ABR/DPOAE publications

Kanamycin (KM), a widely used aminoglycoside antibiotic, is limited by its ototoxic side effects, primarily targeting cochlear outer hair cells (OHCs) and leading to irreversible sensorineural hearing loss. Our previous work showed that silencing AMP‐activated protein kinase alpha 1 (AMPKα1) prevents noise‐induced hearing loss through a redox‐sensitive regulatory mechanism. Given that both noise‐induced and aminoglycoside‐induced hearing loss share oxidative stress as a key pathological feature, we tested whether AMPKα1 silencing could mitigate KM‐induced damage using an acute ototoxic model combining KM with furosemide (KM + FU). KM + FU exposure markedly increased AMPKα phosphorylation at Thr172. Targeted siRNA‐mediated silencing via posterior semicircular canal delivery in CBA/J mice significantly reduced AMPKα1 expression in OHCs and effectively prevented KM + FU‐induced OHC loss and hearing impairment. This preventive effect was independent of stria vascularis permeability or cochlear KM uptake. These findings identify AMPKα as a contributor to KM + FU‐induced acute ototoxicity and suggest that its inhibition may be a promising strategy for hearing preservation.

Optical coherence tomography has allowed in vivo recording of sound-induced vibrations of different regions within the organ of Corti complex (OCC), including the basilar membrane (BM), outer hair cell/Deiters cell (OHC/DC) region, and reticular lamina (RL). In the hook region of the gerbil cochlea, where measurements can be made with a predominantly transverse optical axis, the three regions have different and characteristic motion responses: The OHC/DC region has greater motions than the other two regions at frequencies below the best frequency (sub-BF); the RL region typically has the greatest BF peak and smallest sub-BF motion. The phase of the OHC/DC-region motion increasingly lags BM motion phase as frequency increases; the RL-region motion phase leads BM, but with a relatively small value. All three regions are compressively nonlinear in the BF peak, but only the OHC/DC region shows sub-BF compressive nonlinearity. Here, we describe the strains that arise when the motion transitions between these regions. Large strains (deformations) were observed in the OHC body close to the RL. A region of large strain can be as short as a single 2.7-μm measurement pixel, or it can extend over several pixels, with extensive strains appearing more often at 70 than at 50 dB sound pressure level. Beyond this RL region of large strain, over a distance that can exceed 20 μm, the OHC/DC region displayed nearly spatially unvarying motion; this region vibrated as a relatively rigid body. And beyond this, at sub-BF frequencies, another region of high strain was present, indicating stretching/compressing or bowing/tilting of the DC microtubular stalk.

Objectives. This study aimed to explore the role of the Pcdh15 gene in hearing maintenance and to examine the effects of its deficiency on cochlear structure, cochlear function, and hearing loss in mice.Methods. CRISPR/Cas9 technology was used to generate Pcdh15 knockout (KO) mice. Auditory function was evaluated using hearing tests, while histological approaches were employed to assess morphological changes in cochlear hair cells and spiral ganglion neurons (SGNs). RNA sequencing (RNA-seq) was performed on cochlear tissue from postnatal day 18 (P18) wild-type (WT) and Pcdh15 KO mice to identify differentially expressed genes (DEGs). These DEGs were subjected to functional annotation and pathway analysis, with particular emphasis on oxidative phosphorylation (OXPHOS). Reactive oxygen species (ROS) levels were quantified using flow cytometry and DCFH-DA assays.Results. Pcdh15 KO mice exhibited progressive sensorineural hearing loss, which worsened to profound deafness by P18. Notably, cochlear hair cells and SGNs showed substantial loss and pronounced morphological abnormalities, particularly within the basal high-frequency region. RNA-seq analysis identified 1,359 DEGs, including 1,024 downregulated and 335 upregulated genes. Pathway analysis indicated that Pcdh15 deficiency was associated with inhibition of the OXPHOS pathway, potentially disrupting mitochondrial respiratory chain complexes I, III, IV, and V and thereby impairing energy production and ATP metabolism. In addition, early-stage cochleae from KO mice demonstrated elevated ROS levels and increased oxidative stress, suggesting that redox imbalance may contribute to hearing damage.Conclusion. Pcdh15 plays a critical role in hearing maintenance. Its deficiency is associated with severe hearing loss and marked cochlear abnormalities, which are linked to inhibition of OXPHOS, disruption of energy metabolism, and exacerbation of oxidative stress. Together, these findings provide new insights into the mechanisms underlying hearing loss and suggest potential targets for therapeutic intervention.

Background Stem cell therapy offers promise for the neurodegenerative diseases and has been explored for sensorineural hearing loss (SNHL). However, effective cell delivery strategies remain a critical challenge for SNHL treatment. Methods To address this need, we established an ouabain‐induced SGN injured hearing loss model in rat and evaluated a novel transplantation strategy targeting the cochlear nerve surface via posterior occipital approach, designed to minimize cochlear structural damage and facilitate targeted cell delivery to Rosenthal's canal (RC). The temporal changes in glial cell densities within RC revealed a progressively deteriorating neural microenvironment, supporting early‐stage intervention. Accordingly, hair follicle‐derived neural crest stem cell (HFNCSC) transplantation was performed 3 or 4 days after modeling via two approaches: cochlear nerve surface transplantation (CNT) and round window transplantation (RWT). Results CNT resulted in significant improvements in auditory function, as evidenced by reduced auditory brainstem response (ABR) thresholds, shortened wave I latencies, and preserved wave I amplitudes post‐transplantation. Transplanted cells were distributed along the nerve trunk and within RC. In contrast, RWT failed to improve auditory function and caused cochlear structural damage, with widespread cell dispersion in cochlear fluids. Notably, the CNT group exhibited significantly higher densities of TUJ1‐positive neuron‐like cells and glial cells in the RC, accompanied by enhanced myelin basic protein expression suggestive of remyelination. No such improvements were observed in the RWT group. Conclusions These findings suggest that cochlear nerve surface transplantation enhances stem cell survival and auditory function recovery, and represents a promising delivery approach for stem cell‐based therapy of SGN‐related hearing loss.

Blast-induced hearing loss (BIHL) is a prevalent form of sensory neurotrauma resulting from military and occupational blast exposure. However, effective post-injury interventions remain limited, and progress is hindered by challenges in delivering therapeutics to the injured cochlea. In this study, we developed a reproducible compressed-air blast mouse model with quantified output characteristics and evaluated a localized post-blast TrkB activation strategy using 7,8-dihydroxyflavone (7,8-DHF) delivered to the cochlea via poly(lactide-co-glycolide)-graft-polyethylenimine (PgP) nanoparticles. Device characterization demonstrated a monotonic relationship between regulator-setting pressure (100–250 psi) and acoustic output (72–124 dB). In addition, device modification increased acoustic output while reducing peak pressure relative to the prior configuration, thereby enabling reproducible, pressure-dependent injury. A single unilateral blast exposure produced robust elevations in auditory brainstem response (ABR) thresholds that partially recovered at intermediate regulator settings; however, these thresholds remained elevated over time at higher regulator settings. Since the modified blast paradigm frequently produced tympanic membrane perforation, we leveraged this transient access route to deliver 7,8-DHF-loaded nanoparticles through the ear canal immediately after blast. Compared with vehicle treatment, local 7,8-DHF delivery accelerated functional recovery and yielded significantly improved ABR thresholds, whereas vehicle-treated mice remained persistently impaired at 1 month. Histological analyses revealed preserved inner hair cell density, but significant outer hair cell loss and reduced inner hair cell ribbon synapses in mice exposed to blast. 7,8-DHF-loaded PgP attenuated outer hair cell loss and partially preserved ribbon synapses, with the strongest protection in basal cochlear regions. Together, these findings provide an accessible BIHL platform and support nanoparticle-enabled local TrkB activation as a feasible post-blast strategy that improves both functional and structural endpoints relevant to sensory neurotrauma.

The neural response of the brainstem to brief sounds, known as the auditory brainstem response (ABR), is widely employed in the laboratory and the clinic to diagnose hearing loss. Current ABR approaches are limited to analyzing the average ABR to hundreds of stimulus trials. Historically, trial-by-trial ABR analysis has not been possible owing to each trial's small signal-to-noise ratio. Here we overcome this limitation and show how to score individual ABR trials. We use the distribution of individual-trial ABR scores to define thresholds, which we call auditory brainstem detection (ABD) thresholds, and to assess suprathreshold hearing. Scores based on individual-trial amplitude and similarity produce lower ABD thresholds than scores based on individual-trial amplitude alone. Our method enables us to rank the suprathreshold hearing of individual subjects. Despite having normal ABR thresholds, some subjects appear to have suprathreshold hearing deficits.

Background: Prolonged exposure to moderate and loud noise is known to impair hearing; however, the safety of long-duration exposure to low-level sound, such as that encountered during everyday conversation, remains unclear. This study aimed to determine the effect of continuous exposure to sound at a 65 dB sound pressure level (SPL) on auditory processing. Methods: Auditory brainstem responses (ABRs) were recorded in C57BL/6 mice before and after a 1 h exposure to a continuous pure tone at 65 dB SPL. Changes in ABR thresholds, wave amplitudes, and latencies were analyzed across frequencies and time points. Correlations between amplitude and latency changes across ABR waves were also assessed. Results: Tone exposure induced a significant, frequency-specific increase in ABR thresholds, with a mean elevation of approximately 6 dB and a maximum shift of 15 dB. Significant reductions in amplitudes and prolongations of latencies were observed in Waves I–III, while Wave V amplitude remained relatively stable. A strong negative correlation between amplitude reduction and latency increase was found in Wave I, which progressively weakened from Wave II to Wave V. These functional changes persisted for up to three hours following exposure before gradually returning to baseline. Conclusions: Prolonged exposure to low-level sound at intensities typical of conversational speech can transiently impair auditory function and alter early neural processing in the auditory pathway. These findings suggest that sound levels commonly considered safe may still pose a risk when exposure is sustained, with implications for understanding hidden hearing loss and improving early diagnostic approaches.

Noise-induced hearing loss (NIHL) is classically attributed to glutamate excitotoxicity, yet the precise requirement for synaptic glutamate release remains untested using genetic approaches. Here, we investigated noise-induced cochlear damage using Vglut3 knockout ( Vglut3 -KO) mice, which lack glutamatergic transmission from inner hair cells. Following 116 dB SPL noise exposure, wild-type mice exhibited permanent threshold shifts, persistent ABR wave I amplitude reduction, and significant ribbon synapse loss at the 32 kHz region. Unexpectedly, Vglut3 -KO mice also exhibited significant synapse loss despite their inability to release glutamate, revealing glutamate-independent mechanisms of synaptopathy. Proteomic analysis identified 62 differentially expressed proteins in noise-exposed cochleae, with significant enrichment in synaptic structures and extracellular matrix components, including up-regulation of Rps6ka1 , Cplx2 , and Cdk5 , and downregulation of Ppa2 . Importantly, noise exposure severely compromised AAV8- Vglut3 gene therapy efficacy in Vglut3 -KO mice: while unexposed mutants achieved complete functional recovery, noise-exposed animals failed to attain normal hearing thresholds. These findings demonstrate that high-intensity noise induces synaptopathy through both excitotoxic and non-excitotoxic mechanisms involving synaptic and ECM disruption, and that acoustic trauma creates persistent pathological changes limiting gene therapy outcomes, underscoring the need for comprehensive otoprotective strategies in hereditary deafness patients awaiting treatment.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants linked to adverse health effects. Recent epidemiological data suggest an association between PFAS exposure and hearing loss, but underlying mechanisms remain unclear. This study examined PFAS-induced auditory dysfunction in mice exposed to a mixture of five PFAS compounds (2 mg/L each) in drinking water for seven weeks. Ldlr-/- mice were used due to their susceptibility to metabolic dysfunction, a risk factor for hearing loss. Auditory brainstem responses (ABR) indicated that PFAS exposure significantly elevated hearing thresholds by 18-33 dB across multiple frequencies. Distortion product otoacoustic emissions (DPOAEs) revealed impaired outer hair cell (OHC) function, and immunohistochemical analysis indicated a 20% OHC loss in the basal turn of the cochlea. In addition, PFAS exposure reduced ABR wave-1 amplitudes, and caused a 50% reduction in spiral ganglion cell density, indicating impaired auditory nerve function. Overall, this study provides the first evidence for PFAS-induced high-frequency hearing loss in mice. The findings further indicated that cochlear OHCs and spiral ganglion neurons are potential targets in PFAS-induced hearing loss. Together, these data suggest that PFAS exposure elicits a multifaceted ototoxic response, affecting both sensory and neural elements of the cochlea.

Hereditary hearing loss, the most prevalent genetic sensory disorder, lacks approved pharmacological therapies and represents a compelling target for gene correction. Pathogenic variants in KCNQ4 account for ~9.5% of autosomal dominant nonsyndromic cases. Prior gene-editing strategies disrupting mutant alleles have failed to achieve durable auditory rescue. Here we employed a knock-in mouse model harboring the human KCNQ4 c.961 G > A (p.G321S) mutation to evaluate precise base editing. Dual-AAV delivery of the adenine base editor ABE8e achieved 21.4–28.9% correction in the organ of Corti—the highest efficiency reported for genetic hearing loss. A dose-dependent therapeutic window emerged: higher doses promoted rapid recovery, whereas optimized lower doses minimized long-term toxicity and sustained functional benefit for at least 32 weeks. Treatment reduced auditory brainstem response thresholds by up to 49.09 dB SPL at optimal frequencies, mitigated degeneration of hair cells, spiral ganglion neurons, and auditory nerve fibers, and partially restored outer hair cell electrophysiology. These findings demonstrate the durability of precise mutation correction over allele-disruptive approaches and support clinical translation for KCNQ4 -associated hearing loss.

Hearing is the largest modifiable mid-life risk factor for Alzheimer's disease (AD), yet its link to dementia remains unclear. We identified a neurophysiological biomarker of AD risk using the non-invasive, rapidly acquired, and clinically translatable auditory brainstem response (ABR) in normal hearing knock-in rats (Swedish familial AD risk variant to Amyloid precursor protein, AppS; male and female). Human ABRs have been proposed as a biomarker for AD and related dementias. The novel metric reported here is derived from multidimensional parametric feature extraction on the distribution statistics of repeated single-trial ABR traces. We report accurate prediction of genetic risk for AD risk in young and aged rats: AppS separate clearly from healthy humanized (AppH) in sex- and age-dependent manners. Notably, auditory learning during young adulthood shifted the AppS ABR signature towards a healthy AppH-like state that maintained over time into older age. Altogether the findings support the utility of the ABR to track disease state, progression, and effects of intervention, and point to a central neural generator of auditory dysfunction related to AD risk. ABRs could provide a very early biomarker for detection of AD risk and used to test the synergy of auditory and cognitive functions in human dementia.

Aging and noise over-exposure lead to complex mixtures of cochlear degradation that impair the structure and function of outer hair cells, inner hair cells (IHCs), and the cochlear nerve. However, IHC damage and cochlear synaptopathy (CS) remain pathologies “hidden” from the audiogram. This study aimed to identify and differentiate the physiological signatures of these two distinct pathologies using promising non-invasive assays: Envelope Following Responses (EFRs), Auditory Brainstem Response (ABRs), Wideband middle-ear reflexes (WB-MEMRs), and Distortion Product Otoacoustic Emissions (DPOAEs). We utilized chinchilla models of carboplatin-induced (CA) IHC damage (N = 4) and temporary threshold shift (TTS) noise-induced CS (N = 4) to compare the physiological signatures of each pathology. While both groups showed unchanged ABR thresholds two weeks after exposure, EFRs, ABR Wave V/I ratios, and MEMRs showed distinct effects of exposure. Despite non-elevated ABR-derived audiometric thresholds after exposure, both CA and TTS exposure resulted in severe in EFR “peakiness”, particularly for sharp, short-duty-cycle stimuli and significant elevations in ABR Wave V/I ratios. However, these findings were less-pronounced in the TTS-exposed animals. WB-MEMR amplitudes were decreased with elevated thresholds in both groups; this effect was more pronounced in the TTS group. Opposite trends in DPOAE amplitudes indicated that while both IHC damage and CS result in similar suprathreshold temporal coding deficits, effects on outer-hair-cell integrity and auditory efferent physiology may differ between the two pathologies. Future work and novel diagnostics should aim to distinguish these specific cochlear pathologies in clinical populations, or at the very least consider their overlap. Highlights A multi-metric diagnostic approach was used with chinchilla models of inner-hair-cell (IHC) damage and cochlear synaptopathy (CS). IHC damage and synaptopathy both cause suprathreshold deficits “hidden” from the audiogram. IHC damage results in more severe temporal envelope coding degradation than does synaptopathy. A combination of EFR “peakiness”, ABR Wave V/I ratio, and Wideband Middle Ear Muscle Reflex (WB-MEMR) appear to be useful measures for profiling IHC damage and CS.

SLC26A4 is a major causative gene for hereditary hearing loss, its mutation spectrum shows pronounced population specificity. In Chinese populations, patients predominantly carry biallelic mutations, and compound heterozygous genotypes are prevalent, which results in a wide spectrum of auditory phenotypes. However, how different alleles interact within these contexts to shape phenotypic variability remains poorly understood. We employed cellular and mouse models to explore the allele-specific mechanisms associated with two novel mutations, a frameshift mutation and a missense mutation, in compound heterozygous that share the same splice-site pathogenic allele. In vitro, wild-type (WT) and mutant (c.574delC, c.1211C>A) SLC26A4 constructs were expressed in HeLa cells to assess pendrin localization. Both mutations reduced membrane enrichment and increased intracellular retention. In vivo, compound heterozygous knock-in mouse models (Slc26a4c.574delC/c.919-2A>G and Slc26a4c.1211C>A/c.919-2A>G) were generated using CRISPR/Cas9. The auditory function and cochlear pathology were investigated. Both compound mutants exhibited elevated ABR thresholds, with more severe hearing loss in Slc26a4c.574delC/c.919-2A>G mice. Correspondingly, these mice showed marked hair cell disruption, stereociliary loss, and cochlear structural abnormalities, whereas the Slc26a4c.1211C>A/c.919-2A>G mice displayed milder changes. Transcriptomic profiling examined by bulk RNA-sequencing revealed broader differential expression in Slc26a4c.574delC/c.919-2A>G mice, enriched in structural and developmental pathways, while the missense model showed predominantly immune-related signatures. Our findings demonstrate that allele-specific functional divergence in compound heterozygous SLC26A4 mutations leads to distinct auditory dysfunction, cochlear pathology, and transcriptional programs. These findings provide mechanistic insight into the phenotypic heterogeneity of hearing loss and may indicate future allele-specific interventions or therapeutic strategies.

Oncomodulin (OCM) is the most abundant Ca2+ buffering protein found in mature outer hair cells (OHCs). Cadherin 23 (CDH23) is a crucial component of the tip-links in hair cell stereocilia. The absence or dysfunction of these two proteins contributes to the early onset of age-related hearing loss (AHL). In this study, we investigated the effects of the Cdh23753G→A mutation on OHC function using new Ocm-knockout (KO) mouse models (Ocmtm1a/tm1a) with or without the Cdh23753G→A mutation. Despite having the same genetic background, Ocm-KO mice carrying the Cdh23753G→A mutation displayed a notable decline in OHC function across all measured frequencies as early as three months of age. In contrast, Ocm-KO mice without the Cdh23753G→A mutation did not exhibit comparable hearing loss until they reached twelve months of age. Additionally, we examined the role of OCM in preserving OHC function under ototoxic stress induced by HPβCD (2-hydroxypropyl-β-cyclodextrin). The distortion product otoacoustic emission data show that the administration of HPβCD resulted in a more pronounced decline in OHC function in Ocm-KO mice compared to wild-type (WT) mice. Time-lapse recording also shows that HPβCD treatment led to greater structural deterioration and more rapid rupture events in OHCs from Ocm-KO mice than in those from WT mice. These findings suggest that the Cdh23753G→A mutation, rather than other potential strain-specific genetic factors associated with AHL, significantly exacerbates the early onset of AHL phenotypes in Ocm-KO mice. Furthermore, our data indicates that the OCM protein in OHCs enhances their ability to withstand ototoxic stimuli.

The CUGBP Elav-like family 4 (CELF4), an RNA-binding protein, is dynamically expressed in spiral ganglion neurons (SGNs) in the cochlea over development, but how Celf4 is involved in regulating hearing functions is poorly understood. In this study, we generated a Celf4 ± mouse line and examined changes in the first two synapses along the auditory pathways. Firstly, we found that hearing functions were largely intact in Celf4 ± mice, with exception of reduced amplitude for Wave II of auditory brainstem responses (ABRs) and increased delays for Wave II and IV, both in case of 4 kHz only. Secondly, we found that counts of inner and outer hair cells (IHCs and OHCs) and SGNs remained unchanged in Celf4 ± mice, and that the number and function of ribbon synapses between IHCs and SGNs were comparable between WT and Celf4 ± mice. Lastly, function of the endbulb of Held synapse, formed between auditory nerve fibers (ANFs) of SGNs and bushy cells in the cochlear nucleus, was significantly altered in Celf4 ± mice. Specifically, synaptic vesicle release was subtly reduced, and excitability of bushy cells was significantly dampened, likely due to a hyperpolarized resting membrane potential. Furthermore, we found that spike kinetics was significantly faster in Celf4 ± bushy cells, likely caused by a larger fast-inactivating A-type potassium current (I A ) found in these cells. In conclusion, we found that Celf4 haploinsufficiency altered transmission at the endbulb of Held synapses in the cochlear nucleus in a significant and multifaceted manner, revealing the roles of Celf4 in regulating hearing functions. Graphical Abstract Title: Schematic representation of the experimental approach and major findings. Legend: In mice with Celf4 haploinsufficiency, we found that while function at hair cell ribbon synapses was largely preserved, transmission at the endbulb of Held synapse was significantly altered, including subtly reduced release of synaptic vesicles from the endbulb, dampened excitability in bushy cells, and accelerated spike kinetics caused by enhanced voltage-gated K + current.

Activity-dependent modulation of presynaptic voltage-gated Ca 2+ channels (Ca V 2) regulates Ca 2+ influx to control neuronal circuit output. Although Ca V 2.1 can undergo robust Ca 2+ dependent facilitation (CDF), whether it occurs in native central nervous system neuronal circuits is disputed. Accurate auditory information processing requires precise and reliable synaptic transmission at high activity rates in the auditory brainstem. To determine if Ca V 2.1 CDF is a key regulator of high-fidelity synaptic transmission, we expressed Ca V 2.1 splice variants capable (Ca V 2.1 37a) or incapable (Ca V 2.1 37b) of CDF at the calyx of Held presynaptic terminal in the auditory brainstem. We found no difference in basal Ca V 2.1 currents or synaptic transmission. However, Ca V 2.1 37b terminals lacked CDF, synaptic facilitation and had a decreased reliability and precision of postsynaptic action-potential firing. Additionally, the Wave III amplitude of the auditory brainstem responses was reduced. We propose that Ca V 2.1 CDF is essential for accurate auditory information processing.

Cochlear implants (CIs) have successfully restored hearing in more than one million patients with severe to profound hearing loss worldwide. Although CIs effectively restore speech perception in quiet environments, sound localization remains challenging for bilateral CI users, particularly their ability to utilize interaural time differences (ITDs). The majority of clinical CI processors use a coding strategy that encodes ITD information only in the envelope of electrical pulse trains rather than their pulse timing, which may contribute to the poorer spatial hearing perception of CI users. We recently demonstrated in a behavioural study on early deafened, bilaterally CI‐implanted rats that pulse timing ITDs completely dominate ITD perception, whereas sensitivity to envelope ITDs is almost negligible in comparison. Building on this, we here investigated the neurophysiological sensitivity of the inferior colliculus (IC) to envelope and pulse timing ITDs at two different pulse rates (900 and 4500 pulses/s) and three different stimulation modulations (5, 20 and 100 Hz) in CI rats with different hearing experiences. Our results indicate that IC neurons exhibit far greater sensitivity to pulse timing ITD than envelope ITD independent of pulse rate, modulation rate or hearing experience. These findings suggest that to improve binaural hearing outcomes in bilateral CI users, clinical stimulation strategies should provide informative pulse timing ITDs. image Key points Current bilateral cochlear implants mostly provide time cues of arriving sound in the envelope of stimulation rather than in the timing of pulses. A behavioural study on cochlear implanted rats has shown that pulse timing dominates binaural hearing. Here we examined the neural sensitivity to independent changes in time cues presented on the envelope or pulse timing, respectively, in rats with bilateral cochlear implants and different hearing experiences. Auditory midbrain neurons were consistently more sensitive to changes in pulse timing rather than envelope, irrespective of stimulus pulse rate, modulation rate or an animal's hearing experience.

Oxidative stress and inflammation are interconnected drivers of cellular damage in pathologies ranging from neurodegenerative disorders to sensorineural hearing loss. We aimed to develop a chitosan-Prussian blue nanozyme (CS-PB) to target these processes in sensorineural hearing loss. CS-PB (35 μg/mL) pretreatment of H 2 O 2 -injured HEI-OC1 cochlear cells for 4 h markedly decreased the levels of reactive oxygen species (ROS) inside cells (from 3.8 to 2.4 relative fluorescence intensity) by approximately 37% ( P < 0.001)-and concomitantly significantly decreased expression of the oxidative damage markers 4-HNE and 3-NT (both P < 0.001). The rate of apoptosis decreased from 27.5% in the H 2 O 2 -treated group to 14.1% following CS-PB treatment (P < 0.01). This reduction was accompanied by a significant downregulation of the pro-apoptotic proteins Bax (P < 0.05) and Cleaved-caspase-3 (P < 0.001), as well as an upregulation of the anti-apoptotic protein Bcl-XL (P < 0.01).Western blot analysis confirmed that CS–PB significantly downregulated TLR4 expression and inhibited downstream phosphorylation of p-P65/P65, while upregulating p-IκBα/IκBα protein (all P < 0.01) associated with reduction in the production of inflammatory cytokines like TNF-α, IL-1β, and IL-6. Treatment with the specific TLR4 antagonist TAK-242 (2 μM) mimicked the effect of CS-PB, with coadministration showing no additional ant-inflammatory effect (although HEI-OC1 viability was additionally increased), indicating that the anti-inflammatory properties of CS-PB were facilitated via the TLR4/NF-κB signaling pathway. In vivo , CS-PB pretreatment (2 mg/mL, 2 μL) was administered by carefully applying the solution to the round window membrane (RWM) using a microsyringe in a noise-induced hearing loss rat model, which significantly curtailed the noise-induced activation of the TLR4/NF-κB pathway in the cochlea. After 24 hours, the levels of p-P65/P65 and TLR4 decreased by 54% and 50%, respectively. ( P < 0.001), and levels of the inflammatory cytokines IL-6, TNF-α and IL-1β were decreased by 53%, 48%, and 51% ( P < 0.001). Furthermore, the degree of pathway inhibition showed a strong correlation with cytokine reduction (r = 0.87–0.89, all P < 0.001). CS-PB showed no significant cytotoxicity in vitro or in vivo , suggesting that its combined ROS-scavenging and TLR4/NF-κB–modulating properties may represent a potential therapeutic strategy, pending further mechanistic and translational validation.

Many neurological and psychiatric disorders stem from maladaptive conditioning and resist treatment. Tinnitus, as one example, is the distressing perception of sound in the absence of an external source. One of the network systems implicated in multiple disorders is the 'salience network', associated with positive/negative reward, habit formation, motivation, valuation and mood. However, it is not known whether directly modulating the 'salience network' at its anatomical and physiological core, in the ventral striatum, more specifically the nucleus accumbens, could be beneficial. Here we show that a novel treatment approach, targeting the salience network, can reduce tinnitus perception in a rat model. We found that bilateral rewarding stimulation of the nucleus accumbens, when paired with non-tinnitus tones, was most effective. A combined approach using rewarding accumbens stimulation and dysrewarding lateral habenula stimulation - paired with non-tinnitus and tinnitus tones, respectively - also suppressed tinnitus but caused negative effects. Our results demonstrate how the rewarding stimulation can recondition brain networks to effectively remove the salience of tinnitus frequencies. We anticipate that our unique reconditioning strategy has broader therapeutic applications for other brain disorders rooted in maladaptive learning.

Cisplatin is widely used in oncology yet causes dose-limiting, irreversible hearing loss through cochlear hair-cell injury. Although mitochondrial dysfunction is central to this ototoxicity, how the outer mitochondrial membrane (OMM) fails in hair cells remains unclear. We identify BAX-associated mitochondrial membrane rupture and abnormal permeabilization as key drivers. In HEI-OC1 cells and murine cochlear explants, super-resolution and transmission electron microscopy revealed cisplatin-induced BAX translocation and oligomerization on the OMM, forming ring-like assemblies with ultrastructural damage (rupture, herniation, fragmentation). Subcellular fractionation and functional assays demonstrated consequent mitochondrial outer membrane permeabilization, cytochrome c release, loss of membrane potential, and accumulation of reactive oxygen species (ROS). Pharmacologic inhibition of BAX oligomerization with BAX inhibitor peptide V5 (BipV5) preserved mitochondrial integrity, reduced ROS, and limited hair-cell loss in vitro. In vivo, repeated transtympanic delivery of BipV5 conferred sustained functional protection and reduced outer hair cell loss. These findings establish a structural mechanism linking mitochondrial membrane rupture to redox imbalance and apoptosis in cochlear hair cells and nominate BAX oligomerization as a drug-targetable node for preventing cisplatin ototoxicity without compromising anticancer therapy.