Hidden hearing loss is associated with loss of ribbon synapses of cochlea inner hair cells

The present study aimed to observe the changes in the cochlea ribbon synapses after repeated exposure to moderate-to-high intensity noise. Guinea pigs received 95 dB SPL white noise exposure 4 h a day for consecutive 7 days (we regarded it a medium-term and moderate-intensity noise, or MTMI noise). Animals were divided into four groups: Control, 1DPN (1-day post noise), 1WPN (1-week post noise), and 1MPN (1-month post noise). Auditory function analysis by auditory brainstem response (ABR) and compound action potential (CAP) recordings, as well as ribbon synapse morphological analyses by immunohistochemistry (Ctbp2 and PSD95 staining) were performed 1 day, 1 week, and 1 month after noise exposure. After MTMI noise exposure, the amplitudes of ABR I and III waves were suppressed. The CAP threshold was elevated, and CAP amplitude was reduced in the 1DPN group. No apparent changes in hair cell shape, arrangement, or number were observed, but the number of ribbon synapse was reduced. The 1WPN and 1MPN groups showed that part of ABR and CAP changes recovered, as well as the synapse number. The defects in cochlea auditory function and synapse changes were observed mainly in the high-frequency region. Together, repeated exposure in MTMI noise can cause hidden hearing loss (HHL), which is partially reversible after leaving the noise environment; and MTMI noise-induced HHL is associated with inner hair cell ribbon synapses.     

Chemical neuroprotection in the cochlea: the modulation of dopamine release from lateral olivocochlear efferents

The prevalence of sensorineural hearing loss is increasing worldwide, mainly due to ageing, increased noise exposure and cardiovascular risk factors. Several papers dealt with the mechanisms underlying the primary causes of impaired hearing and eventual deafness, including the damage and loss of auditory hair cells; however, very little is known about the protective mechanisms that exist for hearing. Several recent investigations have implicated dopamine (DA) in a neuroprotective circuit for the cochlea. The lateral olivocochlear (LOC) efferents provide axonal innervation of the inner hair cell afferent synapses and release DA and other substances in response to different stimuli. Under ischemic conditions or during noise exposure, DA has been proven to play a neuroprotective role against glutamate excitotoxicity. This review summarises what is currently known about the modulation of DA release in the cochlea, using primarily in vitro experimental data. Based on recent knowledge, there could be two functional subgroups within the LOC fibres, i.e., the DA- and GABA-containing projections. In this review, we attempt to show the neurochemical interactions between these two subsystems. Other aspects of cochlear neurotransmission are also discussed to provide a complete picture of cochlear dopaminergic function in physiological and pathophysiological cases with particular reference to excitotoxicity.     

Ectopic expression of ROR1 prevents cochlear hair cell loss in guinea pigs with noise‐induced hearing loss

Noise‐induced hearing loss (NIHL) is one of the most frequent disabilities in industrialized countries. Evidence shows that hair cell loss in the auditory end organ is responsible for the majority of various ear pathological conditions. The functional roles of the receptor tyrosine kinase ROR1 have been underscored in various tumours. In this study, we evaluated the ability of ROR1 to influence cochlear hair cell loss of guinea pigs with NIHL. The NIHL model was developed in guinea pigs, with subsequent measurement of the auditory brainstem response (ABR). Gain‐of‐function experiments were employed to explore the role of ROR1 in NIHL. The interaction between ROR1 and Wnt5a and their functions in the cochlear hair cell loss were further analysed in response to alteration of ROR1 and Wnt5a. Guinea pigs with NIHL demonstrated elevated ABR threshold and down‐regulated ROR1, Wnt5a and NF‐κB p65. The up‐regulation of ROR1 was shown to decrease the cochlear hair cell loss and the expression of pro‐apoptotic gene (Bax, p53) in guinea pig cochlea, but promoted the expression of anti‐apoptotic gene (Bcl‐2) and the fluorescence intensity of cleaved‐caspase‐3. ROR1 interacted with Wnt5a to activate the NF‐κB signalling pathway through inducing phosphorylation and translocation of p65. Furthermore, Wnt5a overexpression decreased the cochlear hair cell loss. Collectively, this study suggested the protection of overexpression of ROR1 against cochlear hair cell loss in guinea pigs with NIHL via the Wnt5a‐dependent NF‐κB signalling pathway.     

变压器噪声暴露对SD大鼠听力及应激状态的影响研究

目的:探究短时间内低声级强度低频的变压器噪声暴露对SD大鼠听力及应激状态方面的影响。方法:选取90只SPF级健康无听力障碍的(雌雄各半)SD大鼠作为实验对象,随机分为实验A、B组和对照C组,A、B组分别给予声级上限为65 dB SPL、60 dB SPL(频谱范围:100~800 Hz)的变压器噪声,噪声暴露时程为8周,每日噪声给予时间为22点至次日8点,C组在相同条件下饲养,不给予噪声暴露。噪声暴露结束后,通过DPOAE(畸变耳声发射)、ABR(听性脑干反应)检测、耳蜗铺片及毛细胞计数对SD大鼠听力学状况进行评估;通过血清中促肾上腺皮质激素(ACTH)、血清皮质醇(CORT)对SD大鼠的应激状态进行评估。结果:在变压器噪声暴露的8周内,各组大鼠生长状况良好,体重均呈正常生理性增长,组间无明显差异(P0.05);在变压器噪声暴露8周后,对A、B、C三组大鼠的听力学指标进行两两比较,组间均无明显差异(P0.05),对大鼠血清中促肾上腺皮质激素(ACTH)、血清皮质醇(CORT)的含量进行三组间比较,组间差异均无统计学意义(P0.05)。结论:连续暴露于声压级上限65/60 dB SPL,频谱范围为100~800 Hz的变压器噪声下8周(10小时/天)对SD大鼠听力未产生明显影响,未引发SD大鼠应激状态。     

Prestin Regulation and Function in Residual Outer Hair Cells after Noise-Induced Hearing Loss

The outer hair cell (OHC) motor protein prestin is necessary for electromotility, which drives cochlear amplification and produces exquisitely sharp frequency tuning. Tecta^C1509G transgenic mice have hearing loss, and surprisingly have increased OHC prestin levels. We hypothesized, therefore, that prestin up-regulation may represent a generalized response to compensate for a state of hearing loss. In the present study, we sought to determine the effects of noise-induced hearing loss on prestin expression. After noise exposure, we performed cytocochleograms and observed OHC loss only in the basal region of the cochlea. Next, we patch clamped OHCs from the apical turn (9–12 kHz region), where no OHCs were lost, in noise-exposed and age-matched control mice. The non-linear capacitance was significantly higher in noise-exposed mice, consistent with higher functional prestin levels. We then measured prestin protein and mRNA levels in whole-cochlea specimens. Both Western blot and qPCR studies demonstrated increased prestin expression after noise exposure. Finally, we examined the effect of the prestin increase in vivo following noise damage. Immediately after noise exposure, ABR and DPOAE thresholds were elevated by 30–40 dB. While most of the temporary threshold shifts recovered within 3 days, there were additional improvements over the next month. However, DPOAE magnitudes, basilar membrane vibration, and CAP tuning curve measurements from the 9–12 kHz cochlear region demonstrated no differences between noise-exposed mice and control mice. Taken together, these data indicate that prestin is up-regulated by 32–58% in residual OHCs after noise exposure and that the prestin is functional. These findings are consistent with the notion that prestin increases in an attempt to partially compensate for reduced force production because of missing OHCs. However, in regions where there is no OHC loss, the cochlea is able to compensate for the excess prestin in order to maintain stable auditory thresholds and frequency discrimination.     

噪声所致听力损失现象的物种差异及可能生理机制研究进展

噪声广泛存在于人和动物的生活环境中,从无脊椎动物到哺乳动物乃至人类,都会受到噪声的负面影响.强烈的噪声会损伤听觉系统的结构和功能,引起噪声性听力损失(noise-induced hearing loss,NIHL).本文对噪声性听力损失的类型、影响因素、噪声所致不同程度听力损失形成的可能机制进行了总结,发现NIHL主要与突触结构肿胀、谷氨酸引起的可逆兴奋性中毒以及活性氧引起的氧化应激、细胞凋亡、带状体损伤、α激动型鸟嘌呤核苷酸结合蛋白(guanine nucleotide binding protein alpha stimulating,GNAS)基因的mRNA及其上游lncRNA Sept7的表达量上调等因素有关.比较噪声暴露后不同物种听力损失情况的差异,发现鱼类和鸟类由于具有毛细胞再生能力而能够较快从听力损伤中恢复,啮齿类较容易受到噪声影响,而回声定位鲸类噪声暴露后的暂时性听觉阈移较小,非常有趣的是回声定位蝙蝠在噪声高强度暴露后未表现出暂时性听觉阈移的现象.上述结论提示,对不同物种的比较生理研究可深入揭示NIHL机制,并为听力保护以及噪声所致的听力损伤后修复等提供理论参考.     

Therapeutic effects of orally administrated antioxidant drugs on acute noise-induced hearing loss

Abstract

Objective. The objective of this study was to investigate the dose-dependent therapeutic effect of the orally administrated antioxidant drugs [4-hydroxy alpha-phenyl-tert-butylnitrone (4-OHPBN) and N-acetyl-L-cysteine (NAC)] on acute noise-induced hearing loss because oral administration is the most commonly used method of drug administration due to its convenience, safety, and economical efficiency. Methods. Thirty chinchilla were exposed to a 105 dB octave band noise centered at 4 kHz for 6 h and randomly assigned to a control group (saline only) and three experimental groups [4-OHPBN (10 mg/kg) plus NAC (20 mg/kg), 4-OHPBN (20 mg/kg) plus NAC (50 mg/kg), and 4-OHPBN (50 mg/kg) plus NAC (100 mg/kg)]. The drugs were orally administrated beginning 4 h after noise exposure and then administered twice daily for the next 2 days. Permanent auditory brainstem response threshold shifts, distortion product otoacoustic emission threshold shifts, and the percentage of missing outer hair cell were determined. Results. The oral administration significantly reduced permanent hearing threshold shift, distortion product otoacoustic emission threshold shift, and the percentage of missing outer hair cell in a dose-dependent manner. Discussion. This result demonstrates that orally administered drugs can treat acute noise-induced hearing loss in a dose-dependent manner. This suggests that oral administration was effective in treating acute noise-induced hearing loss as in intraperitoneal administration.     

Reduction in Noise-Induced Functional Loss of the Cochleae in Mice with Pre-Existing Cochlear Dysfunction Due to Genetic Interference of Prestin

Various cochlear pathologies, such as acoustic trauma, ototoxicity and age-related degeneration, cause hearing loss. These pre-existing hearing losses can alter cochlear responses to subsequent acoustic overstimulation. So far, the knowledge on the impacts of pre-existing hearing loss caused by genetic alteration of cochlear genes is limited. Prestin is the motor protein expressed exclusively in outer hair cells in the mammalian cochlea. This motor protein contributes to outer hair cell motility. At present, it is not clear how the interference of prestin function affects cochlear responses to acoustic overstimulation. To address this question, a genetic model of prestin dysfunction in mice was created by inserting an internal ribosome entry site (IRES)-CreER^T2-FRT-Neo-FRT cassette into the prestin locus after the stop codon. Homozygous mice exhibit a threshold elevation of auditory brainstem responses with large individual variation. These mice also display a threshold elevation and a shift of the input/output function of the distortion product otoacoustic emission, suggesting a reduction in outer hair cell function. The disruption of prestin function reduces the threshold shifts caused by exposure to a loud noise at 120 dB (sound pressure level) for 1 h. This reduction is positively correlated with the level of pre-noise cochlear dysfunction and is accompanied by a reduced change in Cdh1 expression, suggesting a reduction in molecular responses to the acoustic overstimulation. Together, these results suggest that prestin interference reduces cochlear stress responses to acoustic overstimulation.     

Gene miles-apart is required for formation of otic vesicle and hair cells in zebrafish

Hearing loss is a serious burden to physical and mental health worldwide. Aberrant development and damage of hearing organs are recognized as the causes of hearing loss, the molecular mechanisms underlining these pathological processes remain elusive. Investigation of new molecular mechanisms involved in proliferation, differentiation, migration and maintenance of neuromast primordium and hair cells will contribute to better understanding of hearing loss pathology. This knowledge will enable the development of protective agents and mechanism study of drug ototoxicity. In this study, we demonstrate that the zebrafish gene miles-apart, a homolog of sphingosine-1-phosphate receptor 2 (s1pr2) in mammals, has an important role in the development of otic vesicle, neuromasts and survival of hair cells. Whole-mount in situ hybridization of embryos showed that miles-apart expression occurred mainly in the encephalic region and the somites at 24 h.p.f. (hour post fertilization), in the midbrain/hindbrain boundary, the brainstem and the pre-neuromast of lateral line at 48 h.p.f. in a strict spatiotemporal regulation. Both up- and downregulation of miles-apart led to abnormal otoliths and semicircular canals, excess or few hair cells and neuromasts, and their disarranged depositions in the lateral lines. Miles-apart (Mil) dysregulation also caused abnormal expression of hearing-associated genes, including hmx2, fgf3, fgf8a, foxi1, otop1, pax2.1 and tmieb during zebrafish organogenesis. Moreover, in larvae miles-apart gene knockdown significantly upregulated proapoptotic gene zBax2 and downregulated prosurvival gene zMcl1b; in contrast, the level of zBax2 was decreased and of zMcl1b enhanced by miles-apart overexpression. Collectively, Mil activity is linked to organization and number decision of hair cells within a neuromast, also to deposition of neuromasts and formation of otic vesicle during zebrafish organogenesis. At the larva stage, Mil as an upstream regulator of bcl-2 gene family has a role in protection of hair cells against apoptosis by promoting expression of prosurvival gene zMcl1b and suppressing proapoptotic gene zBax2.     

A combination of cilostazol and Ginkgo biloba extract protects against cisplatin-induced Cochleo-vestibular dysfunction by inhibiting the mitochondrial apoptotic and ERK pathways

Cisplatin (cis-diammine-dichloroplatinum; CDDP) is an anticancer drug that induces significant hearing loss and balance dysfunction as side effects. Cilostazol (CS, 6-[4-(1-cyclohexyl-1H-tetrazol-5-yl) butoxy]-3, 4-dihydro-2-(1H)-quinolinone) has neuroprotective and antioxidant effects, whereas Ginkgo biloba extract (GbE) has preventive effects on CDDP-induced hearing loss in rats, and GbE enhances the antiatherogenic effect of CS by inhibiting the generation of reactive oxygen species (ROS). The purpose of this study was to investigate the effects of renexin (RXN), which contains GbE and CS, against CDDP-induced cochleo-vestibular dysfunction in rats and to elucidate the mechanism underlying the protective effects of RXN on auditory cells. Rats intraperitoneally injected with CDDP exhibited an increase in hearing threshold and vestibular dysfunction, which agreed with hair cell damage in the Organ of Corti and otoliths. However, these impairments were significantly prevented in a dose-dependent manner by pre- and co-treatment with RXN, and these preventive effects in RXN-treated rats were more prominent than those in GbE-treated rats. In a CDDP pharmacokinetic study, platinum concentration was very similar between CDDP-only treated and RXN+CDDP cotreated rats. RXN markedly attenuated CDDP-induced intracellular ROS and significantly reduced CDDP-activated expression of p-extracellular regulated kinase (ERK), BAX, cytochrome c, cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase, but increased BCL-XL expression. These results show that RXN may have a synergistic effect by strongly protecting hearing and vestibular dysfunction induced by CDDP by inhibiting ROS production, mitochondrial pathways and the ERK pathway, without interfering with CDDP pharmacokinetics. Therefore, RXN could potentially be used to reduce CDDP-related hearing loss and dizziness.     

Mitochondrial Peroxiredoxin 3 Regulates Sensory Cell Survival in the Cochlea

This study delineates the role of peroxiredoxin 3 (Prx3) in hair cell death induced by several etiologies of acquired hearing loss (noise trauma, aminoglycoside treatment, age). In vivo, Prx3 transiently increased in mouse cochlear hair cells after traumatic noise exposure, kanamycin treatment, or with progressing age before any cell loss occurred; when Prx3 declined, hair cell loss began. Maintenance of high Prx3 levels via treatment with the radical scavenger 2,3-dihydroxybenzoate prevented kanamycin-induced hair cell death. Conversely, reducing Prx3 levels with Prx3 siRNA increased the severity of noise-induced trauma. In mouse organ of Corti explants, reactive oxygen species and levels of Prx3 mRNA and protein increased concomitantly at early times of drug challenge. When Prx3 levels declined after prolonged treatment, hair cells began to die. The radical scavenger p-phenylenediamine maintained Prx3 levels and attenuated gentamicin-induced hair cell death. Our results suggest that Prx3 is up-regulated in response to oxidative stress and that maintenance of Prx3 levels in hair cells is a critical factor in their susceptibility to acquired hearing loss.     

Degradation of cochlear Connexin26 accelerate the development of age-related hearing loss

The GJB2 gene, encoding Connexin26 (Cx26), is one of the most common causes of inherited deafness. Clinically, mutations in GJB2 cause congenital deafness or late-onset progressive hearing loss. Recently, it has been reported that Cx26 haploid deficiency accelerates the development of age-related hearing loss (ARHL). However, the roles of cochlear Cx26 in the hearing function of aged animals remain unclear. In this study, we revealed that the Cx26 expression was significantly reduced in the cochleae of aged mice, and further explored the underlying molecular mechanism for Cx26 degradation. Immunofluorescence co-localization results showed that Cx26 was internalized and degraded by lysosomes, which might be one of the important ways for Cx26 degradation in the cochlea of aged mice. Currently, whether the degradation of Cx26 in the cochlea leads directly to ARHL, as well as the mechanism of Cx26 degradation-related hearing loss are still unclear. To address these questions, we generated mice with Cx26 knockout in the adult cochlea as a model for the natural degradation of Cx26. Auditory brainstem response (ABR) results showed that Cx26 knockout mice exhibited high-frequency hearing loss, which gradually progressed over time. Pathological examination also revealed the degeneration of hair cells and spiral ganglions, which is similar to the phenotype of ARHL. In summary, our findings suggest that degradation of Cx26 in the cochlea accelerates the occurrence of ARHL, which may be a novel mechanism of ARHL.     

Effect of c-myc on the ultrastructural structure of cochleae in guinea pigs with noise induced hearing loss

Noise over-stimulation may induce hair cells loss and hearing deficit. The c-myc oncogene is a major regulator for cell proliferation, growth, and apoptosis. However, the role of this gene in the mammalian cochlea is still unclear. The study was designed to firstly investigate its function under noise condition, from the aspect of cochlear ultrastructural changes. We had established the adenoviral vector of c-myc gene and delivered the adenovirus suspension into the scala tympani of guinea pigs 4 days before noise exposure. The empty adenoviral vectors were injected as control. Then, all subjects were exposed to 4-kHz octave-band noise at 110 dB SPL for 8 h/day, 3 days consecutively. Auditory thresholds were assessed by auditory brainstem response, prior to and 7 days following noise exposure. On the seventh days after noise exposure, the cochlear sensory epithelia surface was observed microscopically and the cochleae were taken to study the ultrastructural changes. The results indicated that auditory threshold shift after noise exposure was higher in the ears treated with Ad.EGFP than that treated with Ad.c-myc-EGFP. Stereocilia loss and the disarrangement of outer hair cells were observed, with greater changes found in the Ad.EGFP group. Also, the ultrastructure changes were severe in the Ad.EGFP group, but not obvious in the Ad.c-myc-EGFP group. Therefore, c-myc gene might play an unexpected role in hearing functional and morphological protection from acoustic trauma.     

窄带噪声暴露后豚鼠听阈偏移及毛细胞的损伤

为了探讨听觉损伤与毛细胞损伤的关系,本实验比较了噪声暴露后豚鼠皮层听阈及其与基底膜单位长度上毛细胞损伤率的关系。暴露声源中心频率1000Hz,为1/3倍频程的窄带噪声。强度为136dB作用1小时。108dB每天暴露1小时,5天/周连续1个月。结果表明,毛细胞损伤呈灶性,损伤部位与周围界线十分分明。毛细胞损伤在豚鼠间及左右耳间均存在相当程度的个体差异。136dB暴露,毛细胞损伤最重的部位在基底膜1.5转到2.5转之间,符合1000Hz声音在基底膜的最大振动范围。108dB的损伤部位局限在1.5转,其范围及程度明显轻于136dB。136dB造成的听阈偏移高于108dB,尤其在4、8 KHz高频听阈偏移最明显,但耳蜗底转毛细胞多无明显损伤。     

Reduced acoustic startle response and peripheral hearing loss in the 5xFAD mouse model of Alzheimer's disease

Hearing dysfunction has been associated with Alzheimer's disease (AD) in humans, but there is little data on the auditory function of mouse models of AD. Furthermore, characterization of hearing ability in mouse models is needed to ensure that tests of cognition that use auditory stimuli are not confounded by hearing dysfunction. Therefore, we assessed acoustic startle response and pre‐pulse inhibition in the double transgenic 5xFAD mouse model of AD from 3–4 to 16 months of age. The 5xFAD mice showed an age‐related decline in acoustic startle as early as 3–4 months of age. We subsequently tested auditory brainstem response (ABR) thresholds at 4 and 13–14 months of age using tone bursts at frequencies of 2–32 kHz. The 5xFAD mice showed increased ABR thresholds for tone bursts between 8 and 32 kHz at 13–14 months of age. Finally, cochleae were extracted and basilar membranes were dissected to count hair cell loss across the cochlea. The 5xFAD mice showed significantly greater loss of both inner and outer hair cells at the apical and basal ends of the basilar membrane than wild‐type mice at 15–16 months of age. These results indicate that the 5xFAD mouse model of AD shows age‐related decreases in acoustic startle responses, which are at least partially due to age‐related peripheral hearing loss. Therefore, we caution against the use of cognitive tests that rely on audition in 5xFAD mice over 3–4 months of age, without first confirming that performance is not confounded by hearing dysfunction.