Auditory Neurophysiology publications

Auditory brain areas undergo reorganization resulting from abnormal sensory input during early postnatal development. This is evident from studies at the cortical level but it remains unclear whether there is reorganization in the auditory midbrain in a species similar to the human, that is, with early hearing onset. We have explored midbrain plasticity in the chinchilla, a precocious species that matches the human in terms of hearing development. Neonatal chinchillas were chronically exposed to a 2 kHz narrowband sound at 70 dB SPL for 4 weeks. Tonotopic maps in inferior colliculus (central nucleus) were defined based on single neuron characteristic frequency. We hypothesized an overrepresentation of the 2 kHz region of the maps. However, we observed a significant decrease in the proportion of neurons dedicated to the 2 kHz octave band and also away from the exposure frequency at 8 kHz. In addition, we report a significant increase in low frequency representation (<1 kHz), again a change to tonotopic mapping distant to the 2 kHz region. Thus in a precocious species, tonotopic maps in auditory midbrain are altered following abnormal stimulation during development. However, these changes are more complex than the overrepresentation of exposure related frequency regions that are often reported.

Some combinations of musical tones sound pleasing to Western listeners, and are termed consonant, while others sound discordant, and are termed dissonant. The perceptual phenomenon of consonance has been traced to the acoustic property of harmonicity. It has been repeatedly shown that neural correlates of consonance can be found as early as the auditory brainstem as reflected in the harmonicity of the scalp-recorded frequency-following response (FFR). "Neural Pitch Salience" (NPS) measured from FFRs-essentially a time-domain equivalent of the classic pattern recognition models of pitch-has been found to correlate with behavioral judgments of consonance for synthetic stimuli. Following the idea that the auditory system has evolved to process behaviorally relevant natural sounds, and in order to test the generalizability of this finding made with synthetic tones, we recorded FFRs for consonant and dissonant intervals composed of synthetic and natural stimuli. We found that NPS correlated with behavioral judgments of consonance and dissonance for synthetic but not for naturalistic sounds. These results suggest that while some form of harmonicity can be computed from the auditory brainstem response, the general percept of consonance and dissonance is not captured by this measure. It might either be represented in the brainstem in a different code (such as place code) or arise at higher levels of the auditory pathway. Our findings further illustrate the importance of using natural sounds, as a complementary tool to fully-controlled synthetic sounds, when probing auditory perception.

Identifying sounds is critical for an animal to make appropriate behavioral responses to environmental stimuli, including vocalizations from conspecifics. Identification of vocalizations may be supported by neuronal selectivity in the auditory pathway. The first place in the ascending auditory pathway where neuronal selectivity to vocalizations has been found is in the inferior colliculus (IC), but very few brainstem nuclei have been evaluated. Here, we tested whether selectivity to vocalizations is present in the dorsal cochlear nucleus (DCN). We recorded extracellular neural responses in the DCN of mice and found that fusiform cells responded in a heterogeneous and selective manner to mouse ultrasonic vocalizations. Most fusiform cells responded to vocalizations that contained spectral energy at much higher frequencies than the characteristic frequencies of the cells. To understand this mismatch of stimulus properties and frequency tuning of the cells, we developed a dynamic, nonlinear model of the cochlea that simulates cochlear distortion products on the basilar membrane. We preprocessed the vocalization stimuli through this model and compared responses to these distorted vocalizations with responses to the original vocalizations. We found that fusiform cells in the DCN respond in a heterogeneous manner to vocalizations, and that these neurons can use distortion products as a mechanism for encoding ultrasonic vocalizations. In addition, the selective neuronal responses were dependent on the presence of inhibitory sidebands that modulated the response depending on the temporal structure of the distortion product. These findings suggest that important processing of complex sounds occurs at a very early stage of central auditory processing and is not strictly a function of the cortex.

Stream segregation enables a listener to disentangle multiple competing sequences of sounds. A recent study from our laboratory demonstrated that cortical neurons in anesthetized cats exhibit spatial stream segregation (SSS) by synchronizing preferentially to one of two sequences of noise bursts that alternate between two source locations. Here, we examine the emergence of SSS along the ascending auditory pathway. Extracellular recordings were made in anesthetized rats from the inferior colliculus (IC), the nucleus of the brachium of the IC (BIN), the medial geniculate body (MGB), and the primary auditory cortex (A1). Stimuli consisted of interleaved sequences of broadband noise bursts that alternated between two source locations. At stimulus presentation rates of 5 and 10 bursts per second, at which human listeners report robust SSS, neural SSS is weak in the central nucleus of the IC (ICC), it appears in the nucleus of the brachium of the IC (BIN) and in approximately two-thirds of neurons in the ventral MGB (MGBv), and is prominent throughout A1. The enhancement of SSS at the cortical level reflects both increased spatial sensitivity and increased forward suppression. We demonstrate that forward suppression in A1 does not result from synaptic inhibition at the cortical level. Instead, forward suppression might reflect synaptic depression in the thalamocortical projection. Together, our findings indicate that auditory streams are increasingly segregated along the ascending auditory pathway as distinct mutually synchronized neural populations. SIGNIFICANCE STATEMENT Listeners are capable of disentangling multiple competing sequences of sounds that originate from distinct sources. This stream segregation is aided by differences in spatial location between the sources. A possible substrate of spatial stream segregation (SSS) has been described in the auditory cortex, but the mechanisms leading to those cortical responses are unknown. Here, we investigated SSS in three levels of the ascending auditory pathway with extracellular unit recordings in anesthetized rats. We found that neural SSS emerges within the ascending auditory pathway as a consequence of sharpening of spatial sensitivity and increasing forward suppression. Our results highlight brainstem mechanisms that culminate in SSS at the level of the auditory cortex.

The auditory cortex is necessary for sound localization. The mechanisms that shape bicoordinate spatial representation in the auditory cortex remain unclear. Here, we addressed this issue by quantifying spatial receptive fields (SRFs) in two functionally distinct cortical regions in the pallid bat. The pallid bat uses echolocation for obstacle avoidance and listens to prey-generated noise to localize prey. Its cortex contains two segregated regions of response selectivity that serve echolocation and localization of prey-generated noise. The main aim of this study was to compare 2D SRFs between neurons in the noise-selective region (NSR) and the echolocation region [frequency-modulated sweep-selective region (FMSR)]. The data reveal the following major differences between these two regions: (1) compared with NSR neurons, SRF properties of FMSR neurons were more strongly dependent on sound level; (2) as a population, NSR neurons represent a broad region of contralateral space, while FMSR selectivity was focused near the midline at sound levels near threshold and expanded considerably with increasing sound levels; and (3) the SRF size and centroid elevation were correlated with the characteristic frequency in the NSR, but not the FMSR. These data suggest different mechanisms of sound localization for two different behaviors. Previously, we reported that azimuth is represented by predictable changes in the extent of activated cortex. The present data indicate how elevation constrains this activity pattern. These data suggest a novel model for bicoordinate spatial representation that is based on the extent of activated cortex resulting from the overlap of binaural and tonotopic maps. SIGNIFICANCE STATEMENT Unlike the visual and somatosensory systems, spatial information is not directly represented at the sensory receptor epithelium in the auditory system. Spatial locations are computed by integrating neural binaural properties and frequency-dependent pinna filtering, providing a useful model to study how neural properties and peripheral structures are adapted for sensory encoding. Although auditory cortex is necessary for sound localization, our understanding of how the cortex represents space remains rudimentary. Here we show that two functionally distinct regions of the pallid bat auditory cortex represent 2D space using different mechanisms. In addition, we suggest a novel hypothesis on how the nature of overlap between systematic maps of binaural and frequency selectivity leads to representation of both azimuth and elevation.

Earlier studies demonstrate that when the frequency of rhythmic tone sequences or streams is task relevant, ongoing excitability fluctuations (oscillations) of neuronal ensembles in primary auditory cortex (A1) entrain to stimulation in a frequency dependent way that sharpens frequency tuning. The phase distribution across A1 neuronal ensembles at time points when attended stimuli are predicted to occur reflects the focus of attention along the spectral attribute of auditory stimuli. This study examined how neuronal activity is modulated if only the temporal features of rhythmic stimulus streams are relevant. We presented macaques with auditory clicks arranged in 33 Hz (gamma timescale) quintets, repeated at a 1.6 Hz (delta timescale) rate. Such multi-scale, hierarchically organized temporal structure is characteristic of vocalizations and other natural stimuli. Monkeys were required to detect and respond to deviations in the temporal pattern of gamma quintets. As expected, engagement in the auditory task resulted in the multi-scale entrainment of delta- and gamma-band neuronal oscillations across all of A1. Surprisingly, however, the phase-alignment, and thus, the physiological impact of entrainment differed across the tonotopic map in A1. In the region of 11-16 kHz representation, entrainment most often aligned high excitability oscillatory phases with task-relevant events in the input stream and thus resulted in response enhancement. In the remainder of the A1 sites, entrainment generally resulted in response suppression. Our data indicate that the suppressive effects were due to low excitability phase delta oscillatory entrainment and the phase amplitude coupling of delta and gamma oscillations. Regardless of the phase or frequency, entrainment appeared stronger in left A1, indicative of the hemispheric lateralization of auditory function.

Significance Ionotropic glutamate AMPA receptors (AMPARs) play a fundamental role in normal function and plasticity of the brain, and they are also involved in many brain disorders. Despite the central role of AMPARs in neurobiology, the modulation of synaptic AMPA responses by endogenous modulators remains not well understood. Here, in three synapses found in two different brain areas, we provide the first evidence, to our knowledge, that endogenous zinc is coreleased with glutamate and modulates the strength of synaptic AMPAR responses. Because in many neocortical areas more than 50% of excitatory presynaptic terminals contain zinc within their glutamatergic vesicles, our findings establish zinc as a general neuromodulator that allows for fine-tuning and plasticity of glutamatergic fast synaptic transmission in the brain.

The ability to discriminate tones of different frequencies is fundamentally important for everyday hearing. While neurons in the primary auditory cortex (AC) respond differentially to tones of different frequencies, whether and how AC regulates auditory behaviors that rely on frequency discrimination remains poorly understood. Here, we find that the level of activity of inhibitory neurons in AC controls frequency specificity in innate and learned auditory behaviors that rely on frequency discrimination. Photoactivation of parvalbumin-positive interneurons (PVs) improved the ability of the mouse to detect a shift in tone frequency, whereas photosuppression of PVs impaired the performance. Furthermore, photosuppression of PVs during discriminative auditory fear conditioning increased generalization of conditioned response across tone frequencies, whereas PV photoactivation preserved normal specificity of learning. The observed changes in behavioral performance were correlated with bidirectional changes in the magnitude of tone-evoked responses, consistent with predictions of a model of a coupled excitatory-inhibitory cortical network. Direct photoactivation of excitatory neurons, which did not change tone-evoked response magnitude, did not affect behavioral performance in either task. Our results identify a new function for inhibition in the auditory cortex, demonstrating that it can improve or impair acuity of innate and learned auditory behaviors that rely on frequency discrimination.

The A1555G mutation in the 12S rRNA gene of human mitochondrial DNA causes maternally inherited, nonsyndromic deafness, an extreme case of tissue-specific mitochondrial pathology. A transgenic mouse strain that robustly overexpresses the mitochondrial 12S ribosomal RNA methyltransferase TFB1M (Tg-mtTFB1 mice) exhibits progressive hearing loss that we proposed models aspects of A1555G-related pathology in humans. Although our previous studies of Tg-mtTFB1 mice implicated apoptosis in the spiral ganglion and stria vascularis because of mitochondrial reactive oxygen species-mediated activation of AMP kinase (AMPK) and the nuclear transcription factor E2F1, detailed auditory pathology was not delineated. Herein, we show that Tg-mtTFB1 mice have reduced endocochlear potential, indicative of significant stria vascularis dysfunction, but without obvious signs of strial atrophy. We also observed decreased auditory brainstem response peak 1 amplitude and prolonged wave I latency, consistent with apoptosis of spiral ganglion neurons. Although no major loss of hair cells was observed, there was a mild impairment of voltage-dependent electromotility of outer hair cells. On the basis of these results, we propose that these events conspire to produce the progressive hearing loss phenotype in Tg-mtTFB1 mice. Finally, genetically reducing AMPK α1 rescues hearing loss in Tg-mtTFB1 mice, confirming that aberrant up-regulation of AMPK signaling promotes the observed auditory pathology. The relevance of these findings to human A1555G patients and the potential therapeutic value of reducing AMPK activity are discussed.

Central auditory circuits are influenced by the somatosensory system, a relationship that may underlie tinnitus generation. In the guinea pig dorsal cochlear nucleus (DCN), pairing spinal trigeminal nucleus (Sp5) stimulation with tones at specific intervals and orders facilitated or suppressed subsequent tone-evoked neural responses, reflecting spike timing-dependent plasticity (STDP). Furthermore, after noise-induced tinnitus, bimodal responses in DCN were shifted from Hebbian to anti-Hebbian timing rules with less discrete temporal windows, suggesting a role for bimodal plasticity in tinnitus. Here, we aimed to determine if multisensory STDP principles like those in DCN also exist in primary auditory cortex (A1), and whether they change following noise-induced tinnitus. Tone-evoked and spontaneous neural responses were recorded before and 15 min after bimodal stimulation in which the intervals and orders of auditory-somatosensory stimuli were randomized. Tone-evoked and spontaneous firing rates were influenced by the interval and order of the bimodal stimuli, and in sham-controls Hebbian-like timing rules predominated as was seen in DCN. In noise-exposed animals with and without tinnitus, timing rules shifted away from those found in sham-controls to more anti-Hebbian rules. Only those animals with evidence of tinnitus showed increased spontaneous firing rates, a purported neurophysiological correlate of tinnitus in A1. Together, these findings suggest that bimodal plasticity is also evident in A1 following noise damage and may have implications for tinnitus generation and therapeutic intervention across the central auditory circuit.

Animals require the ability to ignore sensory stimuli that have no consequence yet respond to the same stimuli when they become useful. However, the brain circuits that govern this flexibility in sensory processing are not well understood. Here we show in mouse primary auditory cortex (A1) that daily passive sound exposure causes a long-lasting reduction in representations of the experienced sound by layer 2/3 pyramidal cells. This habituation arises locally in A1 and involves an enhancement in inhibition and selective upregulation in the activity of somatostatin-expressing inhibitory neurons (SOM cells). Furthermore, when mice engage in sound-guided behavior, pyramidal cell excitatory responses to habituated sounds are enhanced, whereas SOM cell responses are diminished. Together, our results demonstrate the bidirectional modulation of A1 sensory representations and suggest that SOM cells gate cortical information flow based on the behavioral relevance of the stimulus.

There has been continued interest in clinical objective measures of binaural processing. One commonly proposed measure is the binaural interaction component (BIC), which is obtained typically by recording auditory brainstem responses (ABRs)—the BIC reflects the difference between the binaural ABR and the sum of the monaural ABRs (i.e., binaural − (left + right)). We have recently developed an alternative, direct measure of sensitivity to interaural time differences, namely, a following response to modulations in interaural phase difference (the interaural phase modulation following response; IPM-FR). To obtain this measure, an ongoing diotically amplitude-modulated signal is presented, and the interaural phase difference of the carrier is switched periodically at minima in the modulation cycle. Such periodic modulations to interaural phase difference can evoke a steady state following response. BIC and IPM-FR measurements were compared from 10 normal-hearing subjects using a 16-channel electroencephalographic system. Both ABRs and IPM-FRs were observed most clearly from similar electrode locations—differential recordings taken from electrodes near the ear (e.g., mastoid) in reference to a vertex electrode (Cz). Although all subjects displayed clear ABRs, the BIC was not reliably observed. In contrast, the IPM-FR typically elicited a robust and significant response. In addition, the IPM-FR measure required a considerably shorter recording session. As the IPM-FR magnitude varied with interaural phase difference modulation depth, it could potentially serve as a correlate of perceptual salience. Overall, the IPM-FR appears a more suitable clinical measure than the BIC.

Models of the active cochlea build upon the underlying passive mechanics. Passive cochlear mechanics is based on physical and geometrical properties of the cochlea and the fluid-tissue interaction between the cochlear partition and the surrounding fluid. Although the fluid-tissue interaction between the basilar membrane and the fluid in scala tympani (ST) has been explored in both active and passive cochleae, there was no experimental data on the fluid-tissue interaction on the scala media (SM) side of the partition. To this aim, we measured sound-evoked intracochlear pressure in SM close to the partition using micropressure sensors. All the SM pressure data are from passive cochleae, likely because the SM cochleostomy led to loss of endocochlear potential. Thus, these experiments are studies of passive cochlear mechanics. SM pressure close to the tissue showed a pattern of peaks and notches, which could be explained as an interaction between fast and slow (i.e., traveling wave) pressure modes. In several animals SM and ST pressure were measured in the same cochlea. Similar to previous studies, ST-pressure was dominated by a slow, traveling wave mode at stimulus frequencies in the vicinity of the best frequency of the measurement location, and by a fast mode above best frequency. Antisymmetric pressure between SM and ST supported the classic single-partition cochlear models, or a dual-partition model with tight coupling between partitions. From the SM and ST pressure we calculated slow and fast modes, and from active ST pressure we extrapolated the passive findings to the active case. The passive slow mode estimated from SM and ST data was low-pass in nature, as predicted by cochlear models.

In recently diverged gray treefrogs (Hyla chrysoscelis and H. versicolor), advertisement calls that differ primarily in pulse shape and pulse rate act as an important premating isolation mechanism. Temporally selective neurons in the anuran inferior colliculus may contribute to selective behavioral responses to these calls. Here we present in vivo extracellular and whole-cell recordings from long-interval-selective neurons (LINs) made during presentation of pulses that varied in shape and rate. Whole-cell recordings revealed that interplay between excitation and inhibition shapes long-interval selectivity. LINs in H. versicolor showed greater selectivity for slow-rise pulses, consistent with the slow-rise pulse characteristics of their calls. The steepness of pulse-rate tuning functions, but not the distributions of best pulse rates, differed between the species in a manner that depended on whether pulses had slow or fast-rise shape. When tested with stimuli representing the temporal structure of the advertisement calls of H. chrysoscelis or H. versicolor, approximately 27 % of LINs in H. versicolor responded exclusively to the latter stimulus type. The LINs of H. chrysoscelis were less selective. Encounter calls, which are produced at similar pulse rates in both species (≈5 pulses/s), are likely to be effective stimuli for the LINs of both species.

Auditory information relayed by auditory nerve fibers and somatosensory information relayed by granule cell parallel fibers converge on the fusiform cells (FCs) of the dorsal cochlear nucleus, the first brain station of the auditory pathway. In vitro, parallel fiber synapses on FCs exhibit spike-timing-dependent plasticity with Hebbian learning rules, partially mediated by the NMDA receptor (NMDAr). Well-timed bimodal auditory-somatosensory stimulation, in vivo equivalent of spike-timing-dependent plasticity, can induce stimulus-timing-dependent plasticity (StTDP) of the FCs spontaneous and tone-evoked firing rates. In healthy guinea pigs, the resulting distribution of StTDP learning rules across a FC neural population is dominated by a Hebbian profile while anti-Hebbian, suppressive and enhancing LRs are less frequent. In this study, we investigate in vivo, the NMDAr contribution to FC baseline activity and long term plasticity. We find that blocking the NMDAr decreases the synchronization of FC- spontaneous activity and mediates differential modulation of FC rate-level functions such that low, and high threshold units are more likely to increase, and decrease, respectively, their maximum amplitudes. Three significant alterations in mean learning-rule profiles were identified: transitions from an initial Hebbian profile towards (1) an anti-Hebbian; (2) a suppressive profile; and (3) transitions from an anti-Hebbian to a Hebbian profile. FC units preserving their learning rules showed instead, NMDAr-dependent plasticity to unimodal acoustic stimulation, with persistent depression of tone-evoked responses changing to persistent enhancement following the NMDAr antagonist. These results reveal a crucial role of the NMDAr in mediating FC baseline activity and long-term plasticity which have important implications for signal processing and auditory pathologies related to maladaptive plasticity of dorsal cochlear nucleus circuitry.

Autism is a neurodevelopmental disorder clinically characterized by deficits in communication, lack of social interaction and repetitive behaviors with restricted interests. A number of studies have reported that sensory perception abnormalities are common in autistic individuals and might contribute to the complex behavioral symptoms of the disorder. In this context, hearing incongruence is particularly prevalent. Considering that some of this abnormal processing might stem from the unbalance of inhibitory and excitatory drives in brain circuitries, we used an animal model of autism induced by valproic acid (VPA) during pregnancy in order to investigate the tonotopic organization of the primary auditory cortex (AI) and its local inhibitory circuitry. Our results show that VPA rats have distorted primary auditory maps with over-representation of high frequencies, broadly tuned receptive fields and higher sound intensity thresholds as compared to controls. However, we did not detect differences in the number of parvalbumin-positive interneurons in AI of VPA and control rats. Altogether our findings show that neurophysiological impairments of hearing perception in this autism model occur independently of alterations in the number of parvalbumin-expressing interneurons. These data support the notion that fine circuit alterations, rather than gross cellular modification, could lead to neurophysiological changes in the autistic brain.

Mating depends on the accurate detection of signals that convey species identity and reproductive state. In African clawed frogs, Xenopus, this information is conveyed by vocal signals that differ in temporal patterns and spectral features between sexes and across species. We characterized spectral sensitivity using auditory-evoked potentials (AEPs), commonly known as the auditory brainstem response, in males and females of four Xenopus species. In female X. amieti, X. petersii, and X. laevis, peripheral auditory sensitivity to their species own dyad-two, species-specific dominant frequencies in the male advertisement call-is enhanced relative to males. Males were most sensitive to lower frequencies including those in the male-directed release calls. Frequency sensitivity was influenced by endocrine state; ovariectomized females had male-like auditory tuning while dihydrotestosterone-treated, ovariectomized females maintained female-like tuning. Thus, adult, female Xenopus demonstrate an endocrine-dependent sensitivity to the spectral features of conspecific male advertisement calls that could facilitate mating. Xenopus AEPs resemble those of other species in stimulus and level dependence, and in sensitivity to anesthetic (MS222). AEPs were correlated with body size and sex within some species. A frequency following response, probably encoded by the amphibian papilla, might facilitate dyad source localization via interaural time differences.

Macrophages are recruited into the cochlea in response to injury caused by acoustic trauma or ototoxicity, but the nature of the interaction between macrophages and the sensory structures of the inner ear remains unclear. The present study examined the role of fractalkine signaling in regulating the injury-evoked behavior of macrophages following the selective ablation of cochlear hair cells. We used a novel transgenic mouse model in which the human diphtheria toxin receptor (huDTR) is selectively expressed under the control of Pou4f3 , a hair cell-specific transcription factor. Administration of diphtheria toxin (DT) to these mice resulted in nearly complete ablation of cochlear hair cells, with no evident pathology among supporting cells, spiral ganglion neurons, or cells of the cochlear lateral wall. Hair cell death led to an increase in macrophages associated with the sensory epithelium of the cochlea. Their numbers peaked at 14 days after DT and then declined at later survival times. Increased macrophages were also observed within the spiral ganglion, but their numbers remained elevated for (at least) 56 d after DT. To investigate the role of fractalkine signaling in macrophage recruitment, we crossed huDTR mice to a mouse line that lacks expression of the fractalkine receptor (CX 3 CR1). Disruption of fractalkine signaling reduced macrophage recruitment into both the sensory epithelium and spiral ganglion and also resulted in diminished survival of spiral ganglion neurons after hair cell death. Our results suggest a fractalkine-mediated interaction between macrophages and the neurons of the cochlea. SIGNIFICANCE STATEMENT It is known that damage to the inner ear leads to recruitment of inflammatory cells (macrophages), but the chemical signals that initiate this recruitment and the functions of macrophages in the damaged ear are unclear. Here we show that fractalkine signaling regulates macrophage recruitment into the cochlea and also promotes the survival of cochlear afferents after selective hair cell lesion. Because these afferent neurons carry sound information from the cochlea to the auditory brainstem, their survival is a key determinant of the success of cochlear prosthetics. Our data suggest that fractalkine signaling in the cochlea is neuroprotective, and reveal a previously uncharacterized interaction between cells of the cochlea and the innate immune system.

A change-related component of the auditory evoked response long thought to be preattentive and preconscious may actually require consciousness.

Zebrafish ( Danio rerio ) is an important model organism in hearing research. However, data on the hearing sensitivity of zebrafish vary across different reports. In the present study, the hearing sensitivity of zebrafish was examined by analysing the auditory evoked potentials (AEPs) over a range of total lengths (TLs) from 12 to 46 mm. Morphological changes in the hair cells (HCs) of the saccule (the main auditory end organ) and their synapses with primary auditory neurons were investigated. The AEPs were detected up to a much higher frequency limit (12 kHz) than previously reported. No significant difference in the frequency response range was observed across the TL range examined. However, the AEP thresholds demonstrated both developmental improvement and age-related loss of hearing sensitivity. The changes in hearing sensitivity were roughly consistent with the morphological changes in the saccule including (1) the number and density of HCs, (2) the organization of stereocilia and (3) the quantity of a main ribbon protein, Ribeye b. The results of this study established a clear baseline for the hearing ability of zebrafish and revealed that the changes in the saccule contribute to the observed changes in TL (age)-related hearing sensitivity.