ABR/DPOAE publications

To identify small molecules that shield mammalian sensory hair cells from the ototoxic side effects of aminoglycoside antibiotics, 10,240 compounds were initially screened in zebrafish larvae, selecting for those that protected lateral-line hair cells against neomycin and gentamicin. When the 64 hits from this screen were retested in mouse cochlear cultures, 8 protected outer hair cells (OHCs) from gentamicin in vitro without causing hair-bundle damage. These 8 hits shared structural features and blocked, to varying degrees, the OHC’s mechano-electrical transducer (MET) channel, a route of aminoglycoside entry into hair cells. Further characterization of one of the strongest MET channel blockers, UoS-7692, revealed it additionally protected against kanamycin and tobramycin and did not abrogate the bactericidal activity of gentamicin. UoS-7692 behaved, like the aminoglycosides, as a permeant blocker of the MET channel; significantly reduced gentamicin–Texas red loading into OHCs; and preserved lateral-line function in neomycin-treated zebrafish. Transtympanic injection of UoS-7692 protected mouse OHCs from furosemide/kanamycin exposure in vivo and partially preserved hearing. The results confirmed the hair-cell MET channel as a viable target for the identification of compounds that protect the cochlea from aminoglycosides and provide a series of hit compounds that will inform the design of future otoprotectants.

Our previous study demonstrated rapamycin added to diet at 4 months of age had significantly less age-related outer hair cell loss in the basal half of the cochlea at 22 months of age compared to mice without rapamycin. The present study tested adding rapamycin to diet later in life, at 14 months of age, and added a longitudinal assessment of auditory brain stem response (ABR). The present study used UMHET4 mice, a 4 way cross in which all grandparental strains lack the Cdh23 753A allele that predisposes to early onset, progressive hearing loss. UMHET4 mice typically have normal hearing until 16–17 months, then exhibit threshold shifts at low frequencies/apical cochlea and later in more basal high frequency regions. ABR thresholds at 4, 12, 24, and 48 kHz were assessed at 12, 18, and 24 months of age and compared to baseline ABR thresholds acquired at 5 months of age to determine threshold shifts (TS). There was no TS at 12 months of age at any frequency tested. At 18 months of age mice with rapamycin added to diet at 14 months had a significantly lower mean TS at 4 and 12 kHz compared to mice on control diet with no significant difference at 24 and 48 kHz. At 24 months of age, the mean 4 kHz TS in rapamycin diet group was no longer significantly lower than the control diet group, while the 12 kHz mean remained significantly lower. Mean TS at 24 and 48 kHz in the rapamycin diet group became significantly lower than in the control diet group at 24 months. Hair cell counts at 24 months showed large loss in the apical half of most rapamycin and control diet mice cochleae with no significant difference between groups. There was only mild outer hair cell loss in the basal half of rapamycin and control diet mice cochleae with no significant difference between groups. The results show that a later life addition of rapamycin can decrease age-related hearing loss in the mouse model, however, it also suggests that this decrease is a delay/deceleration rather than a complete prevention.

Age-related hearing loss (ARHL) is the most common form of hearing loss and the predominant neurodegenerative disease associated with aging. Sirtuin 1 (SIRT1) is associated with the most complex physiological processes, including metabolism, cancer onset, and aging. SIRT1 protein levels are enhanced by the conversion of nicotinamide to N 1 -methylnicotinamide (MNAM), independent of its mRNA levels. Moreover, MNAM has implications in increased longevity achieved through its mitohormetic effects. Nicotinamide N-methyltransferase (Nnmt) is an enzyme involved in MNAM metabolism, and its level increases under caloric restriction (CR) conditions. The CR condition has implications in delaying ARHL onset. In this study, we aimed to determine the relationship between diet, hearing function, SIRT1 and SIRT3 expression levels in the inner ear, and cochlear morphology. Mice fed with a high-fat diet (HFD), HFD + 1% MNAM, and low-fat diet (LFD) were monitored for age-related auditory-evoked brainstem responses, and changes in cochlear histology, metabolism, and protein and mRNA expressions were analyzed. Our results revealed that the HFD- and aging-mediated downregulated expression of SIRT1 and SIRT3 promoted hearing loss that was obfuscated by MNAM supplementation-induced upregulated expression of cochlear SIRT1 and SIRT3. Thus, our results suggest that MNAM can be used as a therapeutic agent for preventing ARHL.

Transmembrane channel-like protein isoform 1 (TMC1) is a major component of the mechano-electrical transducer (MET) channel in cochlear hair cells and is subject to numerous mutations causing deafness. We report a new dominant human deafness mutation, TMC1 p.T422K, and have characterized the homologous mouse mutant, Tmc1 p.T416K, which caused deafness and outer hair cell (OHC) loss by the fourth postnatal week. MET channels showed decreased Ca 2+ permeability and resting open probability, but no change in single-channel conductance or expression. Three adjacent deafness mutations are TMC1 p.L416R, p.G417R, and p.M418K, the last homologous to the mouse Beethoven that exhibits similar channel effects. All substitute a positive for a neutral residue, which could produce charge screening in the channel pore or influence binding of an accessory subunit. Channel properties were compared in mice of both sexes between dominant ( Tmc1 p.T416K, Tmc1 p.D569N) and recessive ( Tmc1 p.W554L, Tmc1 p.D528N) mutations of residues near the putative pore of the channel. Tmc1 p.W554L and p.D569N exhibit reduced maximum current with no effect on single-channel conductance, implying a smaller number of channels transported to the stereociliary tips; this may stem from impaired TMC1 binding to LHFPL5. Tmc1 p.D528N, located in the pore's narrowest region, uniquely caused large reductions in MET channel conductance and block by dihydrostreptomycin (DHS). For Tmc1 p.T416K and Tmc1 p.D528N, transduction loss occurred between P15 and P20. We propose two mechanisms linking channel mutations and deafness: decreased Ca 2+ permeability, common to all mutants, and decreased resting open probability in low Ca 2+ , confined to dominant mutations. SIGNIFICANCE STATEMENT Transmembrane channel-like protein isoform 1 (TMC1) is thought to be a major component of the mechanotransducer channel in auditory hair cells, but the protein organization and channel structure are still uncertain. We made four mouse lines harboring Tmc1 point mutations that alter channel properties, causing hair cell degeneration and deafness. These include a mouse homolog of a new human deafness mutation pT416K that decreased channel Ca 2+ permeability by introducing a positively-charged amino acid in the putative pore. All mutations are consistent with the channel structure predicted from modeling, but only one, p.D528N near the external face of the pore, substantially reduced channel conductance and Ca 2+ permeability and virtually abolished block by dihydrostreptomycin (DHS), strongly endorsing its siting within the pore.

Striatin-interacting protein 1 ( Strip1 ) is a core component of the striatin interacting phosphatase and kinase (STRIPAK) complex, which is involved in embryogenesis and development, circadian rhythms, type 2 diabetes, and cancer progression. However, the expression and role of Strip1 in the mammalian cochlea remains unclear. Here we studied the expression and function of Strip1 in the mouse cochlea by using Strip1 knockout mice. We first found that the mRNA and protein expression of Strip1 increases as mice age starting from postnatal day (P) 3 and reaches its highest expression level at P30 and that the expression of Strip1 can be detected by immunofluorescent staining starting from P14 only in cochlear HCs, and not in supporting cells (SCs). Next, we crossed Strip1 heterozygous knockout ( Strip +/−) mice to obtain Strip1 homozygous knockout ( Strip1 −/−) mice for studying the role of Strip1 in cochlear HCs. However, no Strip1 −/− mice were obtained and the ratio of Strip +/− to Strip1 +/+ mice per litter was about 2:1, which suggested that homozygous Strip1 knockout is embryonic lethal. We measured hearing function and counted the HC number in P30 and P60 Strip +/− mice and found that they had normal hearing ability and HC numbers compared to Strip1 +/+ mice. Our study suggested that Strip1 probably play important roles in HC development and maturation, which needs further study in the future.

Noise-induced hearing loss (NIHL) is a common inner ear disease but has complex pathological mechanisms, one of which is increased oxidative stress in the cochlea. The high-mobility group box 1 (HMGB1) protein acts as an inflammatory mediator and shows different activities with redox modifications linked to the generation of reactive oxygen species (ROS). We aimed to investigate whether manipulation of cochlear HMGB1 during noise exposure could prevent noise-induced oxidative stress and hearing loss. Sixty CBA/CaJ mice were divided into two groups. An intraperitoneal injection of anti-HMGB1 antibodies was administered to the experimental group; the control group was injected with saline. Thirty minutes later, all mice were subjected to white noise exposure. Subsequent cochlear damage, including auditory threshold shifts, hair cell loss, expression of cochlear HMGB1, and free radical activity, was then evaluated. The levels of HMGB1 and 4-hydroxynonenal (4-HNE), as respective markers of reactive nitrogen species (RNS) and ROS formation, showed slight increases on post-exposure day 1 and achieved their highest levels on post-exposure day 4. After noise exposure, the antibody-treated mice showed markedly less ROS formation and lower expression of NADPH oxidase 4 (NOX4), nitrotyrosine, inducible nitric oxide synthase (iNOS), and intercellular adhesion molecule-1 (ICAM-1) than the saline-treated control mice. A significant amelioration was also observed in the threshold shifts of the auditory brainstem response and the loss of outer hair cells in the antibody-treated versus the saline-treated mice. Our results suggest that inhibition of HMGB1 by neutralization with anti-HMGB1 antibodies prior to noise exposure effectively attenuated oxidative stress and subsequent inflammation. This procedure could therefore have potential as a therapy for NIHL.

Blast trauma is a common acoustic/physical insult occurring in modern warfare. Twenty percent of active duty military come into close proximity to explosions and experience mild to severe sensory deficits. The prevalence of such injuries is high but correlating auditory sensitivity changes with the initial insult is difficult because injury and evaluations are often separated by long time periods. Here, auditory sensitivity was measured before and after a traumatic blast in adult CBA/CaJ mice using auditory brainstem responses, distortion production otoacoustic emissions, and behavioral detection of pure tones. These measurements included baseline auditory sensitivity prior to injury in all mice, and again at 3, 30, and 90 days after the blast in the two physiological groups, and daily for up to 90 days in the behavioral group. Mice in all groups experienced an initial deterioration in auditory sensitivity, though physiological measurements showed evidence of recovery that behavioral measurements did not. Amplitudes and latencies of ABR waves may reflect additional changes beyond the peripheral damage shown by the threshold changes and should be explored further. The present work addresses a major gap in the current acoustic trauma literature both in terms of comparing physiological and behavioral methods, as well as measuring the time course of recovery.

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.

Numerous studies indicate that deficits in the proper integration or migration of specific GABAergic precursor cells from the subpallium to the cortex can lead to severe cognitive dysfunctions and neurodevelopmental pathogenesis linked to intellectual disabilities. A different set of GABAergic precursors cells that express Pax2 migrate to hindbrain regions, targeting, for example auditory or somatosensory brainstem regions. We demonstrate that the absence of BDNF in Pax2-lineage descendants of Bdnf Pax 2 KOs causes severe cognitive disabilities. In Bdnf Pax 2 KOs, a normal number of parvalbumin-positive interneurons ( PV-IN s) was found in the auditory cortex ( AC ) and hippocampal regions, which went hand in hand with reduced PV-labeling in neuropil domains and elevated activity-regulated cytoskeleton-associated protein ( Arc/Arg3.1 ; here: Arc ) levels in pyramidal neurons in these same regions. This immaturity in the inhibitory/excitatory balance of the AC and hippocampus was accompanied by elevated LTP, reduced (sound-induced) LTP/LTD adjustment, impaired learning, elevated anxiety, and deficits in social behavior, overall representing an autistic-like phenotype. Reduced tonic inhibitory strength and elevated spontaneous firing rates in dorsal cochlear nucleus ( DCN ) brainstem neurons in otherwise nearly normal hearing Bdnf Pax 2 KOs suggests that diminished fine-grained auditory-specific brainstem activity has hampered activity-driven integration of inhibitory networks of the AC in functional (hippocampal) circuits. This leads to an inability to scale hippocampal post-synapses during LTP/LTD plasticity. BDNF in Pax2-lineage descendants in lower brain regions should thus be considered as a novel candidate for contributing to the development of brain disorders, including autism.

Individual estimates of cochlear compression may provide complementary information to traditional audiometric hearing thresholds in disentangling different types of peripheral cochlear damage. Here we investigated the use of the slope of envelope following response (EFR) magnitude-level functions obtained from four simultaneously presented amplitude modulated tones with modulation frequencies of 80–100 Hz as a proxy of peripheral level compression. Compression estimates in individual normal hearing (NH) listeners were consistent with previously reported group-averaged compression estimates based on psychoacoustical and distortion-product oto-acoustic emission (DPOAE) measures in human listeners. They were also similar to basilar membrane (BM) compression values measured invasively in non-human mammals. EFR-based compression estimates in hearing-impaired listeners were less compressive than those for the NH listeners, consistent with a reduction of BM compression. Cochlear compression was also estimated using DPOAEs in the same NH listeners. DPOAE estimates were larger (less compressive) than EFRs estimates, showing no correlation. Despite the numerical concordance between EFR-based compression estimates and group-averaged estimates from other methods, simulations using an auditory nerve (AN) model revealed that compression estimates based on EFRs might be highly influenced by contributions from off-characteristic frequency (CF) neural populations. This compromises the possibility to estimate on-CF (i.e., frequency-specific or “local”) peripheral level compression with EFRs.

BACKGROUND: Sound normally enters the ear canal, passes through the middle ear, and stimulates the cochlea through the oval window. Alternatively, the cochlea can be stimulated in a reverse manner, namely round window stimulation. The reverse stimulation is not well understood, partly because in classic lumped-parameter models the path of reverse drive during the round window stimulation is usually not considered. OBJECTIVE: The study goal is to gain a better understanding of the hearing mechanism during round window stimulation. METHODS: A piezo actuator was coupled to the oval and round window of the guinea pigs. The auditory brainstem response produced by the forward and reverse stimulation at four frequencies was recorded. RESULTS: The results show that the input voltage of the actuator required at the hearing threshold in the round window drive was higher than that in the oval window drive. In order to understand the data, we designed a lumped-parameter cochlear model that can simulate both forward and reverse drive. The model-predicted results were consistent with the experimental results. CONCLUSIONS: The response of the auditory system to stimulus of oval window and round window was quantified through animal experimentation, and guinea pigs were used as experimental animals. When the same stimulus was applied to the oval window and round window of the cochlea, the ABR signals were compared. A lumped parameter model was designed to incorporate the sound transmission paths in both oval and round window stimulation. The simulated results are consistent with those of animal experiments. This model will be useful in understanding the inner-ear response in round window.

The afferent synapses between inner hair cells (IHC) and spiral ganglion neurons are specialized to faithfully encode sound with sub-millisecond precision over prolonged periods of time. Here, we studied the role of Rab3 interacting molecule-binding proteins (RIM-BP) 1 and 2 – multidomain proteins of the active zone known to directly interact with RIMs, Bassoon and Ca V 1.3 – in IHC presynaptic function and hearing. Recordings of auditory brainstem responses and otoacoustic emissions revealed that genetic disruption of RIM-BPs 1 and 2 in mice ( RIM-BP1/2 –/– ) causes a synaptopathic hearing impairment exceeding that found in mice lacking RIM-BP2 ( RIM-BP2 –/– ). Patch-clamp recordings from RIM-BP1/2 –/– IHCs indicated a subtle impairment of exocytosis from the readily releasable pool of synaptic vesicles that had not been observed in RIM-BP2 –/– IHCs. In contrast, the reduction of Ca 2+ -influx and sustained exocytosis was similar to that in RIMBP2 –/– IHCs. We conclude that both RIM-BPs are required for normal sound encoding at the IHC synapse, whereby RIM-BP2 seems to take the leading role.

Pituitary adenylyl cyclase-activating polypeptide (PACAP) is a member of the vasoactive intestinal polypeptide (VIP)-the secretin-glucagon family of neuropeptides. They act through two classes of receptors: PACAP type 1 (PAC1) and type 2 (VPAC1 and VPAC2). Among their pleiotropic effects throughout the body, PACAP functions as neuromodulators and neuroprotectors, rescuing neurons from apoptosis, mostly through the PAC1 receptor. To explore the potential protective effect of endogenous PACAP against Noise-induced hearing loss (NIHL), we used a knockout mouse model lacking PAC1 receptor expression (PACR1 −/− ) and a transgenic humanized mouse model expressing the human PAC1 receptor (TgHPAC1R). Based on complementary approaches combining electrophysiological, histochemical, and molecular biological evaluations, we show PAC1R expression in spiral ganglion neurons and in cochlear apical cells of the organ of Corti. Wild-type (WT), PAC1R −/− , and TgHPAC1R mice exhibit similar auditory thresholds. For most of the frequencies tested after acute noise damage, however, PAC1R −/− mice showed a larger elevation of the auditory threshold than did their WT counterparts. By contrast, in a transgene copy number-dependent fashion, TgHPAC1R mice showed smaller noise-induced elevations of auditory thresholds compared to their WT counterparts. Together, these findings suggest that PACAP could be a candidate for endogenous protection against noise-induced hearing loss.

Background The SCN11A gene, encoded Nav1.9 TTX resistant sodium channels, is a main effector in peripheral inflammation related pain in nociceptive neurons. The role of SCN11A gene in the auditory system has not been well characterized. We therefore examined the expression of SCN11A in the murine cochlea, the morphological and physiological features of Nav1.9 knockout (KO) ICR mice. Results Nav1.9 expression was found in the primary afferent endings beneath the inner hair cells (IHCs). The relative quantitative expression of Nav1.9 mRNA in modiolus of wild-type (WT) mice remains unchanged from P0 to P60. The number of presynaptic CtBP2 puncta in Nav1.9 KO mice was significantly lower than WT. In addition, the number of SGNs in Nav1.9 KO mice was also less than WT in the basal turn, but not in the apical and middle turns. There was no lesion in the somas and stereocilia of hair cells in Nav1.9 KO mice. Furthermore, Nav1.9 KO mice showed higher and progressive elevated ABR threshold at 16 kHz, and a significant increase in CAP thresholds. Conclusions These data suggest a role of Nav1.9 in regulating the function of ribbon synapses and the auditory nerves. The impairment induced by Nav1.9 gene deletion mimics the characters of cochlear synaptopathy.

Background Aging, noise, infection, and ototoxic drugs are the major causes of human acquired sensorineural hearing loss, but treatment options are limited. CRISPR/Cas9 technology has tremendous potential to become a new therapeutic modality for acquired non-inherited sensorineural hearing loss. Here, we develop CRISPR/Cas9 strategies to prevent aminoglycoside-induced deafness, a common type of acquired non-inherited sensorineural hearing loss, via disrupting the Htra2 gene in the inner ear which is involved in apoptosis but has not been investigated in cochlear hair cell protection. Results The results indicate that adeno-associated virus (AAV)-mediated delivery of CRISPR/SpCas9 system ameliorates neomycin-induced apoptosis, promotes hair cell survival, and significantly improves hearing function in neomycin-treated mice. The protective effect of the AAV–CRISPR/Cas9 system in vivo is sustained up to 8 weeks after neomycin exposure. For more efficient delivery of the whole CRISPR/Cas9 system, we also explore the AAV–CRISPR/SaCas9 system to prevent neomycin-induced deafness. The in vivo editing efficiency of the SaCas9 system is 1.73% on average. We observed significant improvement in auditory brainstem response thresholds in the injected ears compared with the non-injected ears. At 4 weeks after neomycin exposure, the protective effect of the AAV–CRISPR/SaCas9 system is still obvious, with the improvement in auditory brainstem response threshold up to 50 dB at 8 kHz. Conclusions These findings demonstrate the safe and effective prevention of aminoglycoside-induced deafness via Htra2 gene editing and support further development of the CRISPR/Cas9 technology in the treatment of non-inherited hearing loss as well as other non-inherited diseases.

DBA/2 mice are a well-known animal model for hearing loss developed due to intrinsic properties of these animals. However, results on the phenotype of hearing loss in DBA/2 mice have been mainly reported at an early stage in mice aged ≤7 weeks. Instead, the present study evaluated the hearing ability at 5, 13, and 34 weeks of age using DBA/2korl mice. Auditory brainstem response test was performed at 8–32 KHz at 5, 13, and 34 weeks of age, and hearing loss was confirmed to be induced in a time-dependent manner. In addition, histopathological evaluation at the same age confirmed the morphological damage of the cochlea. The findings presented herein are the results of the long-term observation of the phenotype of hearing loss in DBA/2 mice and can be useful in studies related to aging-dependent hearing loss.

Background Noise-induced hearing loss represents a commonly diagnosed type of hearing disability, severely impacting the quality of life of individuals. The current work is aimed at assessing the effects of DNA methylation on noise-induced hearing loss. Methods Blocking DNA methyltransferase 1 (DNMT1) activity with a selective inhibitor RG108 or silencing DNMT1 with siRNA was used in this study. Auditory brainstem responses were measured at baseline and 2 days after trauma in mice to assess auditory functions. Whole-mount immunofluorescent staining and confocal microcopy of mouse inner ear specimens were performed to analyze noise-induced damage in cochleae and the auditory nerve at 2 days after noise exposure. Results The results showed that noise exposure caused threshold elevation of auditory brainstem responses and cochlear hair cell loss. Whole-mount cochlea staining revealed a reduction in the density of auditory ribbon synapses between inner hair cells and spiral ganglion neurons. Inhibition of DNA methyltransferase activity via a non-nucleoside specific pharmacological inhibitor, RG108, or silencing of DNA methyltransferase-1 with siRNA significantly attenuated ABR threshold elevation, hair cell damage, and the loss of auditory synapses. Conclusions This study suggests that inhibition of DNMT1 ameliorates noise-induced hearing loss and indicates that DNMT1 may be a promising therapeutic target. Graphical abstract

Learning of new auditory stimuli often requires repetitive exposure to the stimulus. Fast and implicit learning of sounds presented at random times enables efficient auditory perception. However, it is unclear how such sensory encoding is processed on a neural level. We investigated neural responses that are developed from a passive, repetitive exposure to a specific sound in the auditory cortex of anesthetized rats, using electrocorticography. We presented a series of random sequences that are generated afresh each time, except for a specific reference sequence that remains constant and re-appears at random times across trials. We compared induced activity amplitudes between reference and fresh sequences. Neural responses from both primary and non-primary auditory cortical regions showed significantly decreased induced activity amplitudes for reference sequences compared to fresh sequences, especially in the beta band. This is the first study showing that neural correlates of auditory pattern learning can be evoked even in anesthetized, passive listening animal models.

Attenuation of noise-induced hair cell loss and noise-induced hearing loss (NIHL) by treatment with FK506 (tacrolimus), a calcineurin (CaN/PP2B) inhibitor used clinically as an immunosuppressant, has been previously reported, but the downstream mechanisms of FK506-attenuated NIHL remain unknown. Here we showed that CaN immunolabeling in outer hair cells (OHCs) and nuclear factor of activated T-cells isoform c4 (NFATc4/NFAT3) in OHC nuclei are significantly increased after moderate noise exposure in adult CBA/J mice. Consequently, treatment with FK506 significantly reduces moderate-noise-induced loss of OHCs and NIHL. Furthermore, induction of reactive oxygen species (ROS) by moderate noise was significantly diminished by treatment with FK506. In agreement with our previous finding that autophagy marker microtubule-associated protein light chain 3B (LC3B) does not change in OHCs under conditions of moderate-noise-induced permanent threshold shifts, treatment with FK506 increases LC3B immunolabeling in OHCs after exposure to moderate noise. Additionally, prevention of NIHL by treatment with FK506 was partially abolished by pretreatment with LC3B small interfering RNA. Taken together, these results indicate that attenuation of moderate-noise-induced OHC loss and hearing loss by FK506 treatment occurs not only via inhibition of CaN activity but also through inhibition of ROS and activation of autophagy.

Throughout life, sensory systems adapt to the sensory environment to provide optimal responses to relevant tasks. In the case of a developing system, sensory inputs induce changes that are permanent and detectable up to adulthood. Previously, we have shown that rearing rat pups in a complex acoustic environment (spectrally and temporally modulated sound) from postnatal day 14 (P14) to P28 permanently improves the response characteristics of neurons in the inferior colliculus and auditory cortex, influencing tonotopical arrangement, response thresholds and strength, and frequency selectivity, along with stochasticity and the reproducibility of neuronal spiking patterns. In this study, we used a set of behavioral tests based on a recording of the acoustic startle response (ASR) and its prepulse inhibition (PPI), with the aim to extend the evidence of the persistent beneficial effects of the developmental acoustical enrichment. The enriched animals were generally not more sensitive to startling sounds, and also, their PPI of ASR, induced by noise or pure tone pulses, was comparable to the controls. They did, however, exhibit a more pronounced PPI when the prepulse stimulus was represented either by a change in the frequency of a background tone or by a silent gap in background noise. The differences in the PPI of ASR between the enriched and control animals were significant at lower (55 dB SPL), but not at higher (65-75 dB SPL), intensities of background sound. Thus, rearing pups in the acoustically enriched environment led to an improvement of the frequency resolution and gap detection ability under more difficult testing conditions, i.e., with a worsened stimulus clarity. We confirmed, using behavioral tests, that an acoustically enriched environment during the critical period of development influences the frequency and temporal processing in the auditory system, and these changes persist until adulthood.