ABR/DPOAE publications

Hearing loss in children is common especially in NICU with consequences of worse outcomes in speech, language, education, social functioning, cognitive abilities, and quality of life. Whether neonatal pain is link to increase risks for hearing loss remains to be explored. Here, we implemented Complete Freund's adjuvant (CFA) plantar injection and needle prick model to mimic neonatal pain in NICU during critical period of hearing development. Auditory brainstem response (ABR) test was used to determine the hearing threshold at 4w and 8w postnatal. Sufentanil and Oxycodone were used as analgesic to treat neonatal pain. Hair cell and ribbon synapse stanning were performed to detect cochlear function. Golgi-cox staining and BDNF immunofluorescence of auditory cortex were performed to determine dendritic spine pruning in auditory cortex. The dendritic pruning related protein CaMKII and Rac1/2 level were detected by western blot. We found that CFA induced neonatal pain and ABR threshold increased at 4w and 8w postnatal and the impairment were attenuated after analgesic administration. Neither the inner hair cell (IHC) nor the synapse of CFA mice was damaged in cochlear. CFA mice showed increased dendritic spine density at auditory cortex and reduced BDNF level. Furthermore, Rac1/2 and CaMKII might contributed to the disrupt dendritic spine pruning. Our study suggested that neonatal pain could induced hearing impairment in adulthood ascribed to the reduced BDNF level and AC dendritic spine pruning deficiency, optimal analgesic in early-life could beneficial for hearing development.

Previous psychophysical studies have identified a hierarchy of time-averaged statistics which determine the identity of natural sound textures. However, it is unclear whether the neurons in the inferior colliculus (IC) are sensitive to each of these statistical features in the natural sound textures. We used 13 representative sound textures spanning the space of 3 statistics extracted from over 200 natural textures. The synthetic textures were generated by incorporating the statistical features in a step-by-step manner, in which a particular statistical feature was changed while the other statistical features remain unchanged. The extracellular activity in response to the synthetic texture stimuli was recorded in the IC of anesthetized rats. Analysis of the transient and sustained multiunit activity after each transition of statistical feature showed that the IC units were sensitive to the changes of all types of statistics, although to a varying extent. For example, we found that more neurons were sensitive to the changes in variance than that in the modulation correlations. Our results suggest that the sensitivity of the statistical features in the subcortical levels contributes to the identification and discrimination of natural sound textures.

Pathogenic variants in GJB2, the gene encoding connexin 26, are the most common cause of autosomal-recessive hereditary deafness. Despite this high prevalence, pathogenic mechanisms leading to GJB2-related deafness are not well understood, and cures are absent. Humans with GJB2-related deafness retain at least some auditory hair cells and neurons, and their deafness is usually stable. In contrast, mice with conditional loss of Gjb2 in supporting cells exhibit extensive loss of hair cells and neurons and rapidly progress to profound deafness, precluding the application of therapies that require intact cochlear cells. In an attempt to design a less severe Gjb2 animal model, we generated mice with inducible Sox10iCre ERT2 -mediated loss of Gjb2. Tamoxifen injection led to reduced connexin 26 expression and impaired function, but cochlear hair cells and neurons survived for 2 months, allowing phenotypic rescue attempts within this time. AAV-mediated gene transfer of GJB2 in mature mutant ears did not demonstrate threshold improvement and in some animals exacerbated hearing loss and resulted in hair cell loss. We conclude that Sox10iCre ERT2 ;Gjb2 flox/flox mice are valuable for studying the biology of connexin 26 in the cochlea. In particular, these mice may be useful for evaluating gene therapy vectors and development of therapies for GJB2-related deafness.

Failure in the structural maintenance of the hair cell stereocilia bundle and ribbon synapse causes hearing loss. Here, we have studied how ER stress elicits hair cell pathology, using mouse models with inactivation of Manf (mesencephalic astrocyte-derived neurotrophic factor), encoding an ER-homeostasis-promoting protein. From hearing onset, Manf deficiency caused disarray of the outer hair cell stereocilia bundle and reduced cochlear sound amplification capability throughout the tonotopic axis. In high-frequency outer hair cells, the pathology ended in molecular changes in the stereocilia taper region and in strong stereocilia fusion. In high-frequency inner hair cells, Manf deficiency degraded ribbon synapses. The altered phenotype strongly depended on the mouse genetic background. Altogether, the failure in the ER homeostasis maintenance induced early-onset stereociliopathy and synaptopathy and accelerated the effect of genetic causes driving age-related hearing loss. Correspondingly, MANF mutation in a human patient induced severe sensorineural hearing loss from a young age onward. Thus, we present MANF as a novel protein and ER stress as a mechanism that regulate auditory hair cell maintenance in both mice and humans.

Binaural hearing is critically important for the perception of sound spatial locations. The primary auditory cortex (AI) has been demonstrated to be necessary for sound localization. However, after hearing onset, how the processing of binaural cues by AI neurons develops, and how the binaural processing of AI neurons is affected by reversible unilateral conductive hearing loss (RUCHL), are not fully elucidated. Here, we determined the binaural processing of AI neurons in four groups of rats: postnatal day (P) 14–18 rats, P19–30 rats, P57–70 adult rats, and RUCHL rats (P57–70) with RUCHL during P14–30. We recorded the responses of AI neurons to both monaural and binaural stimuli with variations in interaural level differences (ILDs) and average binaural levels. We found that the monaural response types, the binaural interaction types, and the distributions of the best ILDs of AI neurons in P14–18 rats are already adult-like. However, after hearing onset, there exist developmental refinements in the binaural processing of AI neurons, which are exhibited by the increase in the degree of binaural interaction, and the increase in the sensitivity and selectivity to ILDs. RUCHL during early hearing development affects monaural response types, decreases the degree of binaural interactions, and decreases both the selectivity and sensitivity to ILDs of AI neurons in adulthood. These new evidences help us to understand the refinements and plasticity in the binaural processing of AI neurons during hearing development, and might enhance our understanding in the neuronal mechanism of developmental changes in auditory spatial perception.

Mitochondrial oxidative stress is involved in hair cell damage caused by noise-induced hearing loss (NIHL). Sirtuin-3 (SIRT3) plays an important role in hair cell survival by regulating mitochondrial function; however, the role of SIRT3 in NIHL is unknown. In this study, we used 3-TYP to inhibit SIRT3 and found that this inhibition aggravated oxidative damage in the hair cells of mice with NIHL. Moreover, 3-TYP reduced the enzymatic activity and deacetylation levels of superoxide dismutase 2 (SOD2). Subsequently, we administered adeno-associated virus-SIRT3 to the posterior semicircular canals and found that SIRT3 overexpression significantly attenuated hair cell injury and that this protective effect of SIRT3 could be blocked by 2-methoxyestradiol, a SOD2 inhibitor. These findings suggest that insufficient SIRT3/SOD2 signaling leads to mitochondrial oxidative damage resulting in hair cell injury in NIHL. Thus, ameliorating noise-induced mitochondrial redox imbalance by intervening in the SIRT3/SOD2 signaling pathway may be a new therapeutic target for hair cell injury.

Caffeine is being increasingly used in daily life, such as in drinks, cosmetics, and medicine. Caffeine is known as a mild stimulant of the central nervous system, which is also closely related to neurologic disease. However, it is unknown whether caffeine causes hearing loss, and there is great interest in determining the effect of caffeine in cochlear hair cells. First, we explored the difference in auditory brainstem response (ABR), organ of Corti, stria vascularis, and spiral ganglion neurons between the control and caffeine-treated groups of C57BL/6 mice. RNA sequencing was conducted to profile mRNA expression differences in the cochlea of control and caffeine-treated mice. A CCK-8 assay was used to evaluate the approximate concentration of caffeine. Flow cytometry, TUNEL assay, immunocytochemistry, qRT-PCR, and Western blotting were performed to detect the effects of SGK1 in HEI-OC1 cells and basilar membranes. In vivo research showed that 120 mg/ kg caffeine injection caused hearing loss by damaging the organ of Corti, stria vascularis, and spiral ganglion neurons. RNA-seq results suggested that SGK1 might play a vital role in ototoxicity. To confirm our observations in vitro , we used the HEI-OC1 cell line, a cochlear hair cell-like cell line, to investigate the role of caffeine in hearing loss. The results of flow cytometry, TUNEL assay, immunocytochemistry, qRT-PCR, and Western blotting showed that caffeine caused autophagy and apoptosis via SGK1 pathway. We verified the interaction between SGK1 and HIF-1α by co-IP. To confirm the role of SGK1 and HIF-1α, GSK650394 was used as an inhibitor of SGK1 and CoCl 2 was used as an inducer of HIF-1α. Western blot analysis suggested that GSK650394 and CoCl 2 relieved the caffeine-induced apoptosis and autophagy. Together, these results indicated that caffeine induces autophagy and apoptosis in auditory hair cells via the SGK1/HIF-1α pathway, suggesting that caffeine may cause hearing loss. Additionally, our findings provided new insights into ototoxic drugs, demonstrating that SGK1 and its downstream pathways may be potential therapeutic targets for hearing research at the molecular level.

Current clinical strategies to assess benefits from hearing aids (HAs) are based on self-reported questionnaires and speech-in-noise (SIN) tests; which require behavioural cooperation. Instead, objective measures based on Auditory Brainstem Responses (ABRs) to speech stimuli would not require the individuals’ cooperation. Here, we re-analysed an existing dataset to predict behavioural measures with speech-ABRs using regression trees. Ninety-two HA users completed a self-reported questionnaire (SSQ-Speech) and performed two aided SIN tests: sentences in noise (BKB-SIN) and vowel-consonant-vowels (VCV) in noise. Speech-ABRs were evoked by a 40 ms [da] and recorded in 2x2 conditions: aided vs. unaided and quiet vs. background noise. For each recording condition, two sets of features were extracted: 1) amplitudes and latencies of speech-ABR peaks, 2) amplitudes and latencies of speech-ABR F0 encoding. Two regression trees were fitted for each of the three behavioural measures with either feature set and age, digit-span forward and backward, and pure tone average (PTA) as possible predictors. The PTA was the only predictor in the SSQ-Speech trees. In the BKB-SIN trees, performance was predicted by the aided latency of peak F in quiet for participants with PTAs between 43 and 61 dB HL. In the VCV trees, performance was predicted by the aided F0 encoding latency and the aided amplitude of peak VA in quiet for participants with PTAs ≤ 47 dB HL. These findings indicate that PTA was more informative than any speech-ABR measure, as these were relevant only for a subset of the participants. Therefore, speech-ABRs evoked by a 40 ms [da] are not a clinical predictor of behavioural measures in HA users.

Ca 2+ signaling is a major contributor to sensory hair cell function in the cochlea. Oncomodulin (OCM) is a Ca 2+ binding protein (CaBP) preferentially expressed in outer hair cells (OHCs) of the cochlea and few other specialized cell types. Here, we expand on our previous reports and show that OCM delays hearing loss in mice of two different genetic backgrounds: CBA/CaJ and C57Bl/6J. In both backgrounds, genetic disruption of Ocm leads to early progressive hearing loss as measured by auditory brainstem response (ABR) and distortion product otoacoustic emission (DPOAE). In both strains, loss of Ocm reduced hearing across lifetime (hearing span) by more than 50% relative to wild type (WT). Even though the two WT strains have very different hearing spans, OCM plays a considerable and similar role within their genetic environment to regulate hearing function. The accelerated age-related hearing loss (ARHL) of the Ocm KO illustrates the importance of Ca 2+ signaling in maintaining hearing health. Manipulation of OCM and Ca 2+ signaling may reveal important clues to the systems of function/dysfunction that lead to ARHL.

Although fear memory formation is essential for survival and fear-related mental disorders, the neural circuitry and mechanism are incompletely understood. Here, we utilized trace fear conditioning to study the formation of trace fear memory in mice. We identified the entorhinal cortex (EC) as a critical component of sensory signaling to the amygdala. We adopted both loss-of-function and gain-of-function experiments to demonstrate that release of the cholecystokinin (CCK) from the EC is required for trace fear memory formation. We discovered that CCK-positive neurons project from the EC to the lateral nuclei of the amygdala (LA), and inhibition of CCK-dependent signaling in the EC prevented long-term potentiation of the auditory response in the LA and formation of trace fear memory. In summary, high-frequency activation of EC neurons triggers the release of CCK in their projection terminals in the LA, potentiating auditory response in LA neurons. The neural plasticity in the LA leads to trace fear memory formation.

Calcitonin-gene-related peptide (CGRP) is a lateral olivocochlear (LOC) efferent neurotransmitter. Depression of sound-driven auditory brainstem response amplitude in CGRP-null mice suggests the potential for endogenous CGRP release to upregulate spontaneous and/or sound-driven auditory nerve (AN) activity. We chronically infused CGRP into the guinea pig cochlea and evaluated changes in AN activity as well as outer hair cell (OHC) function. The amplitude of both round window noise (a measure of ensemble spontaneous activity) and the synchronous whole-nerve response to sound (compound action potential, CAP) were enhanced. Lack of change in both onset adaptation and steady state amplitude of sound-evoked distortion product otoacoustic emission (DPOAE) responses indicated CGRP had no effect on OHCs, suggesting the origin of the observed changes was neural. Combined with results from the CGRP-null mice, these results appear to confirm that endogenous CGRP enhances auditory nerve activity when released by the LOC neurons. However, infusion of the CGRP receptor antagonist CGRP (8–37) did not reliably influence spontaneous or sound-driven AN activity, or OHC function, results that contrast with the decreased ABR amplitude measured in CGRP-null mice.

Recent studies have identified sex-differences in auditory physiology and in the susceptibility to noise-induced hearing loss (NIHL). We hypothesize that 17β-estradiol (E2), a known modulator of auditory physiology, may underpin sex-differences in the response to noise trauma. Here, we gonadectomized B6CBAF1/J mice and used a combination of electrophysiological and histological techniques to study the effects of estrogen replacement on peripheral auditory physiology in the absence of noise exposure and on protection from NIHL. Functional analysis of auditory physiology in gonadectomized female mice revealed that E2-treatment modulated the peripheral response to sound in the absence of changes to the endocochlear potential compared to vehicle-treatment. E2-replacement in gonadectomized female mice protected against hearing loss following permanent threshold shift (PTS)- and temporary threshold shift (TTS)-inducing noise exposures. Histological analysis of the cochlear tissue revealed that E2-replacement mitigated outer hair cell loss and cochlear synaptopathy following noise exposure compared to vehicle-treatment. Lastly, using fluorescent in situ hybridization, we demonstrate co-localization of estrogen receptor-2 with type-1C, high threshold spiral ganglion neurons, suggesting that the observed protection from cochlear synaptopathy may occur through E2-mediated preservation of these neurons. Taken together, these data indicate the estrogen signaling pathways may be harnessed for the prevention and treatment of NIHL.

Usher syndrome type I (USH1) is characterized by deafness, vestibular areflexia, and progressive retinal degeneration. The protein-truncating p.Arg245* founder variant of PCDH15 (USH1F) has an ~2% carrier frequency amongst Ashkenazi Jews accounts for ~60% of their USH1 cases. Here, longitudinal phenotyping in 13 USH1F individuals revealed progressive retinal degeneration, leading to severe vision loss with macular atrophy by the sixth decade. Half of the affected individuals were legally blind by their mid-50s. The mouse Pcdh15 R250X variant is equivalent to human p.Arg245*. Homozygous Pcdh15 R250X mice also have visual deficits and aberrant light-dependent translocation of the phototransduction cascade proteins, arrestin, and transducin. Retinal pigment epithelium (RPE)-specific retinoid cycle proteins, RPE65 and CRALBP, were also reduced in Pcdh15 R250X mice, indicating a dual role for protocadherin-15 in photoreceptors and RPE. Exogenous 9- cis retinal improved ERG amplitudes in Pcdh15 R250X mice, suggesting a basis for a clinical trial of FDA-approved retinoids to preserve vision in USH1F patients.

A common complaint of older adults is difficulty understanding speech, particularly in challenging listening conditions. Accumulating evidence suggests that these difficulties may reflect a loss and/or dysfunction of auditory nerve (AN) fibers. We used a novel approach to study age-related changes in AN structure and several measures of AN function, including neural synchrony, in 58 older adults and 42 younger adults. AN activity was measured in response to an auditory click (compound action potential; CAP), presented at stimulus levels ranging from 70 to 110 dB pSPL. Poorer AN function was observed for older than younger adults across CAP measures at higher but not lower stimulus levels. Associations across metrics and stimulus levels were consistent with age-related AN disengagement and AN dyssynchrony. High-resolution T2-weighted structural imaging revealed age-related differences in the density of cranial nerve VIII, with lower density in older adults with poorer neural synchrony. Individual differences in neural synchrony were the strongest predictor of speech recognition, such that poorer synchrony predicted poorer recognition of time-compressed speech and poorer speech recognition in noise for both younger and older adults. These results have broad clinical implications and are consistent with an interpretation that age-related atrophy at the level of the AN contributes to poorer neural synchrony and may explain some of the perceptual difficulties of older adults. SIGNIFICANCE STATEMENT Differences in auditory nerve (AN) pathophysiology may contribute to the large variations in hearing and communication abilities of older adults. However, current diagnostics focus largely on the increase in detection thresholds, which is likely because of the absence of indirect measures of AN function in standard clinical test batteries. Using novel metrics of AN function, combined with estimates of AN structure and auditory function, we identified age-related differences across measures that we interpret to represent age-related reductions in AN engagement and poorer neural synchrony. Structure-function associations are consistent with an explanation of AN deficits that arise from age-related atrophy of the AN. Associations between neural synchrony and speech recognition suggest that individual and age-related deficits in neural synchrony contribute to speech recognition deficits.

The medial olivocochlear (MOC) efferent feedback circuit projecting to the cochlear outer hair cells (OHCs) confers protection from noise-induced hearing loss and is generally thought to be driven by inner hair cell (IHC) - type I spiral ganglion afferent (SGN) input. Knockout of the Prph gene ( Prph KO) encoding the peripherin type III intermediate filament disrupted the OHC - type II SGN innervation and virtually eliminated MOC – mediated contralateral suppression from noise delivered to the opposite ear, measured as a reduction in cubic distortion product otoacoustic emissions. Electrical stimulation of the MOC pathway elicited contralateral suppression indistinguishable between wildtype (WT) and Prph KO mice, indicating that the loss of contralateral suppression was not due to disruption of the efferent arm of the circuit; IHC – type I SGN input was also normal, based on auditory brainstem responses. High-intensity, broadband noise (108 dB SPL, 1 hour) produced permanent hearing loss in Prph KO mice, but not in WT littermates. These findings associate OHC-type II input with MOC efferent - based otoprotection at loud sound levels.

The binaural interaction component (BIC) is a sound-evoked electrophysiological signature of binaural processing in the auditory brainstem that has received attention as a potential biomarker for spatial hearing deficits. Yet the number of trials necessary to evoke the BIC, or its measurability, seems to vary across species: while it is easily measured in small rodents, it has proven to be highly variable and less reliably measured in humans. This has hindered its potential use as a diagnostic tool. Further measurements of the BIC across a wide range of species could help us better understand its origin and the possible reasons for the variation in its measurability. Statistical analysis on the function relating BIC DN1 amplitude and the interaural time difference has been performed in only a few small rodent species, thus it remains to be shown how the results apply to more taxonomically diverse mammals, and those with larger heads. To fill this gap, we measured BICs in rhesus macaque. We show the overall behavior of the BIC is the same as in smaller rodents, suggesting that the brainstem circuit responsible for the BIC is conserved across a wider range of mammals. We suggest that differences in measurability are likely because of differences in head size.

Auditory brainstem response (ABR) serves as an objective indication of auditory perception at a given sound level and is nowadays widely used in hearing function assessment. Despite efforts for automation over decades, ABR threshold determination by machine algorithms remains unreliable and thereby one still relies on visual identification by trained personnel. Here, we described a procedure for automatic threshold determination that can be used in both animal and human ABR tests. The method terminates level averaging of ABR recordings upon detection of time-locked waveform through cross-correlation analysis. The threshold level was then indicated by a dramatic increase in the sweep numbers required to produce "qualified" level averaging. A good match was obtained between the algorithm outcome and the human readouts. Moreover, the method varies the level averaging based on the cross-correlation, thereby adapting to the signal-to-noise ratio of sweep recordings. These features empower a robust and fully automated ABR test.

Cochlear implantation (CI) is the major treatment for severe sensorineural hearing loss. However, the fibrotic tissue forming around the electrodes reduces the treatment effectiveness of CI. Dexamethasone (DEX) is usually applied routinely in perioperative treatment of cochlear implantation (CI), but its diffusion in the inner ear after systemic administration is limited. In the present study, an electrode coated with polycaprolactone (PCL) loaded with dexamethasone was developed with a simple preparation process to maintain the stability of the electrode itself. The DEX-loaded PCL coating has good biocompatibility and does not change the smoothness, flexibility, or compliance of the implant electrode. Stable and effective DEX concentrations were maintained for more than 9 months. Compared with the pristine electrode, decreasing intracochlear fibrosis, protection of hair cells and spiral ganglion cells, and better residual hearing were observed 5 weeks after PCL-DEX electrode implantation. The PCL-DEX electrode has great potential in preventing hearing loss and fibrosis by regulating macrophages and inhibiting the expression of the fibrosis-related factors IL-1β, TNF-α, IL-4, and TGF-β1. In conclusion, the PCL-DEX electrode coating shows promising application in CI surgery.

Hearing loss is the most common human sensory deficit. Hearing relies on stereocilia, inserted into the cuticular plate of hair cells (HCs), where they play an important role in the perception of sound and its transmission. Although numerous genes have been associated with hearing loss, the function of many hair cell genes has yet to be elucidated. Herein, we focused on nonerythroid spectrin αII (SPTAN1), abundant in the cuticular plate, surrounding the rootlets of stereocilia and along the plasma membrane. Interestingly, mice with HC-specific Sptan1 knockout exhibited rapid deafness, abnormal formation of stereocilia and cuticular plates, and loss of HCs from middle and apical turns of the cochlea during early postnatal stages. Additionally, Sptan1 deficiency led to the decreased spreading of House Ear Institute-Organ of Corti 1 cells, and induced abnormal formation of focal adhesions and integrin signaling in mouse HCs. Altogether, our findings highlight SPTAN1 as a critical molecule for HC stereocilia morphology and auditory function via regulation of focal adhesion signaling.

Little is known about the ontogeny of lingual echolocation. We examined the echolocation development of Rousettus aegyptiacus , the Egyptian fruit bat, which uses rapid tongue movements to produce hyper-short clicks and steer the beam's direction. We recorded from day 0 to day 35 postbirth and assessed hearing and beam-steering abilities. On day 0, R. aegyptiacus pups emit isolation calls and hyper-short clicks in response to acoustic stimuli, demonstrating hearing. Auditory brainstem response recordings show that pups are sensitive to pure tones of the main hearing range of adult Rousettus and to brief clicks. Newborn pups produced clicks in the adult paired pattern and were able to use their tongues to steer the sonar beam. As they aged, pups produced click pairs faster, converging with adult intervals by age of first flights (7–8 weeks). In contrast with laryngeal bats, Rousettus echolocation frequency and duration are stable through to day 35, but shift by the time pups begin to fly, possibly owing to tongue-diet maturation effects. Furthermore, frequency and duration shift in the opposite direction of mammalian laryngeal vocalizations. Rousettus lingual echolocation thus appears to be a highly functional sensory system from birth and follows a different ontogeny from that of laryngeal bats.