Engraftment of Human Stem Cell-Derived Otic Progenitors in the Damaged Cochlea.

Most cases of sensorineural deafness are caused by degeneration of hair cells. Although stem/progenitor cell therapy is becoming a promising treatment strategy in a variety of organ systems, cell engraftment in the adult mammalian cochlea has not yet been demonstrated. In this study, we generated human otic progenitor cells (hOPCs) from induced pluripotent stem cells (iPSCs) in vitro and identified these cells by the expression of known otic markers. We showed successful cell transplantation of iPSC-derived-hOPCs in an in vivo adult guinea pig model of ototoxicity. The delivered hOPCs migrated throughout the cochlea, engrafted in non-sensory regions, and survived up to 4 weeks post-transplantation. Some of the engrafted hOPCs responded to environmental cues within the cochlear sensory epithelium and displayed molecular features of early sensory differentiation. We confirmed these results with hair cell progenitors derived from Atoh1-GFP mice as donor cells. These mouse otic progenitors transplanted using the same in vivo delivery system migrated into damaged cochlear sensory epithelium and adopted a partial sensory cell fate. This is the first report of the survival and differentiation of hOPCs in ototoxic-injured mature cochlear epithelium, and it should stimulate further research into cell-based therapies for treatment of deafness.

Adjustment of the dynamic weight distribution as a sensitive parameter for diagnosis of postural alteration in a rodent model of vestibular deficit.

Vestibular disorders, by inducing significant posturo-locomotor and cognitive disorders, can significantly impair the most basic tasks of everyday life. Their precise diagnosis is essential to implement appropriate therapeutic countermeasures. Monitoring their evolution is also very important to validate or, on the contrary, to adapt the undertaken therapeutic actions. To date, the diagnosis methods of posturo-locomotor impairments are restricted to examinations that most often lack sensitivity and precision. In the present work we studied the alterations of the dynamic weight distribution in a rodent model of sudden and complete unilateral vestibular loss. We used a system of force sensors connected to a data analysis system to quantify in real time and in an automated way the weight bearing of the animal on the ground. We show here that sudden, unilateral, complete and permanent loss of the vestibular inputs causes a severe alteration of the dynamic ground weight distribution of vestibulo lesioned rodents. Characteristics of alterations in the dynamic weight distribution vary over time and follow the sequence of appearance and disappearance of the various symptoms that compose the vestibular syndrome. This study reveals for the first time that dynamic weight bearing is a very sensitive parameter for evaluating posturo-locomotor function impairment. Associated with more classical vestibular examinations, this paradigm can considerably enrich the methods for assessing and monitoring vestibular disorders. Systematic application of this type of evaluation to the dizzy or unstable patient could improve the detection of vestibular deficits and allow predicting better their impact on posture and walk. Thus it could also allow a better follow-up of the therapeutic approaches for rehabilitating gait and balance.

Forward Masking in Cochlear Implant Users: Electrophysiological and Psychophysical Data Using Pulse Train Maskers.

Electrical stimulation of auditory nerve fibers using cochlear implants (CI) shows psychophysical forward masking (pFM) up to several hundreds of milliseconds. By contrast, recovery of electrically evoked compound action potentials (eCAPs) from forward masking (eFM) was shown to be more rapid, with time constants no greater than a few milliseconds. These discrepancies suggested two main contributors to pFM: a rapid-recovery process due to refractory properties of the auditory nerve and a slow-recovery process arising from more central structures. In the present study, we investigate whether the use of different maskers between eCAP and psychophysical measures, specifically single-pulse versus pulse train maskers, may have been a source of confound.In experiment 1, we measured eFM using the following: a single-pulse masker, a 300-ms low-rate pulse train masker (LTM, 250 pps), and a 300-ms high-rate pulse train masker (HTM, 5000 pps). The maskers were presented either at same physical current (Φ) or at same perceptual (Ψ) level corresponding to comfortable loudness. Responses to a single-pulse probe were measured for masker-probe intervals ranging from 1 to 512 ms. Recovery from masking was much slower for pulse trains than for the single-pulse masker. When presented at Φ level, HTM produced more and longer-lasting masking than LTM. However, results were inconsistent when LTM and HTM were compared at Ψ level. In experiment 2, masked detection thresholds of single-pulse probes were measured using the same pulse train masker conditions. In line with our eFM findings, masked thresholds for HTM were higher than those for LTM at Φ level. However, the opposite result was found when the pulse trains were presented at Ψ level.Our results confirm the presence of slow-recovery phenomena at the level of the auditory nerve in CI users, as previously shown in animal studies. Inconsistencies between eFM and pFM results, despite using the same masking conditions, further underline the importance of comparing electrophysiological and psychophysical measures with identical stimulation paradigms.

Effects of Pulse Shape and Polarity on Sensitivity to Cochlear Implant Stimulation: A Chronic Study in Guinea Pigs.

Most cochlear implants (CIs) stimulate the auditory nerve with trains of symmetric biphasic pulses consisting of two phases of opposite polarity. Animal and human studies have shown that both polarities can elicit neural responses. In human CI listeners, studies have shown that at suprathreshold levels, the anodic phase is more effective than the cathodic phase. In contrast, animal studies usually show the opposite trend. Although the reason for this discrepancy remains unclear, computational modelling results have proposed that the degeneration of the peripheral processes of the neurons could lead to a higher efficiency of anodic stimulation. We tested this hypothesis in ten guinea pigs who were deafened with an injection of sysomycin and implanted with a single ball electrode inserted in the first turn of the cochlea. Animals were tested at regular intervals between 1 week after deafening and up to 1 year for some of them. Our hypothesis was that if the effect of polarity is determined by the presence or absence of peripheral processes, the difference in polarity efficiency should change over time because of a progressive neural degeneration. Stimuli consisted of charge-balanced symmetric and asymmetric pulses allowing us to observe the response to each polarity individually. For all stimuli, the inferior colliculus evoked potential was measured. Results show that the cathodic phase was more effective than the anodic phase and that this remained so even several months after deafening. This suggests that neural degeneration cannot entirely account for the higher efficiency of anodic stimulation observed in human CI listeners.

Erratum to: Effects of Pulse Shape and Polarity on Sensitivity to Cochlear Implant Stimulation: A Chronic Study in Guinea Pigs.

In the original version of the chapter, the labels on the x-axis of Figure 2, panels A and B were wrong. This incorrect figure has been replaced with the below figure.

KCNK5 channels mostly expressed in cochlear outer sulcus cells are indispensable for hearing.

In the cochlea, K(+) is essential for mechano-electrical transduction. Here, we explore cochlear structure and function in mice lacking K(+) channels of the two-pore domain family. A profound deafness associated with a decrease in endocochlear potential is found in adult Kcnk5(-/-) mice. Hearing occurs around postnatal day 19 (P19), and completely disappears 2 days later. At P19, Kcnk5(-/-) mice have a normal endolymphatic [K(+)] but a partly lowered endocochlear potential. Using Lac-Z as a gene reporter, KCNK5 is mainly found in outer sulcus Claudius', Boettcher's and root cells. Low levels of expression are also seen in the spiral ganglion, Reissner's membrane and stria vascularis. Essential channels (KCNJ10 and KCNQ1) contributing to K(+) secretion in stria vascularis have normal expression in Kcnk5(-/-) mice. Thus, KCNK5 channels are indispensable for the maintenance of hearing. Among several plausible mechanisms, we emphasize their role in K(+) recycling along the outer sulcus lateral route.

Temporal-envelope reconstruction for hearing-impaired listeners.

Recent studies suggest that normal-hearing listeners maintain robust speech intelligibility despite severe degradations of amplitude-modulation (AM) cues, by using temporal-envelope information recovered from broadband frequency-modulation (FM) speech cues at the output of cochlear filters. This study aimed to assess whether cochlear damage affects this capacity to reconstruct temporal-envelope information from FM. This was achieved by measuring the ability of 40 normal-hearing listeners and 41 listeners with mild-to-moderate hearing loss to identify syllables processed to degrade AM cues while leaving FM cues intact within three broad frequency bands spanning the range 65-3,645 Hz. Stimuli were presented at 65 dB SPL for both normal-hearing listeners and hearing-impaired listeners. They were presented as such or amplified using a modified half-gain rule for hearing-impaired listeners. Hearing-impaired listeners showed significantly poorer identification scores than normal-hearing listeners at both presentation levels. However, the deficit shown by hearing-impaired listeners for amplified stimuli was relatively modest. Overall, hearing-impaired data and the results of a simulation study were consistent with a poorer-than-normal ability to reconstruct temporal-envelope information resulting from a broadening of cochlear filters by a factor ranging from 2 to 4. These results suggest that mild-to-moderate cochlear hearing loss has only a modest detrimental effect on peripheral, temporal-envelope reconstruction mechanisms.

Long-term tinnitus suppression with linear octave frequency transposition hearing AIDS.

Over the last three years of hearing aid dispensing, it was observed that among 74 subjects fitted with a linear octave frequency transposition (LOFT) hearing aid, 60 reported partial or complete tinnitus suppression during day and night, an effect still lasting after several months or years of daily use. We report in more details on 38 subjects from whom we obtained quantified measures of tinnitus suppression through visual analog scaling and several additional psychoacoustic and audiometric measures. The long-term suppression seems independent of subject age, and of duration and subjective localization of tinnitus. A small but significant correlation was found with audiogram losses but not with high frequency loss slope. Long-term tinnitus suppression was observed for different etiologies, but with a low success rate for sudden deafness. It should be noted that a majority of subjects (23) had a history of noise exposure. Tinnitus suppression started after a few days of LOFT hearing aid use and reached a maximum after a few weeks of daily use. For nine subjects different amounts of frequency shifting were tried and found more or less successful for long-term tinnitus suppression, no correlation was found with tinnitus pitch. When the use of the LOFT hearing aid was stopped tinnitus reappeared within a day, and after re-using the LOFT aid it disappeared again within a day. For about one third of the 38 subjects a classical amplification or a non linear frequency compression aid was also tried, and no such tinnitus suppression was observed. Besides improvements in audiometric sensitivity to high frequencies and in speech discrimination scores, LOFT can be considered as a remarkable opportunity to suppress tinnitus over a long time scale. From a pathophysiological viewpoint these observations seem to fit with a possible re-attribution of activity to previously deprived cerebral areas corresponding to high frequency coding.

Age-related impairment of ultrasonic vocalization in Tau.P301L mice: possible implication for progressive language disorders.

BACKGROUND: Tauopathies, including Alzheimer's Disease, are the most frequent neurodegenerative diseases in elderly people and cause various cognitive, behavioural and motor defects, but also progressive language disorders. For communication and social interactions, mice produce ultrasonic vocalization (USV) via expiratory airflow through the larynx. We examined USV of Tau.P301L mice, a mouse model for tauopathy expressing human mutant tau protein and developing cognitive, motor and upper airway defects. METHODOLOGY/PRINCIPAL FINDINGS: At age 4-5 months, Tau.P301L mice had normal USV, normal expiratory airflow and no brainstem tauopathy. At age 8-10 months, Tau.P301L mice presented impaired USV, reduced expiratory airflow and severe tauopathy in the periaqueductal gray, Kolliker-Fuse and retroambiguus nuclei. Tauopathy in these nuclei that control upper airway function and vocalization correlates well with the USV impairment of old Tau.P301L mice. CONCLUSIONS: In a mouse model for tauopathy, we report for the first time an age-related impairment of USV that correlates with tauopathy in midbrain and brainstem areas controlling vocalization. The vocalization disorder of old Tau.P301L mice could be, at least in part, reminiscent of language disorders of elderly suffering tauopathy.

Ensemble spontaneous activity alterations detected by CISA approach.

In this paper, we propose a method for detecting alterations in the Ensemble Spontaneous Activity (ESA), a random signal representing the composite spontaneous contribution of the auditory nerve recorded on the round window. The proposed method is based on shape analysis of the ESA amplitude histogram. For this task, we use a recent approach, the Corrected Integral Shape Averaging (CISA). Using this approach, a shape clustering algorithm is proposed to classify healthy and pathological ESA signals generated by a recent ESA model. This model allows a precise simulation of neural mechanisms occurring in the auditory nerve. The obtained results demonstrate that this shape analysis is very sensitive for detecting a small number of fibers with correlated firing, supposed to occur during a particular type of tinnitus. In comparison, the classical spectral index fails in this detection.

Experimental estrogen-induced hyperprolactinemia results in bone-related hearing loss in the guinea pig.

Our group (Horner KC, Guieu R, Magnan J, Chays A, Cazals Y. Neuropsychopharmacology 26: 135-138, 2002) has earlier described hyperprolactinemia in some patients presenting inner ear dysfunction. However, in that study, it was not possible to determine whether hyperprolactinemia was a cause or an effect of the symptoms. To investigate the effect of hyperprolactinemia on inner ear function, we first developed a model of hyperprolactinemia in estrogen-primed Fischer 344 rats and then performed functional studies on pigmented guinea pigs. Hyperprolactinemia induced, after 2 mo, a hearing loss of approximately 30-40 dB across all frequencies, as indicated by the compound action potential audiogram. During the 3rd mo, the hearing loss continued to deteriorate. The threshold shifts were more substantial in males than in females. Observations under a dissection microscope revealed bone dysmorphology of the bulla and the cochlea. Light microscopy observations of cryostat sections confirmed bone-related pathology of the bony cochlear bulla and the cochlear wall and revealed morphopathology of the stria vascularis and spiral ligament. Scanning electron microscopy revealed loss of hair cells and stereocilia damage, in particular in the upper three cochlear turns and the two outermost hair cell rows. The data provide the first evidence of otic capsule and hair cell pathology associated with estrogen-induced prolonged hyperprolactinemia and suggest that conditions such as pregnancy, anti-psychotic drug treatment, aging, and/or stress might lead to similar ear dysfunctions.

Loudness reduction induced by a contralateral tone.

The induced reduction in the loudness (ILR) of a weaker tone caused by a preceding stronger tone was measured with both tones in the same ear (ipsilateral ILR) and also in opposite ears (contralateral ILR). The two tones were always equal in duration and were presented repeatedly over several minutes. When the tone duration was 200 ms, for 24 listeners the loudness reduction averaged 11 dB under ipsilateral ILR and 6 dB under contralateral ILR. When the duration was 5 ms, ILR was 8 dB whether ipsilateral or contralateral. For each duration, ipsilateral and contralateral ILR were strongly correlated (r around 0.80).

Hyperbaric oxygenation with corticoid in experimental acoustic trauma.

Among possible therapies after acute acoustic trauma, hyperbaric oxygenation (HBO) combined with corticoid was found effective in several animal studies. Such evidence was obtained for moderate 20-25 dB losses. The aim of this study was to further assess this therapy for noise-induced hearing losses greater than previously examined. Sixty-five ears from thirty-six adult guinea pigs were used. Acoustically evoked responses from intracranial electrodes chronically implanted bilaterally into the ventral cochlear nucleus were used to assess acoustic sensitivity alterations. Trauma sound was a third-octave noise-band around 8 kHz presented bilaterally at 115 dB SPL for 45 min. One control group received no treatment, one group was treated with HBO only and another with corticoid only both starting within one day post-trauma, two groups were treated with both HBO and corticoid starting for one group within one day post-trauma, and for the second group at 6 days post-trauma. Acoustic thresholds were measured between the 6th and the 16th days after acoustic trauma. Animals treated with HBO alone or corticoid alone did not differ from controls. Combined HBO and corticoid therapy provided significant protection from noise-induced loss of auditory thresholds, especially when started one day post-exposure. Hearing loss reduction induced by HBO combined with corticoid was of similar magnitude (about 10-15 dB) as in previous studies although the induced hearing loss was considerably greater (about 40 dB instead of 20-25 dB).

Enhanced cochlear acoustic sensitivity and susceptibility to endotoxin are induced by adrenalectomy and reversed by corticosterone supplementation in rat.

Glucocorticoid receptors are widely distributed in the cochlea but their role remains poorly known. Previous studies provided contradictory reports on a possible cochlear acoustic hypersensitivity induced by adrenal insufficiency, while several experiments agree on a significant action of glucocorticoid receptors in adverse conditions such as acoustic trauma and restraint stress. The present experiments confirmed a cochlear acoustic hypersensitivity induced by adrenalectomy and reversed by corticosterone supplementation. These observations point to a significant role of corticosteroids in basal cochlear functioning. The glucocorticoids are known to be essential for limiting and resolving inflammatory processes. The endotoxin Escherichia coli lipopolysaccharide is widely used to induce inflammatory reactions. However, in various organs several toxic processes of this endotoxin are not influenced by glucocorticoids. From previous experiments on the cochlea there is no evidence that glucocorticoids are an essential factor against endotoxin cochlear toxicity. In the present experiments it was found that adrenalectomy greatly increased the cochlear susceptibility to endotoxin; the effect was reversed by providing corticosterone supplementation. This shows the essential role of corticosterone in this cochlear inflammation model. In previous studies local administration (at the cochlear base) of endotoxin was used and losses of cochlear acoustic sensitivity were found predominantly at high frequencies; in contrast, the systemic injection used in this study produced a cochlear loss of acoustic sensitivity at all frequencies, indicating a uniform cochlear sensitivity to the toxic effects of endotoxin.

Stress hormones in Ménière's disease and acoustic neuroma.

Stress has been postulated to trigger or contribute to inner ear pathologies but there is little objective evidence. We investigated stress hormones in Ménière's patients and patients with acoustic neuroma. Data were compared with those from a control group of patients with facial spasm. We assayed classic stress hormones including adrenocorticotropic hormone, cortisol, growth hormone and prolactin. We found a strong positive correlation between cortisol and adrenocorticotropic hormone in Ménière patients and patients with acoustic neuroma but no correlation in patients with facial spasm. The data also revealed in female patients with Ménière's disease or with acoustic neuroma an unexpected significant positive correlation between cortisol and prolactin. The data showed the expected negative correlation or no correlation between cortisol and prolactin associated with males and females in the other patient groups. Both cortisol and prolactin increases are known to represent alternative strategies to cope with stress, and our data point to prolactin being possibly more dominant in Ménière's disease and cortisol in acoustic neuroma. These data provide further evidence for modification of different stress hormones in audiovestibular pathologies, which might provide a valuable diagnostic or prognostic tool in the future.

Hexachlorobenzene, a dioxin-like compound, disrupts auditory function in rat.

Hexachlorobenzene (HCB) is a dioxin-like compound widely distributed in the environment. In this study, we investigated the effects of HCB on the cochlea. Conscious free-moving rats were given HCB per os daily for 4 weeks at doses of 0.16, 4 or 16 mg/kg in olive oil, whereas the control group received olive oil only. The effects of HCB were evaluated at various time intervals, by measuring auditory nerve acoustic thresholds and plasma thyroid hormone concentration by radioimmunoassay. Histological evaluation involved surface preparation and scanning electron microscopy observations of cochlear hair cells. At a dose of 0.16 mg/kg, HCB induced no loss of acoustic sensitivity, whereas at 4 mg/kg, it induced cochlear sensitivity deficits at the mid-frequencies (2-16 kHz) with complete recovery once treatment was stopped. At a dose of 16 mg/kg, permanent threshold shifts were observed at all frequencies tested (from 1 to 32 kHz). Morphological studies showed no cochlear hair cell loss or alteration of stereocilia. HCB treatment reduced circulating thyroxine concentrations. Thyroidectomy had no effect on cochlear sensitivity in control animals. Thus, HCB is a potent oto-toxicant, and its ototoxicity may be independent of its thyroidal effects.

Does repeated hyperbaric exposure to 4 atmosphere absolute cause hearing impairment? Study in Guinea pigs and clinical incidences.

HYPOTHESIS: Direct pressure applied on the inner ear cannot induce hearing loss. BACKGROUND: Three possible causes have been described in the literature for inner ear permanent lesions during scuba diving: pressure imbalance between the middle ear and the external ear, appearance of microbubbles in the internal ear, and direct effect of pressure on the inner ear. We seek to determine whether this last factor can be involved. METHODS: We submitted two groups of guinea pigs previously implanted with an electrode in the round window to a protocol of air diving in a hyperbaric chamber. Eardrums of animals in one of the two groups had been perforated beforehand. Twenty dives were practiced over 4 weeks. We chose dive parameters consistent with common sport diving: maximal pressure of 4 atmosphere absolute and duration of 30 minutes. Auditory threshold and cochlear spontaneous activity were recorded at regular intervals. Furthermore, we recorded spontaneous cochlear activity in Heliox 400-m and 600-m dives to determine whether our conclusions hold for "extreme" diving. RESULTS: In the group with perforated eardrums, no variation of those parameters were recorded, even in extreme diving. Important variations were noticed in the other group. CONCLUSIONS: Pressure applied directly on the inner ear during diving does not disturb cochlear activity.

Similar half-octave TTS protection of the cochlea by xylazine/ketamine or sympathectomy.

Cochlear efferents, sympathetic control and stress conditions have been shown to influence sound-induced hearing loss. These factors are also known to be modified by sedation/anesthesia. We tested here the effect of sedation/anesthesia on temporary threshold shift (TTS) compared to that in the same awake animals. The effect of sympathectomy was also tested. We employed awake guinea pigs with a chronically implanted electrode on the round window of each of the cochleae. Each ear was tested for its sensitivity to TTS induced by a 1 min or a 10 min exposure to an 8 kHz pure tone at 96 dB sound pressure level. After an intramuscular injection of xylazine or ketamine together with xylazine, TTS at half-octave frequencies was reduced compared to that in awake animals. The second half-octave frequencies were less affected. This specific pattern of protection was also observed here after surgical ablation of a superior cervical ganglion. The data lead to the speculation that protection from TTS under sedation/anesthesia might be due to diminished sympathetic influence. Xylazine is a pre-synaptic alpha2-adrenoreceptor agonist which blocks noradrenaline release from the sympathetic system. Ketamine is a N-methyl-D-aspartic acid receptor antagonist which could reduce glutamate excitotoxicity as well as reduce sympathetic activity.

Prolactinoma in some Ménière's patients--is stress involved?

Dizziness is a common complaint in primary care clinics and can enter the diagnostic profile of different pathologies spanning from psychiatric problems to vestibular dysfunction. Episodes of vertigo in Ménière's patients are often reported to be triggered by stress but no physiological data are available to account for the subjective link. The study involved 42 Ménière's patients hospitalized for neurectomy of the vestibular nerve for relief of incapacitating vertigo. In addition 18 patients with neurinoma of the vestibular nerve and 12 patients with facial spasm, who underwent surgery, served as controls. A blood sample was taken on the day of surgery in order to determine the level of battery of different stress hormones. The most striking observation was the presence of hyperprolactinemia (above 20 microg/l) in 14 Ménière's patients. The presence of prolactinoma was confirmed by MRI in six cases out of six investigated and the others have not yet been followed up in this retrospective study. These observations are clearly indicative for systematic determination of prolactin levels before opting for surgery in Ménière's patients.