Publications using the ES1 & EC1

Combined electric and acoustic stimulation has proven to be an effective strategy to improve hearing in some cochlear implant users. We describe an acoustic microactuator to directly deliver stimuli to the perilymph in the scala tympani. The 800 µm by 800 µm actuator has a silicon diaphragm driven by a piezoelectric thin film (e.g., lead-zirconium-titanium oxide or PZT). This device could also be used as a component of a bimodal acoustic-electric electrode array. In the current study, we established a guinea pig model to test the actuator for its ability to deliver auditory signals to the cochlea in vivo. The actuator was placed through the round window of the cochlea. Auditory brainstem response (ABR) thresholds, peak latencies, and amplitude growth were calculated for an ear canal speaker versus the intracochlear actuator for tone burst stimuli at 4, 8, 16, and 24 kHz. An ABR was obtained after removal of the probe to assess loss of hearing related to the procedure. In some animals, the temporal bone was harvested for histologic analysis of cochlear damage. We show that the device is capable of stimulating ABRs in vivo with latencies and growth functions comparable to stimulation in the ear canal. Further experiments will be necessary to evaluate the efficiency and safety of this modality in long-term auditory stimulation and its ability to be integrated with conventional cochlear implant arrays.

Receptive fields of sensory neurons are considered to be dynamic and depend on the stimulus history. In the auditory system, evidence of dynamic frequency-receptive fields has been found following stimulus-specific adaptation (SSA). However, the underlying mechanism and circuitry of SSA have not been fully elucidated. Here, we studied how frequency-receptive fields of neurons in rat inferior colliculus (IC) changed when exposed to a biased tone sequence. Pure tone with one specific frequency (adaptor) was presented markedly more often than others. The adapted tuning was compared with the original tuning measured with an unbiased sequence. We found inhomogeneous changes in frequency tuning in IC, exhibiting a center-surround pattern with respect to the neuron's best frequency. Central adaptors elicited strong suppressive and repulsive changes while flank adaptors induced facilitative and attractive changes. Moreover, we proposed a two-layer model of the underlying network, which not only reproduced the adaptive changes in the receptive fields but also predicted novelty responses to oddball sequences. These results suggest that frequency-specific adaptation in auditory midbrain can be accounted for by an adapted frequency channel and its lateral spreading of adaptation, which shed light on the organization of the underlying circuitry.

The reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) requires adequate normalization in order to ensure accurate results. The use of reference genes is the most common method to normalize RT-qPCR assays; however, many studies have reported that the expression of frequently used reference genes is more variable than expected, depending on experimental conditions. Consequently, proper validation of the stability of reference genes is an essential step when performing new gene expression studies. Despite the fact that RT-qPCR has been widely used to elucidate molecular correlates of noise-induced hearing loss (NIHL), up to date there are no reports demonstrating validation of reference genes for the evaluation of changes in gene expression after NIHL. Therefore, in this study we evaluated the expression of some commonly used reference genes (Arbp, b-Act, b2m, CyA, Gapdh, Hprt1, Tbp, Tfrc and UbC) and examined their suitability as endogenous control genes for RT-qPCR analysis in the adult Wistar rat in response to NIHL. Four groups of rats were noise-exposed to generate permanent cochlear damage. Cochleae were collected at different time points after noise exposure and the expression level of candidate reference genes was evaluated by RT-qPCR using geNorm, NormFinder and BestKeeper software to determine expression stability. The three independent applications revealed Tbp as the most stably expressed reference gene. We also suggest a group of top-ranked reference genes that can be combined to obtain suitable reference gene pairs for the evaluation of the effects of noise on gene expression in the cochlea. These findings provide essential basis for further RT-qPCR analysis in studies of NIHL using Wistar rats as animal model.

Auditory hair cells have repeatedly been shown to be susceptible to ototoxicity from a multitude of drugs including aminoglycoside antibiotics. Here, we found that systemic HDAC inhibition using suberoylanilide hydroxamic acid (SAHA) on adult mice offers almost complete protection against hair cell loss and hearing threshold shifts from acute ototoxic insult from kanamycin potentiated with furosemide. We also found that the apparent lack of hair cell loss was completely independent of spontaneous or facilitated (ectopic Atoh1 induction) hair cell regeneration. Rather, SAHA treatment correlated with RelA acetylation (K310) and subsequent activation of the Nf- κ B pro-survival pathway leading to expression of pro-survival genes such as Cflar ( cFLIP ) and Bcl2l1 ( Bcl-xL ). In addition, we also detected increased expression of pro-survival genes Cdkn1a ( p21 ) and Hspa1a ( Hsp70 ), and decreased expression of the pro-apoptosis gene Bcl2l11 ( Bim ). These data combined provide evidence that class I HDACs control the transcriptional activation of pro-survival pathways in response to ototoxic insult by regulating the acetylation status of transcription factors found at the crossroads of cell death and survival in the mammalian inner ear.

In this study, we investigated the effects of varying intensity levels of repeated moderate noise exposures on hearing. The aim was to define an appropriate intensity level that could be repeated several times without giving rise to a permanent hearing loss, and thus establish a model for early adulthood moderate noise exposure in rats. Female Sprague-Dawley rats were exposed to broadband noise for 90 min, with a 50 % duty cycle at levels of 101, 104, 107, or 110 dB sound pressure level (SPL), and compared to a control group of non-exposed animals. Exposure was repeated every 6 weeks for a maximum of six repetitions or until a permanent hearing loss was observed. Hearing was assessed by the auditory brainstem response (ABR). Rats exposed to the higher intensities of 107 and 110 dB SPL showed permanent threshold shifts following the first exposure, while rats exposed to 101 and 104 dB SPL could be exposed at least six times without a sustained change in hearing thresholds. ABR amplitudes decreased over time for all groups, including the non-exposed control group, while the latencies were unaffected. A possible change in noise susceptibility following the repeated moderate noise exposures was tested by subjecting the animals to high-intensity noise exposure of 110 dB for 4 h. Rats previously exposed repeatedly to 104 dB SPL were slightly more resistant to high-intensity noise exposure than non-exposed rats or rats exposed to 101 dB SPL. Repeated moderate exposure to 104 dB SPL broadband noise is a viable model for early adulthood noise exposure in rats and may be useful for the study of noise exposure on age-related hearing loss.

Defects in the rat sarcoma viral oncogene homolog (Ras)/extracellular-signal-regulated kinase and the phosphatidylinositol 3-kinase-mammalian target of rapamycin (mTOR) signaling pathways are responsible for several neurodevelopmental disorders. These disorders are an important cause for intellectual disability; additional manifestations include autism spectrum disorder, seizures, and brain malformations. Changes in synaptic function are thought to underlie the neurological conditions associated with these syndromes. We therefore studied morphology and in vivo synaptic transmission of the calyx of Held synapse, a relay synapse in the medial nucleus of the trapezoid body (MNTB) of the auditory brainstem, in mouse models of tuberous sclerosis complex (TSC), Fragile X syndrome (FXS), Neurofibromatosis type 1 (NF1), and Costello syndrome. Calyces from both Tsc1(+/-) and from Fmr1 knock-out (KO) mice showed increased volume and surface area compared to wild-type (WT) controls. In addition, in Fmr1 KO animals a larger fraction of calyces showed complex morphology. In MNTB principal neurons of Nf1 (+/) (-) mice the average delay between EPSPs and APs was slightly smaller compared to WT controls, which could indicate an increased excitability. Otherwise, no obvious changes in synaptic transmission, or short-term plasticity were observed during juxtacellular recordings in any of the four lines. Our results in these four mutants thus indicate that abnormalities of mTOR or Ras signaling do not necessarily result in changes in in vivo synaptic transmission.

Pannexin1 (Panx1) is a gap junction gene in vertebrates whose proteins mainly function as non-junctional channels on the cell surface. Panx1 channels can release ATP under physiological conditions and play critical roles in many physiological and pathological processes. Here, we report that Panx1 deficiency can reduce ATP release and endocochlear potential (EP) generation in the cochlea inducing hearing loss. Panx1 extensively expresses in the cochlea, including the cochlear lateral wall. We found that deletion of Panx1 in the cochlear lateral wall almost abolished ATP release under physiological conditions. Positive EP is a driving force for current through hair cells to produce auditory receptor potential. EP generation requires ATP. In the Panx1 deficient mice, EP and auditory receptor potential as measured by cochlear microphonics (CM) were significantly reduced. However, no apparent hair cell loss was detected. Moreover, defect of connexin hemichannels by deletion of connexin26 (Cx26) and Cx30, which are predominant connexin isoforms in the cochlea, did not reduce ATP release under physiological conditions. These data demonstrate that Panx1 channels dominate ATP release in the cochlea ensuring EP and auditory receptor potential generation and hearing. Panx1 deficiency can reduce ATP release and EP generation causing hearing loss.

In an ever changing auditory scene, change detection is an ongoing task performed by the auditory brain. Neurons in the midbrain and auditory cortex that exhibit stimulus-specific adaptation (SSA) may contribute to this process. Those neurons adapt to frequent sounds while retaining their excitability to rare sounds. Here, we test whether neurons exhibiting SSA and those without are part of the same networks in the inferior colliculus (IC). We recorded the responses to frequent and rare sounds and then marked the sites of these neurons with a retrograde tracer to correlate the source of projections with the physiological response. SSA neurons were confined to the non-lemniscal subdivisions and exhibited broad receptive fields, while the non-SSA were confined to the central nucleus and displayed narrow receptive fields. SSA neurons receive strong inputs from auditory cortical areas and very poor or even absent projections from the brainstem nuclei. On the contrary, the major sources of inputs to the neurons that lacked SSA were from the brainstem nuclei. These findings demonstrate that auditory cortical inputs are biased in favor of IC synaptic domains that are populated by SSA neurons enabling them to compare top-down signals with incoming sensory information from lower areas.

The dynamic adjustment of hearing sensitivity and frequency selectivity is mediated by the medial olivocochlear efferent reflex, which suppresses the gain of the ‘cochlear amplifier’ in each ear. Such efferent feedback is important for promoting discrimination of sounds in background noise, sound localization and protecting the cochleae from acoustic overstimulation. However, the sensory driver for the olivocochlear reflex is unknown. Here, we resolve this longstanding question using a mouse model null for the gene encoding the type III intermediate filament peripherin ( Prph) . Prph (−/−) mice lacked type II spiral ganglion neuron innervation of the outer hair cells, whereas innervation of the inner hair cells by type I spiral ganglion neurons was normal. Compared with Prph (+/+) controls, both contralateral and ipsilateral olivocochlear efferent-mediated suppression of the cochlear amplifier were absent in Prph (−/−) mice, demonstrating that outer hair cells and their type II afferents constitute the sensory drive for the olivocochlear efferent reflex.

There are two types of sensory cells in the mammalian cochlea, inner hair cells, which make synaptic contact with auditory-nerve afferent fibers, and outer hair cells that are innervated by crossed and uncrossed medial olivocochlear (MOC) efferent fibers. Contralateral acoustic stimulation activates the uncrossed efferent MOC fibers reducing cochlear neural responses, thus modifying the input to the central auditory system. The chinchilla, among all studied mammals, displays the lowest percentage of uncrossed MOC fibers raising questions about the strength and frequency distribution of the contralateral-sound effect in this species. On the other hand, MOC effects on cochlear sensitivity have been mainly studied in anesthetized animals and since the MOC-neuron activity depends on the level of anesthesia, it is important to assess the influence of anesthesia in the strength of efferent effects. Seven adult chinchillas (Chinchilla laniger) were chronically implanted with round-window electrodes in both cochleae. We compared the effect of contralateral sound in awake and anesthetized condition. Compound action potentials (CAP) and cochlear microphonics (CM) were measured in the ipsilateral cochlea in response to tones in absence and presence of contralateral sound. Control measurements performed after middle-ear muscles section in one animal discarded any possible middle-ear reflex activation. Contralateral sound produced CAP amplitude reductions in all chinchillas, with suppression effects greater by about 1-3 dB in awake than in anesthetized animals. In contrast, CM amplitude increases of up to 1.9 dB were found in only three awake chinchillas. In both conditions the strongest efferent effects were produced by contralateral tones at frequencies equal or close to those of ipsilateral tones. Contralateral CAP suppressions for 1-6 kHz ipsilateral tones corresponded to a span of uncrossed MOC fiber innervation reaching at least the central third of the chinchilla cochlea.

The work presents the results of experimental study on the possibilities of determining the source of an ultrasonic signal in two-dimensional space (distance, horizontal angle). During the research the team used a self-constructed linear array of MEMS microphones. Knowledge in the field of sonar systems was utilized to analyse and design a location system based on a microphone array. Using the above mentioned transducers and broadband ultrasound sources allows a quantitative comparison of estimation of the location of an ultrasonic wave source with the use of broadband modulated signals (modelled on bats' echolocation signals) to be performed. During the laboratory research the team used various signal processing algorithms, which made it possible to select an optimal processing strategy, where the sending signal is known.

The most common reason for sensorineural deafness is death of hair cells (HCs). Heat shock proteins (HSPs) are molecular chaperones that participate in folding, targeting, and degrading proteins. HSP expression is increased in response to various environmental stresses to protect cells from damage. Here, we tested whether viral-mediated overexpression of HSP70 can protect HCs and hearing from severe ototoxicity (kanamycin and furosemide) in guinea pigs. Adenovirus-HSP70 mCherry (Ad.HSP70-mCherry) was injected to experimental animals and adenovirus-mCherry to controls, 4 days before the ototoxic insult. Hearing thresholds were measured by auditory brainstem response before the insult and again before sacrificing the animals, 14 days after the insult. Epi-fluorescence immunocytochemistry showed that injection of Ad.HSP70-mCherry resulted in mCherry fluorescence in nonsensory cells of the organ of Corti. The ototoxic insult eliminated both outer HCs and inner HCs throughout most of the cochlea of control (adenovirus-mCherry-injected) ears and contralateral (uninjected) ears. Ad.HSP70-mCherry-injected ears exhibited a significant preservation of inner HCs compared to control and contralateral ears, but outer HCs were not protected. Auditory brainstem response thresholds were significantly better in Ad.HSP70-mCherry-injected ears than in control and contralateral ears. Our data show that HSP70 augmentation may represent a potential therapy attenuating ototoxic inner HC loss.

The interaural level difference (ILD) cue to sound location is first encoded in the lateral superior olive (LSO). ILD sensitivity results because the LSO receives excitatory input from the ipsilateral cochlear nucleus and inhibitory input indirectly from the contralateral cochlear nucleus via glycinergic neurons of the ipsilateral medial nucleus of the trapezoid body (MNTB). It is hypothesized that in order for LSO neurons to encode ILDs, the sound spectra at both ears must be accurately encoded via spike rate by their afferents. This spectral-coding hypothesis has not been directly tested in MNTB, likely because MNTB neurons have been mostly described and studied recently in regards to their abilities to encode temporal aspects of sounds, not spectral. Here, we test the hypothesis that MNTB neurons and their inputs from the cochlear nucleus and auditory nerve code sound spectra via discharge rate. The Random Spectral Shape (RSS) method was used to estimate how the levels of 100-ms duration spectrally stationary stimuli were weighted, both linearly and non-linearly, across a wide band of frequencies. In general, MNTB neurons, and their globular bushy cell inputs, were found to be well-modeled by a linear weighting of spectra demonstrating that the pathways through the MNTB can accurately encode sound spectra including those resulting from the acoustical cues to sound location provided by head-related directional transfer functions (DTFs). Together with the anatomical and biophysical specializations for timing in the MNTB-LSO complex, these mechanisms may allow ILDs to be computed for complex stimuli with rapid spectrotemporally-modulated envelopes such as speech and animal vocalizations and moving sound sources.

Assemblies of vertically connected neurons in the cerebral cortex form information processing units (columns) that participate in the distribution and segregation of sensory signals. Despite well-accepted models of columnar architecture, functional mechanisms of inter-laminar communication remain poorly understood. Hence, the purpose of the present investigation was to examine the effects of sensory information features on columnar response properties. Using acute recording techniques, extracellular response activity was collected from the right hemisphere of eight mature cats (felis catus). Recordings were conducted with multichannel electrodes that permitted the simultaneous acquisition of neuronal activity within primary auditory cortex columns. Neuronal responses to simple (pure tones), complex (noise burst and frequency modulated sweeps), and ecologically relevant (con-specific vocalizations) acoustic signals were measured. Collectively, the present investigation demonstrates that despite consistencies in neuronal tuning (characteristic frequency), irregularities in discharge activity between neurons of individual A1 columns increase as a function of spectral (signal complexity) and temporal (duration) acoustic variations.

It is generally thought that background noise can mask auditory information. However, how the noise specifically transforms neuronal auditory processing in a level-dependent manner remains to be carefully determined. Here, with in vivo loose-patch cell-attached recordings in layer 4 of the rat primary auditory cortex (A1), we systematically examined how continuous wideband noise of different levels affected receptive field properties of individual neurons. We found that the background noise, when above a certain critical/effective level, resulted in an elevation of intensity threshold for tone-evoked responses. This increase of threshold was linearly dependent on the noise intensity above the critical level. As such, the tonal receptive field (TRF) of individual neurons was translated upward as an entirety toward high intensities along the intensity domain. This resulted in preserved preferred characteristic frequency (CF) and the overall shape of TRF, but reduced frequency responding range and an enhanced frequency selectivity for the same stimulus intensity. Such translational effects on intensity threshold were observed in both excitatory and fast-spiking inhibitory neurons, as well as in both monotonic and nonmonotonic (intensity-tuned) A1 neurons. Our results suggest that in a noise background, fundamental auditory representations are modulated through a background level-dependent linear shifting along intensity domain, which is equivalent to reducing stimulus intensity.

This study describes two experiments that were conducted in search for a behavioral paradigm to test for tinnitus in guinea pigs. Conditioning paradigms are available to determine the presence of tinnitus in animals and are based on the assumption that tinnitus impairs their ability to detect silent intervals in continuous noise. Guinea pigs have not been subjected to these paradigms yet; therefore, we investigated whether guinea pigs could be conditioned in the two-way shuttle-box paradigm to respond to silent intervals in noise. Even though guinea pigs could be trained relatively easy to respond to the presence of a noise interval, training guinea pigs to silent intervals in noise was unsuccessful. Instead, it appeared that they became immobile when the continuous stimulus was suddenly stopped. This was confirmed by the next experiment, in which we subjected guinea pigs to alternating intervals of noise and silence with a random duration between 30 and 120 s. Indeed, guinea pigs were significantly longer immobile during silence compared to during noise. By interpreting immobility as a signature of perceiving silence, we hypothesized that the presence of tinnitus would reduce immobility in silence. Therefore, we unilaterally exposed one group of guinea pigs to an 11-kHz tone of 124 dB sound pressure level for 1 h. A subset of the exposed animals was significantly more active in silence, but also more active in noise, as compared to the control group. The increased mobility during silent intervals might represent tinnitus. However, the increased mobility in noise of this group implies that the observed behavior could have derived from, e.g., an overall increase in activity. Therefore, conducting validation experiments is very important before implementing this method as a new screening tool for tinnitus. Follow-up experiments are discussed to further elucidate the origin of the increased mobility in both silence and noise.

Conductive hearing loss causes a progressive decline in cochlear activity that may result in functional and structural modifications in auditory neurons. However, whether these activity-dependent changes are accompanied by a glial response involving microglia, astrocytes, or both has not yet been fully elucidated. Accordingly, the present study was designed to determine the involvement of glial related mechanisms in the anteroventral cochlear nucleus (AVCN) of adult rats at 1, 4, 7, and 15 d after removing middle ear ossicles. Quantitative immunohistochemistry analyses at light microscopy with specific markers of microglia or astroglia along with immunocytochemistry at the electron microscopy level were used. Also, in order to test whether trophic support by neurotrophins is modulated in glial cells by auditory activity, the expression and distribution of neurotrophin-3 (NT-3) and its colocalization with microglial or astroglial markers was investigated. Diminished cochlear activity after middle ear ossicle removal leads to a significant ipsilateral increase in the mean gray levels and stained area of microglial cells but not astrocytes in the AVCN at 1 and 4 d post-lesion as compared to the contralateral side and control animals. These results suggest that microglial cells but not astrocytes may act as dynamic modulators of synaptic transmission in the cochlear nucleus immediately following unilateral hearing loss. On the other hand, NT-3 immunostaining was localized mainly in neuronal cell bodies and axons and was upregulated at 1, 4 and 7 d post-lesion. Very few glial cells expressed this neurotrophin in both control and experimental rats, suggesting that NT-3 is primarily activated in neurons and not as much in glia after limiting auditory activity in the AVCN by conductive hearing loss.

The mechanism of loss of consciousness (LOC) under anesthesia is unknown. Because consciousness depends on activity in the cortico-thalamic network, anesthetic actions on this network are likely critical for LOC. Competing theories stress the importance of anesthetic actions on bottom-up "core" thalamo-cortical (TC) vs. top-down cortico-cortical (CC) and matrix TC connections. We tested these models using laminar recordings in rat auditory cortex in vivo and murine brain slices. We selectively activated bottom-up vs. top-down afferent pathways using sensory stimuli in vivo and electrical stimulation in brain slices, and compared effects of isoflurane on responses evoked via the two pathways. Auditory stimuli in vivo and core TC afferent stimulation in brain slices evoked short latency current sinks in middle layers, consistent with activation of core TC afferents. By contrast, visual stimuli in vivo and stimulation of CC and matrix TC afferents in brain slices evoked responses mainly in superficial and deep layers, consistent with projection patterns of top-down afferents that carry visual information to auditory cortex. Responses to auditory stimuli in vivo and core TC afferents in brain slices were significantly less affected by isoflurane compared to responses triggered by visual stimuli in vivo and CC/matrix TC afferents in slices. At a just-hypnotic dose in vivo, auditory responses were enhanced by isoflurane, whereas visual responses were dramatically reduced. At a comparable concentration in slices, isoflurane suppressed both core TC and CC/matrix TC responses, but the effect on the latter responses was far greater than on core TC responses, indicating that at least part of the differential effects observed in vivo were due to local actions of isoflurane in auditory cortex. These data support a model in which disruption of top-down connectivity contributes to anesthesia-induced LOC, and have implications for understanding the neural basis of consciousness.

The transgenic P23H line 1 (P23H-1) rat expresses a variant of rhodopsin with a mutation that leads to loss of visual function. This rat strain is an experimental model usually employed to study photoreceptor degeneration. Although the mutated protein should not interfere with other sensory functions, observing severe loss of auditory reflexes in response to natural sounds led us to study auditory brain response (ABR) recording. Animals were separated into different hearing levels following the response to natural stimuli (hand clapping and kissing sounds). Of all the analyzed animals, 25.9% presented auditory loss before 50 days of age (P50) and 45% were totally deaf by P200. ABR recordings showed that all the rats had a higher hearing threshold than the control Sprague-Dawley (SD) rats, which was also higher than any other rat strains. The integrity of the central and peripheral auditory pathway was analyzed by histology and immunocytochemistry. In the cochlear nucleus (CN), statistical differences were found between SD and P23H-1 rats in VGluT1 distribution, but none were found when labeling all the CN synapses with anti-Syntaxin. This finding suggests anatomical and/or molecular abnormalities in the auditory downstream pathway. The inner ear of the hypoacusic P23H-1 rats showed several anatomical defects, including loss and disruption of hair cells and spiral ganglion neurons. All these results can explain, at least in part, how hearing impairment can occur in a high percentage of P23H-1 rats. P23H-1 rats may be considered an experimental model with visual and auditory dysfunctions in future research.

Previously, we showed that age-related hearing loss (AHL) was delayed in C57BL6 mice overexpressing X-Linked Inhibitor of Apoptotic Protein (XIAP), and the delayed AHL was associated with attenuated hair cell (HC) loss in XIAP-overexpressing mice. Similar to other reports, the HC loss in aged mice was restricted to the basal turn in this previous study, and occurred slightly at the apical end of the cochlea, showing considerably less spread than the frequency region of hearing loss. In the present study, we examined whether and how AHL is related to the degeneration of neuronal innervation of the cochlea and whether the overexpression of XIAP exerts a protective effect against age-related degeneration in both afferent and efferent cochlear neurites. In contrast to HC loss, degeneration of both afferent and efferent neurites spread to the middle turns of the cochlea. Moreover, XIAP-overexpressing mice lost fewer HC afferent dendrites and efferent axons, as well as fewer spiral ganglion neurons between 3 and 14 months of age in comparison with wild-type littermates. The results suggest that age-related degeneration of cochlear neurites may be independent of HC loss. Further, the inhibition of apoptosis by XIAP appears to reduce degeneration of both afferent and efferent cochlear neurites.