncaging caused a corresponding modulation of Ca(2+) transients in central vestibular neurons. These experiments revealed specific cellular properties as well as synaptic interactions between excitatory and inhibitory inputs, responsible for spatio-temporal-specific sensory signal processing. Location-specific GABA-uncaging revealed a potent inhibitory shunt of vestibular nerve afferent input in the predominating population of tonic vestibular neurons, indicating a considerable impact of local and commissural inhibitory circuits on the processing of head/body motion-related signals. The discovery of these previously unknown properties of vestibular computations demonstrates the merits of our novel microscope system for experimental applications in the field of neurobiology., (© 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

The primary vestibular neurons convey afferent information from hair cells in the inner ear to the vestibular nuclei and the cerebellum. The intrinsic firing properties of vestibular ganglion cells (VGCs) are heterogeneous to sustained membrane depolarization, and undergo marked developmental changes from phasic to tonic types during the early postnatal period. Previous studies have shown that low-voltage-activated potassium channels, Kv1 and Kv7, play a critical role in determining the firing pattern of VGCs. In the present study, we explored the developmental changes in the properties of hyperpolarization-activated current (Ih) in rat VGCs and the role played by Ih in determining the firing properties of VGCs. Tonic firing VGCs showed a larger current density of Ih as compared to phasic firing VGCs, and tonic firing VGCs became phasic firing in the presence of ZD7288, an Ih channel blocker, indicating that Ih contributes to control the firing pattern of VGCs. The amplitude of Ih increased and the activation kinetics of Ih became faster during the developmental period. Analysis of developmental changes in the expression of hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels revealed that expression of HCN1 protein and its mRNA increased during the developmental period, whereas expression of HCN2-4 protein and its mRNA did not change. Our results suggest that HCN1 channels as well as Kv1 channels are critical in determining the firing pattern of rat VGCs and that developmental up-regulation of HCN1 transforms VGCs from phasic to tonic firing phenotypes., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

During the development of the nervous system, the perinatal period is particularly sensitive as neuronal connections are still forming in the brain of the neonate. Alpha2-adrenergic receptors are overexpressed temporarily in proliferative zones in the developing brain, reaching a peak during the first postnatal week of life. Both stimulation and blocking of these receptors during this period alter the development of neural circuits, affecting synaptic connectivity and neuronal responses. They even affect motor and cognitive skills later on in the adult. It's especially important to look for the early neurological consequences resulting from such modifications, because they may go unnoticed. The main objective of the present study has been to reaffirm the importance of the maturation of alpha-adrenergic system in mice, by carrying out a comprehensive examination of motor, behavioral and cognitive effects in neonates, during early postnatal development, following chronic administration of the drug Clonidine, an alpha2 adrenergic system agonist. Our study shows that mice treated postnatally with clonidine present a temporal delay in the appearance of developmental markers, a slow execution of vestibular reflexes during first postnatal week of life and a blockade of the short term memory in the novel object recognition task. Shortly after the treatment the startle response is hyperreactive.

Both TRPV1 and TRPA1 are non-selective cation channels. They are co-expressed, and interact in sensory neurons such as dorsal root ganglia (DRG) and trigeminal ganglia (TG), and are involved in nociception, being activated by nociceptive stimuli. Immunohistological localization of TRPV1 in vestibular ganglion (VG) neurons has been reported. Although TRPA1 is co-expressed with TRPV1 in DRG and TG neurons, it is unclear whether TRPA1 channels are expressed in VG neurons. Moreover, it is unknown whether TRPV1 and TRPA1 channels are functional in VG neurons. We investigated the expression of TRPV1 and TRPA1 in rat VG neurons by RT-PCR, in situ hybridization, immunohistochemistry, and Ca(2+) imaging experiments. Both TRPV1 and TRPA1 RT-PCR products were amplified from the mRNA of rat VG neurons. In situ hybridization experiments showed TRPV1 and TRPA1 mRNA expression in the majority of VG neurons. Immunohistochemistry experiments confirmed TRPV1 protein expression. In Ca(2+) imaging experiments, capsaicin, a TRPV1 agonist, induced a significant increase in intracellular calcium ion concentration ([Ca(2+)]i) in rat primary cultured VG neurons, which was almost completely blocked by capsazepine, a TRPV1-specific antagonist. Cinnamaldehyde, a TRPA1 agonist, also caused an increase in [Ca(2+)]i, which was completely inhibited by HC030031, a TRPA1-specific antagonist. Moreover, in some VG neurons, a [Ca(2+)]i increase was evoked by both capsaicin and cinnamaldehyde in the same neuron. In summary, our histological and physiological studies reveal that TRPV1 and TRPA1 are expressed in VG neurons. It is suggested that TRPV1 and TRPA1 in VG neurons might participate in vestibular function and/or dysfunction such as vertigo., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Mechanosensory hair cells of the chicken inner ear are innervated by the peripheral processes of statoacoustic ganglion (SAG) neurons. Members of several morphogen families are expressed within and surrounding the chick inner ear during stages of SAG axon outgrowth and pathfinding. On the basis of their localized expression patterns, we hypothesized that bone morphogenetic proteins (BMPs), fibroblast growth factors (FGFs), and sonic hedgehog (Shh) may function as guidance cues for growing axons and/or may function as trophic factors once axons have reached their targets. To test this hypothesis, three-dimensional collagen cultures were used to grow Embryonic Day 4 (E4) chick SAG explants for 24 h in the presence of purified proteins or beads soaked in proteins. The density of neurite outgrowth was quantified to determine effects on neurite outgrowth. Explants displayed enhanced neurite outgrowth when cultured in the presence of purified BMP4, BMP7, a low concentration of Shh, FGF8, FGF10, or FGF19. In contrast, SAG neurons appeared unresponsive to FGF2. Collagen gel cultures were labeled with terminal dUTP nick-end labeling and immunostained with anti-phosphohistone H3 to determine effects on neuron survival and proliferation, respectively. Treatments that increased neurite outgrowth also yielded significantly fewer apoptotic cells, with no effect on cell proliferation. When presented as focal sources, BMP4, Shh, and FGFs -8, -10, and -19 promoted asymmetric outgrowth from the ganglion in the direction of the beads. BMP7-soaked beads did not induce this response. These results suggest that a subset of morphogens enhance both survival and axon outgrowth of otic neurons., (Copyright © 2011 Wiley Periodicals, Inc.)

Infrared laser irradiation has been established as an appropriate stimulus for primary sensory neurons under conditions where sensory receptor cells are impaired or lost. Yet, development of clinical applications has been impeded by lack of information about the molecular mechanisms underlying the laser-induced neural response. Here, we directly address this question through pharmacological characterization of the biological response evoked by midinfrared irradiation of isolated retinal and vestibular ganglion cells from rodents. Whole cell patch-clamp recordings reveal that both voltage-gated calcium and sodium channels contribute to the laser-evoked neuronal voltage variations (LEVV). In addition, selective blockade of the LEVV by micromolar concentrations of ruthenium red and RN 1734 identifies thermosensitive transient receptor potential vanilloid channels as the primary effectors of the chain reaction triggered by midinfrared laser irradiation. These results have the potential to facilitate greatly the design of future prosthetic devices aimed at restoring neurosensory capacities in disabled patients.

Unlabelled: Motion sickness presents a challenge due to its high incidence and unknown pathogenesis although it is a known fact that a functioning vestibular system is essential for the perception of motion sickness. Recent studies show that the efferent vestibular neurons contain calcitonin gene-related peptide (CGRP). It is a possibility that the CGRP immunoreactivity (CGRPi) fibers of the efferent vestibular system modulate primary afferent input into the central nervous system; thus, making it likely that CGRP plays a key role in motion sickness. To elucidate the relationship between motion sickness and CGRP, the effects of CGRP on the vestibular efferent nucleus and the vestibular nucleus were investigated in rats with motion sickness., Methods: An animal model of motion sickness was created by subjecting rats to rotary stimulation for 30 minutes via a trapezoidal stimulation pattern. The number of CGRPi neurons in the vestibular efferent nucleus at the level of the facial nerve genu and the expression level of CGRPi in the vestibular nucleus of rats were measured. Using the ABC method of immunohistochemistry technique, measurements were taken before and after rotary stimulation. The effects of anisodamine on the expression of CGRP in the vestibular efferent nucleus and the vestibular nucleus of rats with motion sickness were also investigated., Results and Discussion: Both the number of CGRPi neurons in the vestibular efferent nucleus and expression level in the vestibular nucleus increased significantly in rats with motion sickness compared to that of controls. The increase of CGRP expression in rats subjected to rotary stimulation 3 times was greater than those having only one-time stimulation. Administration of anisodamine decreased the expression of CGRP within the vestibular efferent nucleus and the vestibular nucleus in rats subjected to rotary stimulation. In conclusion, CGRP possibly plays a role in motion sickness and its mechanism merits further investigation.

We examined the responsiveness of rat vestibular ganglion neurons (VGNs) to exogenous neurotrophic factors, BDNF, NT-3, and GDNF, during postnatal development in dissociated cultures. VGNs were obtained from postnatal days (P) 1, 3, 7, and 14. After two days in culture, the survival of VGNs in control cultures without any exogenous neurotrophic factors was greater in younger (P1 or 3) rats than older (P7 or 14) rats. None of the three neurotrophic factors used facilitated survival of VGNs from older rats while only BDNF facilitated survival of VGNs from younger rats. BDNF was also effective both for increasing neurite sprouting in VGNs from younger rats and for neurite extension in VGNs from each of the postnatal ages. VGNs from P1 rats showed responsiveness to all three neurotrophic factors in their neurite sprouting. Developmental changes of VGNs in their responsiveness to exogenous neurotrophic factors should be considered in treating or preventing neuronal degeneration caused by peripheral vestibular lesions., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Vestibular loss induces a combination of postural, oculomotor, and perceptive symptoms that are compensated over time. The aim of this study was to analyze the influence of betahistine dihydrochloride on vestibular compensation. A randomized, double-blind, placebo-controlled study was performed in Menière's disease patients who underwent a curative unilateral vestibular neurotomy (UVN). The effects of betahistine treatment were investigated on a broad spectrum of vestibular-induced changes resulting from vestibular loss: body sway, head orientation, ocular cyclotorsion, spontaneous nystagmus, verticality perception, and self-evaluation of the postural stability. The time course of the recovery was compared in 16 patients who received either a placebo or betahistine (24 mg b.i.d.) from 3 days up to 3 months after UVN. Patients were examined before (day -1) and after UVN (days 7, 30, and 90). Results indicate that betahistine reduces the time to recovery by 1 month or more depending on the tested functions. Betahistine was effective as soon as 4 days after treatment administration, and the effect remained during the whole compensation period (up to 3 months). The observed clinical effects may be attributed to an action of betahistine in rebalancing the neuronal activity between contralateral vestibular nuclei.

Background: Posturography methods have been applied in clinical neurootology to evaluate the equilibrium function of patients. Methods of statistical analysis play an important role for improving data processing and to support the interpretation of the results. In contrast to conventional statistics, artificial neural networks are model-free and non-parametric. The aim of the presented study was to investigate how accurately these methods are able to discriminate between healthy and equilibrium-disturbed subjects., Patients and Methods: 51 healthy volunteers participated in this study. 2 static posturography measurements were recorded before and 40 min after alcohol intake (0.4‰-0.6‰). Recorded signals were processed by 4 different methods in order to estimate power spectral densities (0 Hz-25 Hz). 11 different methods of artificial neural networks were investigated. The ability of artificial neural networks for classification was evaluated in patients with an acute unilateral vestibular loss., Results: It turned out that estimating power spectral densities by means of autoregressive modelling and subsequent classification by Support-Vector Machine or by Learning Vector Quantization Networks are most accurate. Validation analysis yielded mean classification errors for the test set of 4.2 ± 2.2%., Conclusions: Analysis of neurootological data by artificial neural networks proved to be a sensitive recognition method of even small changes of the postural system., (© Georg Thieme Verlag KG Stuttgart · New York.)

Melatonin has been reported to decrease nerve activity of medial vestibular nuclei in the rat and is associated with attenuated muscle sympathetic nerve activity (MSNA) responses to baroreceptor unloading in humans. The purpose of this study was to determine if melatonin alters the vestibulosympathetic reflex (VSR) and vestibulocollic reflex (VCR) in humans. In study 1, MSNA, arterial blood pressure, and heart rate were measured in 12 healthy subjects (28 ± 1 yr; 6 men, 6 women) during head-down rotation (HDR) before and 45 min after ingestion of either melatonin (3 mg) or placebo (sucrose). Subjects returned at least 2 days later at the same time of day to repeat the trial after ingesting the opposite treatment (melatonin or placebo). Melatonin significantly attenuated MSNA responses during HDR compared with placebo (burst frequency Δ 4 ± 1 vs. Δ 7 ± 1 bursts/min, and total MSNA Δ 51 ± 20 and Δ 96 ± 15%, respectively; P < 0.02). In study 2, vestibular evoked myogenic potentials (VEMP) were measured in 10 healthy subjects (26 ± 1 yr; 4 men and 6 women) before and after ingestion of 3 mg melatonin. Melatonin did not alter the timing of the p13 and n23 peaks (pre-melatonin 13.2 ± 0.4 and 21.3 ± 0.6 ms vs. post-melatonin 13.5 ± 0.4 and 21.4 ± 0.7 ms, respectively) or the p13-n23 interpeak amplitudes [pre-melatonin 22.5 ± 4.6 arbitrary units (au) and post-melatonin 22.7 ± 4.6 au]. In summary, melatonin attenuates the VSR and supports the concept that melatonin negatively affects orthostatic tolerance. However, melatonin does not alter the VCR in humans suggesting melatonin's effect on the VSR appears to be mediated by the utricles.

Previous studies have demonstrated that following intratympanic gentamicin application in the guinea pigs, vestibular evoked myogenic potentials (VEMPs) were absent regardless of stimulation mode using either air-conducted sound (ACS) stimuli or galvanic vestibular stimulation (GVS). Ultrastructurally, both type I hair cells and their calyx terminals were distorted in the saccular macula. However, little is known about the toxic effects of gentamicin on the vestibular ganglion (VG). In this study, absent ACS- and GVS-VEMPs were noted in all the gentamicin-treated ears (100%), which were confirmed by the substantial loss of sensory hair cells in the saccular macula. Moreover, dramatic up-regulation of growth associated protein-43 (GAP-43) expression was detected in the ipsilateral VG neurons. The mean percentage of substance P-like immunoreactive (SP-LI) neurons in the treated VG (81.8±1.9%) was significantly higher than that in the control VG (68.6±3.3%). Conversely, the mean percentage of neuropeptide Y-like immunoreactive (NPY-LI) neurons in the treated VG (13.7±3.8%) was dramatically lower than that in the control VG (49.0±3.8%). Double labeling results shown 82% of SP-LI and 16% of NPY-LI neurons coexpressed with GAP-43, suggested that SP accumulating coincided with NPY decreasing in regenerating VG neurons after gentamicin treatment. Overall, the changes in SP and NPY expression in VG neurons after gentamicin treatment were like to those in the superior cervical ganglion following sympathectomy., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Conclusion: These findings indicate that application of 4-16-fold diluted Omniscan could have depressant effects on the frog vestibular end organs. Based on cochlear studies on cytotoxicity using 8-fold diluted Omniscan, 16-fold diluted Omniscan is optimal for the inner ear and its application could be useful for visualization of endolymphatic hydrops as well as the control of vertiginous attacks in Meniere's disease., Objectives: Endolymphatic hydrops could be visualized recently by intratympanic injection of Omniscan diluted 8- or 16-fold with saline using three-dimensional fluid attenuated inversion recovery (3-D FLAIR) MRI. However, the effects of the Omniscan on vestibular end organs are not clearly understood. The purpose of this study was to investigate the effects of Omniscan on vestibular end organs both morphologically and physiologically., Materials and Methods: Vestibular hair cells isolated from bullfrogs were loaded with 4-32-fold diluted Omniscan (gadolinium hydrate) and morphological changes were observed. Moreover, compound action potentials of posterior semicircular canals were also observed after application of Omniscan., Results: Application of 4-16-fold diluted Omniscan caused morphological damage of isolated vestibular hair cells and decreased action potentials of the posterior semicircular canal. After application of 32-fold diluted Omniscan, no apparent changes were noted even after 20 min, both morphologically and physiologically.

Conclusion: It is suggested that transient receptor potential vanilloid (TRPV)-1 and -2 may be of pathological significance for sensory cells and ganglions, while TRPV-3 and -4 may play an important part in neuroprotection of the inner ear., Objective: Changes in the expression of TRPV-1, -2, -3, and -4 in gentamicin (GM)-treated mouse inner ear were studied., Materials and Methods: CBA/J mice were used in this study. The localization of TRPV-1, -2, -3, and -4 in the inner ear of both untreated and GM-treated CBA/J animals (intratympanic injection of 5 mg GM) was investigated by immunohistochemistry., Results: TRPV-1, -2, and -3 were co-expressed in the inner ear sensory and ganglion cells, while TRPV-4 was also expressed in the stria vascularis and vestibular dark cells. Following GM treatment, the intensity of immunofluorescent reaction to TRPV-1 and TRPV-2 increased, while that to TRPV-3 and TRPV-4 decreased.

The effect of human and rabbit neutrophilic defensins NP-1 and amonoglycoside antibiotic gentamicin on the synaptic transmission in the afferent synapse of isolated vestibular apparatus of the frog has been comparatively studied. Both defensins proved active in the concentration range of 0.0001 to 1 nM and efficiently decreased the impulse frequency in the afferent nerve fibers in a concentration-dependent manner. No significant differences in the efficiency of rabbit and human defensin NP-1 have been revealed in these experiments. Gentamicin also had an inhibitory effect on the afferent discharge in the concentration range of 10-500 microM (0.5-25 mg/kg). The inhibitory effect of gentamicin on the impulse activity of the vestibular nerve was observed at therapeutic doses. The excitatory effect of the putative neurotransmitter L-glutamate was considerably inhibited by defensin NP-1. These findings suggest that the mechanism of defensin action involves a modification of the synaptic transmission the hair receptor and is mediated by L-glutamate.

Sensitivity to sound and vertigo are often components of migraine. Recent studies suggest that plasma extravasation from intradural blood vessels may contribute to migraine pain. This study documented plasma extravasation in the mouse inner ear after intravenous administration of serotonin (5-HT). Horseradish peroxidase (HRP) was injected intravenously to trace protein extravasation in mice, followed 15 min later by intravenous 5-HT or saline. Forty-five minutes later, mice were euthanized. HRP extravasation was visualized immunohistochemically and quantified densitometrically. Baseline and evoked extravasation in stria vascularis and tectorial membrane were indistinguishable from skin, dura mater and tympanic membrane. Brain parenchyma, Scarpa's ganglion, basal spiral ganglion and modiolus, and the central vestibular nerve segment showed no significant 5-HT-induced extravasation. In contrast, 5-HT produced extravasation in the apical spiral ganglion, modiolus, and intralabyrinthine superior and inferior vestibular nerve. Thus, inner ear plasma extravasation is a potential mechanism for migraine-associated vertigo and sound sensitivity.

Multiunit activity during horizontal sinusoidal motion was recorded from pairs of oculomotor, trochlear, or abducens nerves of an in vitro turtle brainstem preparation that received inputs from intact semicircular canals. Responses of left oculomotor, right trochlear and right abducens nerves were approximately aligned with leftward head velocity, and that of the respective contralateral nerves were in-phase with rightward velocity. We examined the effect of sectioning or injecting lidocaine (1-2 microL of 0.5%) into the right vestibular nerve. Nerve block caused a striking phase shift in the evoked response of right oculomotor and left trochlear nerves, in which (rightward) control responses were replaced by a smaller-amplitude response to leftward table motion. Such "phase-reversed" responses were poorly defined in abducens nerve recordings. Frequency analysis demonstrated that this activity was advanced in phase relative to post-block responses of the respective contralateral nerves, which were in turn phase-advanced relative to pre-block controls. Phase differences were largest (approximately 10 degrees) at low frequencies (approximately 0.1 Hz) and statistically absent at 1 Hz. The phase-reversed responses were further investigated by eliminating individual canal input from the left labyrinth following right nVIII block, which indicated that the activation of the vertical canal afferents is the source of this activity.

It is now well-established that an active cross-talk occurs between neurons and glial cells, in the adult as well as in the developing and regenerating nervous systems. These functional interactions not only actively modulate synaptic transmission, but also support neuronal growth and differentiation. We have investigated the possible existence of a reciprocal interaction between inner ear vestibular neurons and Schwann cells maintained in primary cultures. We show that ATP released by the extending vestibular axons elevates intracellular calcium levels within Schwann cells. Purinergic activation of the Schwann P2X(7) receptor induces the release of neurotrophin BDNF, which occurs via a regulated, tetanus-toxin sensitive, vesicular pathway. BDNF, in turn, is required by the vestibular neuron to support its own survival and growth. Given the massive release of ATP during tissue damage, cross-talk between vestibular neurons and Schwann cells could play a primary role during regeneration.

The time course of the recovery of subjective visual horizontal (SVH) after unilateral vestibular deafferentation by intratympanic instillation of gentamicin was studied. Six patients who underwent intratympanic gentamicin instillation therapy for Meniere's disease (1 man and 5 women, 32 to 69 years of age) were enrolled in this study. For comparison, SVH in 23 healthy subjects (12 men and 11 woman, 23 to 48 years of age) was also measured. The mean +/- SD of SVH in healthy subjects was 0.0 +/- 1.1 deg. All of the 6 patients showed significantly deviated SVH toward the injected side-down at the early stage after the therapy. Although one patient showed recovery of SVH to the normal range 25 days after the injection, the other patients required more time for recovery. Three patients did not show recovery to the normal range after 1 year. On the other hand, spontaneous nystagmus observed using an infrared CCD camera in total dark disappeared after 35 days (median). Patients who had normal vestibular evoked myogenic potentials before the therapy showed a tendency of delay of recovery of SVH. The reasons why the recovery of SVH took longer than the disappearance of spontaneous nystagmus are discussed in this report.

Objective: To measure the frequency dynamics of the vestibular evoked myogenic potential in patients with endolymphatic hydrops., Study Design: A prospective study., Setting: A university hospital., Subjects: The endolymphatic hydrops group consisted of 28 affected ears of patients with definite unilateral Ménière's disease and a control group of 36 ears of 20 healthy volunteers., Interventions: Vestibular evoked myogenic potentials generated by tone bursts at 250, 500, 700, 1,000, 1,500, 2,000, and 4,000 Hz were measured in both groups. Vestibular evoked myogenic potentials were also measured after furosemide administration in six patients in the endolymphatic hydrops group., Main Outcome Measure: The frequency sensitivity of vestibular evoked myogenic potential, as evaluated by p13-n23 normalized amplitude., Results: Peak amplitudes were noted at 500 Hz in the control group and at 1,000 Hz in the endolymphatic hydrops group. After furosemide loading, peak amplitude shifted to a lower frequency in four of six ears., Conclusion: The peak amplitude of vestibular evoked myogenic potentials in the endolymphatic hydrops group was at a higher frequency than in the control group. The frequency of the saccule (nu) should be proportional to radical(tau/sigma), where tau is the tension of membrane and sigma is its density. We advocate the hypothesis that the shift in frequency dynamics of vestibular evoked myogenic potential in patients with endolymphatic hydrops originates from the morphologic features of the saccule, analogous to an expanded balloon.

The transient receptor potential cation channel subfamily V (TRPV) is a non-specific cation ion channel receptor family that is gated by heat, protons, low extracellular osmolarity and arachidonic acid derivatives. Since some of these endogenous agonists of TRPV receptors are reactive oxygen intermediates produced by lipoxygenases, it has been hypothesized that some members of the TRPV family may respond to challenges by reactive oxygen species. This study used real-time PCR to quantitatively track changes in TRPV1-4 mRNA expression in the spiral, vestibular, and trigeminal ganglia and the kidney from kanamycin (KM)-treated mice. TRPV1, TRPV2, TRPV3 and TRPV4 mRNAs were expressed in spiral and vestibular ganglia, and TRPV2 and TRPV1 mRNAs were most predominant in control mice. After KM (700 mg/kg s.c. b.i.d., 14 days), TRPV1 mRNA and protein expression were significantly up-regulated both in the spiral and vestibular ganglia, but expression was unaffected in the trigeminal ganglion and kidney. Real-time PCR also demonstrated a significant down-regulation in TRPV4 mRNA expression in the inner ear ganglia and kidney after KM treatment. All these mRNA and protein expression changes were eliminated by simultaneous administration of dihydroxybenzoate (300 mg/kg s.c. b.i.d., 14 days), an anti-oxidant that blocks KM ototoxicity. It is proposed that up-regulated TRPV1 expression during KM exposure may promote ganglion cell survival by contributing to neuronal depolarization, with KM-induced tinnitus and dizziness as consequences.

Mitochondrial uncoupling proteins are a proton transporter family involved in regulation mitochondrial superoxide and ATP production. Uncoupling proteins are expressed by rat spiral ganglion and vestibular ganglion cells [Hear Res 196 (2004) 39]. This study tests the hypothesis that uncoupling protein expression is up-regulated in response to the reactive oxygen species challenge imposed by kanamycin and antioxidant (2,3-dihydroxybenzoate) treatment in mice. In control C57BL/6, CBA/J and BALB/c mice, mRNA for uncoupling protein 1, uncoupling protein 2, uncoupling protein 3, Slc25a27 (uncoupling protein 4) and Slc25a14 (uncoupling protein 5/BMCP1) was expressed in the spiral and vestibular ganglia. After kanamycin-treatment (700 mg/kg twice daily for 14 days s.c.), uncoupling protein 2 and uncoupling protein 3 mRNA expression increased significantly in spiral and vestibular ganglia and kidney, but was unaffected in cerebral cortex. Significant Slc25a27 (uncoupling protein 4) mRNA up-regulation was also observed in spiral and vestibular ganglia, but not in kidney or cerebral cortex. These effects were blocked by simultaneous administration of kanamycin and 2,3-dihydroxybenzoate (300 mg/kg twice daily for 14 days s.c.). Western immunoblotting and immunohistochemistry confirmed the uncoupling protein 2 and uncoupling protein 3 up-regulation in inner ear. Finally, 2,3-dihydroxybenzoate treatment alone produced an upregulation of uncoupling protein 1 mRNA in the spiral ganglion, vestibular ganglion and cerebral cortex, but not the kidney. Uncoupling protein 2 and uncoupling protein 3 upregulation in the kidney and inner ear ganglia likely reflects their general role as a feedback pathway to reduce mitochondrial superoxide generation. Slc25a27 (uncoupling protein 4) upregulation in the inner ear ganglia, by contrast, is likely to be a secondary response to kanamycin-induced hair cell death. We propose that increased uncoupling protein 2, uncoupling protein 3 and Slc25a27 expression has several neuroprotective effects via reduction in mitochondrial superoxide generation and local thermogenesis, including: (1) reducing mean ROS load to prevent apoptosis, (2) increasing signal-to-noise characteristics of intracellular ROS signaling pathways (e.g. lipoxygenases, growth factor and transcription factors), (3) heat-related alteration of enzyme kinetics and (4) promotion of cell depolarization (activation of heat-gated ion channels).

While most studies have demonstrated damage to the cochlear and vestibular endorgan as the primary site of aminoglycoside toxicity, the effect on the primary afferent neurons of the vestibular ganglion remains to be determined. This study used the unbiased stereology-optical fractionator method to obtain estimates of the vestibular ganglion neuronal number. Archival temporal bone specimens from seven subjects with a history of gentamicin (n=3) and streptomycin (n=4) aminoglycoside ototoxicity were used. The post-ototoxicity survival time ranged from two months to 8 years, with an average of 2.2 years. Seven archival human temporal bone specimens from age-matched subjects with no history of audiovestibular symptoms or ototoxicity served as controls. Group means were compared using unpaired, two-tailed student's t test. The average vestibular ganglion neuronal number in the aminoglycoside ototoxicity group was 20, 733 neurons (CV=0.073), which was significantly lower (p<0.005) than the average number in the age-matched control group of 24, 902 neurons (CV=0.109). These findings may be consistent with either retrograde degeneration or a direct neurotoxic effect of the aminoglycosides on the vestibular ganglion neuron.

Multiple sclerosis is characterized by the presence of multiple plaques within the central nervous system, manifesting as remission and exacerbation of neurologic dysfunction over variable time courses. We present the case of a 20-year-old woman. Before treatment, her auditory brain stem response (ABR) test revealed bilateral prolongation. A caloric test showed canal paresis of the right ear and a normal response on the left. A vestibular evoked myogenic potential (VEMP) test displayed an absent response in the right ear and a delayed response in the left. A magnetic resonance imaging (MRI) scan demonstrated multiple diffuse high signal lesions in the hemispheres, brain stem, and cerebellum. Six months after treatment, the demyelinating plaques were shown to have resolved spontaneously on MRI. Recovery of caloric responses was anticipated. Bilateral prolongation of ABRs remained, but the VEMP test disclosed a normal response in the right ear and a delayed response in the left. Accordingly, in addition to MRI, caloric tests and ABR and VEMP tests are useful in monitoring the evolution of audiovestibular function in patients with multiple sclerosis.

Characterization of synaptic transmission between the inner ear sensory cells and primary neuron dendrites has been hampered by the limited access to the postsynaptic terminals. Because direct physiological recording of postsynaptic currents are difficult to achieve, no information regarding the synaptic and dendritic events are available. This is due to the small size of the postsynaptic afferent nerve endings that do not allow a clear identification, and thus compromise direct electrophysiological recordings of the buttons. To study the physiology of afferent nerve endings, we have developed a two-photon imaging technique in cochlear and vestibular slice preparations from neonatal rats and turtles. This technique is based on a retrograde labeling of afferent nerve endings with high-affinity calcium-sensitive dyes. Dye filling was achieved by 6 h application of the dextran-amine conjugate of calcium green-1. Calcium changes were measured in afferent nerve endings in line scan and time lap mode. To address recording in a near-physiological situation, iontophoretic application of K+ was performed in the area of the stereocilia whereas glutamate was applied at the basal pole of sensory hair cells. Both types of application cause a reversible and sustained increase of Ca2+ in the button of afferent nerve fibers. Typical recordings are presented and potential interests for pharmacological studies of inner ear sensory cell synapses are discussed., (Copyright 2004 Elsevier B.V.)

Extracellular unit responses were recorded from the vestibular nucleus (VN) and medial longitudinal fasciculus during horizontal head rotation of an in vitro turtle brainstem in which the temporal bones remained attached. Units were characterized as type I or type II based on the responses to ipsiversive or contraversive rotation, respectively. Lidocaine injections (0.5-2 microl of 0.5%) into the root of the eighth cranial nerve within the cranium caused rapid effects on unit responses to head rotation. Responses of type I units were reduced by ipsilateral injection but enhanced following contralateral injection. On the other hand, type II units had their responses increased by ipsilateral injections yet decreased by contralateral injections. In approximately half of the type II cells, decrease of the contraversive response was accompanied by the appearance of latent ipsiversive activity. Our findings not only confirm that each eighth nerve has afferents that drive ipsiversive excitation of both vestibular nuclei but also suggest that both nerves compete to dominate a central neuron's vestibular response. These results may be inconsistent with the push-pull vestibular model in which each nerve drives the central neuron with a complementary response that enhances the vestibular output. An alternate model is described in which vestibular neurons receive bilateral excitation, and that excitatory input is antagonized by crossed inhibition during contraversive motion.

Purpose of Review: Ménière's disease is characterized by spontaneous attacks of vertigo, fluctuating sensorineural hearing loss, aural fullness, and tinnitus. The pathologic process involves distortion of the membranous labyrinth with the formation of endolymphatic hydrops. This review describes the pathogenesis and etiology as well as the diagnosis and treatment of Ménière's disease., Recent Findings: Initial management of Ménière's disease can involve a low-salt diet and a diuretic. Treatment with intratympanic injection of gentamicin can be beneficial when vertigo persists despite optimal medical management. Recent studies have shown that gentamicin reduces vestibular function in the treated ear, although complete ablation of this vestibular function is not typically required in order to achieve control of vertigo., Summary: Vertigo is often the most debilitating symptom associated with Ménière's disease. Many treatment options exist for the management of vertigo. Intratympanic injection of gentamicin (low dose) can be used in patients for whom vertigo has not been controlled by medical measures. Ongoing research is providing a greater understanding of the effects of gentamicin on vestibular function and of the mechanisms through which gentamicin leads to control of vertigo.

Background: Ethanol led to disturbed dynamic visual acuity (DVA) during vertical linear acceleration (VLA; amplitude, 5 cm; frequency, 1.2 Hz). The aim of this study was to analyze whether suppression of visual-oculomotor or vestibular pathway is responsible for the disturbance of DVA., Methods: Twenty volunteers were investigated before and after ethanol consumption (mean breath alcohol concentration, 0.32 mg/liter). Vertical eye movements and linear head acceleration were recorded. Tested stimuli were vestibular (VLA in the dark), visual (smooth pursuit), and combined (VLA plus fixation on an earth-fixed target) stimulation; visual suppression (VLA plus fixation of a head-fixed target); static visual acuity; and DVA. Parameters of analysis were gain, sensitivity, eye velocity and amplitude, latency between onset of head acceleration and start of eye movement, correct and wrong answers during static visual acuity and DVA testing, feeling of drunkenness (FOD), and breath alcohol concentration., Results: Both during isolated visual and during combined visual-vestibular stimulation, alcohol induced a significant latency increase. Furthermore, DVA was disturbed after ethanol consumption. Test subjects with a strong alcohol-induced disturbance of DVA presented during isolated visual stimulation a significantly higher latency change than volunteers with a minor alcohol-induced disturbance of DVA. On the basis of the FOD, two groups were formed (one with a slight and one with a strong FOD). The two groups differed significantly concerning the alcohol-induced latency increase during isolated visual stimulation and the alcohol-induced disturbance of DVA., Conclusions: Ethanol leads to a disturbance of the visual-oculomotor system and, thus, even during combined visual-vestibular stimulation, to a latency increase. This "delay" is responsible for the disturbance of DVA. This alcohol-induced suppression of the visual-oculomotor system and the disturbance of DVA show a significantly positive correlation with the subjective FOD.

Objective: To describe possible cases of cochleosaccular endolymphatic hydrops without vertigo., Study Design: Retrospective case report., Setting: University hospital., Patients: Three patients with possible cochleosaccular hydrops without vertigo were studied. The basis of diagnosis was positive result of the furosemide-loading vestibular evoked myogenic potential test, no canal paresis in the caloric test, and recurrent cochlear symptoms or fluctuating low-tone hearing loss., Case Report: In case 1, a 47-year-old woman had recurrent left aural fullness and tinnitus and a few weeks later complained of a floating sensation and could not stand up. The furosemide-loading vestibular evoked myogenic potential test showed a positive result in the left ear. In case 2, a 24-year-old woman complained of a backward falling sensation lasting several seconds; subsequently, a severe floating sensation persisted and she could not stand up for several days. Audiography showed fluctuating low-tone hearing loss in the left ear, and the furosemide-loading vestibular evoked myogenic potential test showed a positive result. In case 3, a 41-year-old woman had a floating sensation while walking and subsequently complained of tinnitus in the left ear. She could not stand up because of a severe floating sensation and, moreover, complained of a sudden falling sensation lasting for several seconds. The furosemide-loading vestibular evoked myogenic potential test indicated a positive result in the left ear., Conclusions: The patients in cases 2 and 3 complained of a short-lasting sensation of falling down. Severe disequilibrium that prohibited standing up was noted in all cases. It was suggested that these symptoms were caused by saccular hydrops.

It has been assumed that aminoglycoside-induced damage to the vestibular inner ear ('vestibulotoxicity') is solely due to the inhibition of mitochondrial protein synthesis. There is now substantial evidence, however, to suggest that this form of neurotoxicity is due, at least in part, to the production of free radicals; either in response to excessive stimulation of the N-methyl-D-aspartate (NMDA) receptor by aminoglycosides, or from the formation of oxidative compounds following the binding of aminoglycosides to iron. If activation of NMDA receptors by aminoglycosides contributes to this hair cell death, it should be possible to modify the chemical structures of the aminoglycosides in ways that minimize the unwanted action. Researchers are also currently evaluating drugs that could be co-administered with aminoglycosides in order to interfere with the apoptotic cascade. For example, NMDA receptor antagonists, nitric oxide synthase inhibitors, caspase inhibitors, neurotrophins and free radical scavengers have been demonstrated to protect against aminoglycoside-induced vestibulotoxicity in experimental studies. The next step will be to pursue these effects using in vivo models and drugs that are likely to have favorable pharmacokinetic and side effect profiles in humans.

For 40 years, the amino acid acetyl-DL-leucine (or isoleucine/Tanganil) has been used in clinical practice to reduce the imbalance and autonomic signs associated with acute vertigo crises. In animal models, acetyl-DL-leucine was shown to accelerate vestibular compensation following unilateral labyrinthectomy, while having only minor effects on normal vestibular function. However, the underlying mechanisms are unknown. In this study, the effect of acetyl-DL-leucine on the activity of central vestibular neurons of the medial vestibular nucleus (MVN) and/or the overall activity of vestibular-related networks was electrophysiologically measured in brainstem slices and in the isolated, in vitro whole brain (IWB) of guinea-pig. Only moderate effects were obtained in normal animals, where both excitatory and inhibitory actions of acetyl-DL-leucine were obtained. However, intracellular recordings from MVN neurons revealed that the nature of the response depended on the resting membrane potential. The neurons excited by acetyl-DL-leucine were significantly hyperpolarized compared to nonsensitive cells, whereas the neurons inhibited by this compound tended to display higher than normal membrane potentials. In accordance with these data, acetyl-DL-leucine reduced the prominent asymmetry characterizing the vestibular-related networks of IWBs taken from previously labyrinthectomized animals, by decreasing the activity of the abnormally depolarized neurons on the hyperactive side. Altogether, our results suggest that acetyl-DL-leucine might act mainly on abnormally hyperpolarized and/or depolarized MVN neurons, by bringing back their membrane potential towards a mean value of -65 to -60 mV. Since in animal models, acute vestibular disorders are associated with asymmetrical spontaneous activities of MVN neurons, this study suggests how acetyl-DL-leucine may reduce acute, vestibular-related imbalances in humans.

We investigated the roles of ionotropic glutamate receptor subtypes in the development and recovery of spontaneous nystagmus (SN) after unilateral labyrinthectomy (UL) in guinea pigs. When administered at 3 h after UL, N-methyl-D-aspartate (NMDA) and kainate (KA), which are NMDA and non-NMDA receptor agonists, respectively, increased the frequency of SN. The effect of KA was more potent than that of NMDA. In contrast to these agonists, MK-801 and CNQX decreased the frequency of SN. Although the administration of KA at 48 h after UL increased the frequency of SN, it did not exhibit any effects at 72 h after UL. MK-801 caused a recurrence of SN following administration at 48 and 72 h after UL. Neither NMDA nor CNQX exhibited any effects after administration at 48 or 72 h after UL. A newly synthesized compound, NC-1200, which has inhibitory action on the glutamate response, decreased the frequency of SN in a dose-dependent manner following administration at 3 h after UL, but did not exhibit any effects when administered at 48 and 72 h after UL. From these results, it was found that NMDA and non-NMDA receptors play important roles in the development of SN after UL, and that the NMDA receptor contributes to the development of ocular motor compensation.

We evaluated the effect of steroid hormone on vestibular function in a guinea pig of peripheral vestibular disorder. The right lateral semicircular canal was surgically damaged, and after surgery, animals were treated with 0.1 mg/ml of betamethasone in saline, 1 mg/ml of betamethasone in saline, or saline only, which was administrated directly into the scala tympani by osmotic pump. Rotation tests were performed, and the post-rotatory nystagmus (PRN) ratio (PRN number after counterclockwise rotation/PRN number after clockwise rotation) was calculated. The PRN ratio was recovered to normal at 5 days after treatment in the betamethasone administrated groups, but it did not recover to normal until 14 days after treatment in the saline administrated group. Results indicate that in the vestibular periphery steroid hormones may play an important role in recover of vestibular function.

The way a rabbit moves its eyes in response to a stimulus consisting of two moving random dot patterns largely depends on the relative luminances of the two patterns. Concurrent rotation of the animal enhances the response to the visual pattern that represents the same head movement as the vestibular stimulation. In this paper we investigate the role that the flocculus plays in this behaviour. We injected the non-selective acetylcholine agonist carbachol into the flocculus. These injections are known to increase the gain of the optokinetic reflex, but have a smaller effect on the vestibulo-ocular reflex. We investigated the effect on the oculomotor response to (vestibulo-) transparent stimuli, where one pattern oscillated sinusoidally and the other pattern was stable with respect to the head. We found that the injections caused a higher response gain at a lower luminance of the oscillating pattern. Furthermore the influence of concurrent vestibular stimulation decreased. These findings agree with a role of the flocculus that is downstream of the visual normalisation, but upstream of the visual-vestibular interaction.

In the past year significant advances have been made in our understanding of the neurochemistry and neuropharmacology of the peripheral and central vestibular systems. The recognition of the central importance of excitatory amino acids and their receptors at the level of the hair cells, vestibular nerve and vestibular nucleus has progressed further, and the role of nitric oxide in relation to activation of the N-methyl-D-aspartate receptor subtype is becoming increasingly clear. Increasing evidence suggests that excessive N-methyl-D-aspartate receptor activation and nitric oxide production after exposure to aminoglycoside antibiotics is a critical part of hair cell death, and new pharmacological strategies for preventing aminoglycoside ototoxicity are emerging as a result. Conversely, the use of aminoglycosides to lesion the peripheral vestibular system in the treatment of Meniere's disease has been studied intensively. In the vestibular nucleus, new studies suggest the importance of opioid, nociceptin and glucocorticoid receptors in the control of vestibular reflex function. Finally, the mechanisms of action and optimal use of antihistamines in the treatment of vestibular disorders has also received a great deal of attention.

Avian auditory and vestibular hair cells regenerate after damage by ototoxic drugs, but until recently there was little evidence that regenerated vestibular hair cells function normally. In an earlier study we showed that the vestibuloocular reflex (VOR) is eliminated with aminoglycoside antibiotic treatment and recovers as hair cells regenerate. The VOR, which stabilizes the eye in the head, is an open-loop system that is thought to depend largely on regularly firing afferents. Recovery of the VOR is highly correlated with the regeneration of type I hair cells. In contrast, the vestibulocolic reflex (VCR), which stabilizes the head in space, is a closed-loop, negative-feedback system that seems to depend more on irregularly firing afferent input and is thought to be subserved by different circuitry than the VOR. We examined whether this different reflex also of vestibular origin would show similar recovery after hair cell regeneration. Lesions of the vestibular hair cells of 10-day-old chicks were created by a 5-day course of streptomycin sulfate. One day after completion of streptomycin treatment there was no measurable VCR gain, and total hair cell density was approximately 35% of that in untreated, age-matched controls. At 2 wk postlesion there was significant recovery of the VCR; at this time two subjects showed VCR gains within the range of control chicks. At 3 wk postlesion all subjects showed VCR gains and phase shifts within the normal range. These data show that the VCR recovers before the VOR. Unlike VOR gain, recovering VCR gain correlates equally well with the density of regenerating type I and type II vestibular hair cells, except at high frequencies. Several factors other than hair cell regeneration, such as length of stereocilia, reafferentation of hair cells, and compensation involving central neural pathways, may be involved in behavioral recovery. Our data suggest that one or more of these factors differentially affect the recovery of these two vestibular reflexes.

Microtubule associated proteins (MAPs) are essential cytoskeletal proteins in developing neurons. The present study was undertaken to analyze the expression of MAP2 and its isoforms (a,b,c) during the embryonal and early post-hatching development of chicken cochleovestibular ganglion (CVG) neurons. Moreover, we have investigated MAP2 expression in primary cultures of CVG neurons, and whether it is regulated by neurotrophin-3 (NT3). The expression of MAP2 immunoreactivity (IR) was studied using both Western blot and immunohistochemistry on tissue sections and primary cultures. In vivo MAP2c was expressed from incubation day 4 (E4) to E10, and MAP2b was found in all embryonal stages studied and at post-hatching day 10 (P10), whereas MAP2a was restricted to the post-hatching periods. The cellular localization of IR was in the neuronal perikarya and their peripheral processes (dendrites) but not in axons. Primary cultures matched the in vivo pattern of MAP2 expression, and IR was localized in neuronal cell bodies and the initial segment of the neuronal processes. Exogenous NT3 regulated the expression of MAP2 isoforms in a dose dependent manner. At the survival dose of 0.5 ng/ml NT3, the main MAP2 expression was MAP2c. Conversely, at the neuritogenic dose of 5 ng/ml NT3 increased MAP2b and MAP2a expression, but not MAP2c. The present results demonstrate that MAP2 isoforms are developmentally regulated, thus suggesting that each isoform is specifically involved in CVG neuron maturation. Furthermore, we provide evidence of MAP2 regulation in culture by the neurotrophic factor NT3.

This study was designed to examine the role of flavin-containing monooxygenase (FMO) on the auditory and vestibular neurotoxicity of 3,3'-iminodipropionitrile (IDPN) using the FMO substrate and competitive inhibitor methimazole (MMI). Specifically, the purpose was to block the FMO-mediated conversion of IDPN to the putative neurotoxic metabolite N-hydroxy3,3'-iminodipropionitrile (HOIDPN). In three separate experiments, adult male Long-Evans hooded rats were administered (ip) saline (vehicle), MMI, IDPN, or HOIDPN individually, or a combination of IDPN and MMI or HOIDPN and MMI. Animals were observed daily for signs of the ECC syndrome (excitation with choreiform and circling movements) for 10 days. One to 2 weeks after exposure, a battery of behavioral tests was used to examine vestibular and auditory function. MMI completely blocked the neurotoxicity associated with a 600 mg/kg dose of IDPN and partially blocked the effects of a 1000 mg/kg dose of IDPN. In contrast, MMI failed to block, and instead increased, the neurotoxicity associated with HOIDPN. These data suggest that FMO-mediated metabolism of IDPN is necessary for the generation of a metabolite responsible for the vestibular and auditory neurotoxicities.

The action of streptomycin sulfate (SM) on the regenerative process of the vestibular nerve and posture recovery was studied, using bull frogs. The vestibular nerve was sectioned in various conditions with intact endorgan or with SM intoxication. When the nerve was sectioned with the hair cells left intact, the nerve regenerated well and body balance recovered to normal. However, when neural regeneration was blocked, recovery was incomplete. SM intoxication resulted in various degrees of hair cell damage. Degree of posture recovery correlated well with the number of hair cells. When the nerve was sectioned after damaging the hair cell, the nerve failed to regenerate and posture recovery was incomplete. These results suggest that the degree of posture recovery depends on hair cell function and neural regeneration. Furthermore, neural regeneration is strongly influenced by hair cell function.

Carboplatin, a second-generation platinum drug used in the treatment of cancer, can damage the hair cells in the vestibular system; however, little is known about the time course of its vestibulotoxic effects. The present study examined the acute vestibulotoxic effects of carboplatin (50 mg/kg) in the chinchilla. The duration of the nystagmus response evoked by cold caloric stimulation was significantly reduced 6 h following carboplatin treatment and showed a maximum, permanent reduction of approximately 50% by 24 h after injection. Light-microscopic observations at 6 h subsequent to injection revealed swollen afferent dendrites beneath type-I hair cells and the appearance of small vacuoles within the type-I hair cells; these changes were most pronounced in the crista ampullaris of the semicircular canals compared to the maculae of the utricle and saccule. Many mitochondria were swollen and partially depleted of their membranous infoldings. The mitochondrial abnormalities tended to be somewhat more severe in the hair cells than in their afferent terminals. The structural abnormalities in the mitochondria were more severe at 24 h following injection resulting in the appearance of larger and more numerous vacuoles in the hair cells. By 3 days after injection, many type-I hair cells were filled with large vacuoles which often caused severe distortion of the nucleus and disruption of the plasma membrane. Small vacuoles were occasionally observed in type-II hair cells, mainly in the crista ampullaris. These results indicate that the vestibulotoxic effects of carboplatin occur quite rapidly and cause significant disruption of the mitochondria in hair cells and their afferent terminals.

Microtubule-associated proteins (MAPs) are essential cytoskeletal components during development for neurogenesis and neuronal plasticity. Inner ear innervation is accomplished by cochleovestibular ganglion (CVG) neurons in a highly specific, well-defined pattern, which is regulated by neurotrophic factors belonging to the neurotrophin family. The inner ear offers a suitable model for studying the expression of MAPs and assessing their role in neurotrophin-induced effects that are required for neuron-target innervation. The present study was undertaken to analyze the expression and localization of MAP5 isoforms during development of CVG neurons in vivo an in vitro; as well as the regulation of MAP5 by neurotrophin-3 (NT3) in cell culture. MAP5 expression in the inner ear of chick embryos and postnatal specimens was monitored using immunoblots and immunohistochemistry on frozen sections. MAP5 was highly expressed during the early stages of CVG development, at embryonic day (E)4, being located in both neuronal perikarya and neurites. Expression was maintained during the neurite outgrowth phase (E6-E12), when strong MAP5 immunostaining was observed at the same cellular locations. MAP5 expression decreased suddenly at E14, after the establishment of specific connections between the CVG neurons and their targets, the sensory epithelium of the inner ear. In cultured CVG neurons addition of NT3 led to increased MAP5 expression and produced neurite outgrowth. Both effects are differentially regulated in parallel by low (0.5 ng/ml) and high (5 ng/ml) NT3 concentrations. Present results suggest that MAP5 may be involved in neurotrophin-induced microtubule bundling during neurite outgrowth of auditory neurons.