Auditory Research publications

Mutations in the GJB2 and GJB6 genes, respectively, coding for connexin26 (Cx26) and connexin30 (Cx30) proteins, are the most common cause for prelingual non-syndromic deafness in humans. In the inner ear, Cx26 and Cx30 are expressed in different non-sensory cell types, where they largely co-localize and may form heteromeric gap junction channels. Here, we describe the generation and characterization of a mouse model for human bilateral middle/high-frequency hearing loss based on the substitution of an evolutionarily conserved threonine by a methionine residue at position 5 near the N-terminus of Cx30 (Cx30T5M). The mutation was inserted in the mouse genome by homologous recombination in mouse embryonic stem cells. Expression of the mutated Cx30T5M protein in these transgenic mice is under the control of the endogenous Cx30 promoter and was analysed via activation of the lacZ reporter gene. When probed by auditory brainstem recordings, Cx30(T5M/T5M) mice exhibited a mild, but significant increase in their hearing thresholds of about 15 dB at all frequencies. Immunolabelling with antibodies to Cx26 or Cx30 suggested normal location of these proteins in the adult inner ear, but western blot analysis showed significantly down-regulated the expression levels of Cx26 and Cx30. In the developing cochlea, electrical coupling, probed by dual patch-clamp recordings, was normal. However, transfer of the fluorescent tracer calcein between cochlear non-sensory cells was reduced, as was intercellular Ca(2+) signalling due to spontaneous ATP release from connexin hemichannels. Our findings link hearing loss to decreased biochemical coupling due to the point-mutated Cx30 in mice.

Cave fish have rarely been investigated with regard to their inner ear morphology, hearing abilities, and acoustic communication. Based on a previous study that revealed morphological differences in the saccular otolith between a cave and two surface populations of Poecilia mexicana, we checked for additional differences in utricular and lagenar otoliths and tested whether different populations have similar hearing sensitivities. We found pronounced differences in the shape of all three otoliths. Otoliths of the saccule and lagena from cave fish differed from those of surface fish in the features of the face oriented towards the sensory epithelium. In addition, otoliths of the utricle and lagena were significantly heavier in cave fish. Auditory sensitivities were measured between 100 and 1500Hz, utilizing the auditory evoked potential recording technique. We found similar hearing abilities in cave and surface fish, with greatest sensitivity between 200 and 300Hz. An acoustic survey revealed that neither ecotype produced species-specific sounds. Our data indicate that cave dwelling altered the otolith morphology in Atlantic mollies, probably due to metabolic differences. Different otolith morphology, however, did not affect general auditory sensitivity or acoustic behavior.

Sensory systems are known to adapt their coding strategies to the statistics of their environment, but little is still known about the perceptual implications of such adjustments. We investigated how auditory spatial processing adapts to stimulus statistics by presenting human listeners and anesthetized ferrets with noise sequences in which interaural level differences (ILD) rapidly fluctuated according to a Gaussian distribution. The mean of the distribution biased the perceived laterality of a subsequent stimulus, whereas the distribution's variance changed the listeners' spatial sensitivity. The responses of neurons in the inferior colliculus changed in line with these perceptual phenomena. Their ILD preference adjusted to match the stimulus distribution mean, resulting in large shifts in rate-ILD functions, while their gain adapted to the stimulus variance, producing pronounced changes in neural sensitivity. Our findings suggest that processing of auditory space is geared toward emphasizing relative spatial differences rather than the accurate representation of absolute position.

A growing number of studies of auditory processing are being carried out in awake, behaving animals, creating a need for precisely controlled sound delivery without restricting head movements. We have designed a system for closed-field stimulus presentation in freely moving ferrets, which comprises lightweight, adjustable headphones that can be consistently positioned over the ears via a small, skull-mounted implant. The invasiveness of the implant was minimized by simplifying its construction and using dental adhesive only for attaching it to the skull, thereby reducing the surgery required and avoiding the use of screws or other anchoring devices. Attaching the headphones to a chronic implant also reduced the amount of contact they had with the head and ears, increasing the willingness of the animals to wear them. We validated sound stimulation via the headphones in ferrets trained previously in a free-field task to localize stimuli presented from one of two loudspeakers. Noise bursts were delivered binaurally over the headphones and interaural level differences (ILDs) were introduced to allow the sound to be lateralized. Animals rapidly transferred from the free-field task to indicate the perceived location of the stimulus presented over headphones. They showed near perfect lateralization with a 5 dB ILD, matching the scores achieved in the free-field task. As expected, the ferrets' performance declined when the ILD was reduced in value. This closed-field system can easily be adapted for use in other species, and provides a reliable means of presenting closed-field stimuli whilst monitoring behavioral responses in freely moving animals.

Neurons in the auditory cortex of anesthetized animals are generally considered to generate phasic responses to simple stimuli such as tones or noise bursts. In this paper, we show that under ketamine/medetomidine anesthesia, neurons in ferret auditory cortex usually exhibit complex sustained responses. We presented 100-ms broad-band noise bursts at a range of interaural level differences (ILDs) and average binaural levels (ABLs), and used extracellular electrodes to monitor evoked activity over 700 ms poststimulus onset. We estimated the degree of randomness (noise) in the response functions of individual neurons over poststimulus time; we found that neural activity was significantly modulated by sound for up to ∼500 ms following stimulus offset. Pooling data from all neurons, we found that spiking activity carries significant information about stimulus identity over this same time period. However, information about ILD decayed much more quickly over time compared with information about ABL. In addition, ILD and ABL are coded independently by the neural population even though this is not the case at individual neurons. Though most neurons responded more strongly to ILDs corresponding to the opposite side of space, as a population, they were equally informative about both contra- and ipsilateral stimuli.

Background: Surveys of ontogenetic development of hearing and sound production in fish are scarce, and the ontogenetic development of acoustic communication has been investigated in only two fish species so far. Studies on the labyrinth fish Trichopsis vittata and the toadfish Halobatrachus didactylus show that the ability to detect conspecific sounds develops during growth. In otophysine fish, which are characterized by Weberian ossicles and improved hearing sensitivities, the ontogenetic development of sound communication has never been investigated. We analysed the ontogeny of the auditory sensitivity and vocalizations in the mochokid catfish Synodontis schoutedeni. Mochokid catfishes of the genus Synodontis are commonly called squeakers because they produce broadband stridulation sounds during abduction and adduction of pectoral fin spines. Fish from six different size groups - from 22 mm standard length to 126 mm - were studied. Hearing thresholds were measured between 50 Hz and 6 kHz using the auditory evoked potentials recording technique; stridulation sounds were recorded and their sound pressure levels determined. Finally, absolute sound power spectra were compared to auditory sensitivity curves within each size group. Results: The smallest juveniles showed the poorest hearing abilities of all size groups between 50 and 1,000 Hz and highest hearing sensitivity at 5 and 6 kHz. The duration of abduction and adduction sounds and the pulse period increased and sound pressure level (in animals smaller than 58 mm) increased, while the dominant frequency of sounds decreased with size in animals larger than 37 mm. Comparisons between audiograms and sound spectra revealed that the most sensitive frequencies correlate with the dominant frequencies of stridulation sounds in all S. schoutedeni size groups and that all specimens are able to detect sounds of all size groups. Conclusions: This study on the squeaker catfish S. schoutedeni is the first to demonstrate that absolute hearing sensitivity changes during ontogeny in an otophysine fish. This contrasts with prior studies on two cypriniform fish species in which no such change could be observed. Furthermore, S. schoutedeni can detect conspecific sounds at all stages of development, again contrasting with prior findings in fishes.

Insulin-like growth factor-I (IGF-I) belongs to the family of insulin-related peptides that fulfils a key role during the late development of the nervous system. Human IGF1 mutations cause profound deafness, poor growth and mental retardation. Accordingly, Igf1(-/-) null mice are dwarfs that have low survival rates, cochlear alterations and severe sensorineural deafness. Presbycusis (age-related hearing loss) is a common disorder associated with aging that causes social and cognitive problems. Aging is also associated with a decrease in circulating IGF-I levels and this reduction has been related to cognitive and brain alterations, although there is no information as yet regarding the relationship between presbycusis and IGF-I biodisponibility. Here we present a longitudinal study of wild type Igf1(+/+) and null Igf1(-/-) mice from 2 to 12 months of age comparing the temporal progression of several parameters: hearing, brain morphology, cochlear cytoarchitecture, insulin-related factors and IGF gene expression and IGF-I serum levels. Complementary invasive and non-invasive techniques were used, including auditory brainstem-evoked response (ABR) recordings and in vivo MRI brain imaging. Igf1(-/-) null mice presented profound deafness at all the ages studied, without any obvious worsening of hearing parameters with aging. Igf1(+/+) wild type mice suffered significant age-related hearing loss, their auditory thresholds and peak I latencies augmenting as they aged, in parallel with a decrease in the circulating levels of IGF-I. Accordingly, there was an age-related spiral ganglion degeneration in wild type mice that was not evident in the Igf1 null mice. However, the Igf1(-/-) null mice in turn developed a prematurely aged stria vascularis reminiscent of the diabetic strial phenotype. Our data indicate that IGF-I is required for the correct development and maintenance of hearing, supporting the idea that IGF-I-based therapies could contribute to prevent or ameliorate age-related hearing loss.

Mutations in the CLRN1 gene cause Usher syndrome type 3 (USH3), a human disease characterized by progressive blindness and deafness. Clarin 1, the protein product of CLRN1, is a four-transmembrane protein predicted to be associated with ribbon synapses of photoreceptors and cochlear hair cells, and recently demonstrated to be associated with the cytoskeleton. To study Clrn1, we created a Clrn1 knockout (KO) mouse and characterized the histological and functional consequences of Clrn1 deletion in the retina and cochlea. Clrn1 KO mice do not develop a retinal degeneration phenotype, but exhibit progressive loss of sensory hair cells in the cochlea and deterioration of the organ of Corti by 4 months. Hair cell stereocilia in KO animals were longer and disorganized by 4 months, and some Clrn1 KO mice exhibited circling behavior by 5-6 months of age. Clrn1 mRNA expression was localized in the retina using in situ hybridization (ISH), laser capture microdissection (LCM), and RT-PCR. Retinal Clrn1 transcripts were found throughout development and adulthood by RT-PCR, although expression peaked at P7 and declined to undetectable levels in adult retina by ISH. LCM localized Clrn1 transcripts to the retinas inner nuclear layer, and WT levels of retinal Clrn1 expression were observed in photoreceptor-less retinas. Examination of Clrn1 KO mice suggests that CLRN1 is unnecessary in the murine retina but essential for normal cochlear development and function. This may reflect a redundancy in the mouse retina not present in human retina. In contrast to mouse KO models of USH1 and USH2, our data indicate that Clrn1 expression in the retina is restricted to the Müller glia. This is a novel finding, as most retinal degeneration associated proteins are expressed in photoreceptors, not in glia. If CLRN1 expression in humans is comparable to the expression pattern observed in mice, this is the first report of an inner retinal protein that, when mutated, causes retinal degeneration.

Exogenous delivery of neurotrophic factors into the cochlea of deafened animals rescues spiral ganglion cells (SGCs) from degeneration. To be clinically relevant for human cochlear implant candidates, the protective effect of neurotrophins should persist after cessation of treatment and the treated SGCs should remain functional. In this study, the survival and functionality of SGCs were investigated after temporary treatment with brain-derived neurotrophic factor (BDNF). Guinea pigs in the experimental group were deafened, and 2 weeks later, the right cochleae were implanted with an electrode array and drug delivery cannula. BDNF was administered to the implanted cochleae during a 4-week period via a mini-osmotic pump. After completion of the treatment, the osmotic pumps were removed. Two weeks later, the animals were killed and the survival of SGCs was analyzed. To monitor the functionality of the auditory nerve, electrically evoked auditory brainstem responses (eABRs) were recorded in awake animals throughout the experiment. BDNF treatment resulted in enhanced survival of SGCs 2 weeks after cessation of the treatment and prevented the decreases in size and circularity that are seen in the untreated contralateral cochleae. The amplitude of the suprathreshold eABR response in BDNF-treated animals was significantly larger than in deafened control animals and comparable to that in normal-hearing control animals. The amplitude in the BDNF-treated group did not decrease significantly after cessation of treatment. The eABR latency in BDNF-treated animals was longer than normal and comparable to that in deafened control animals. These morphological and functional findings demonstrate that neurotrophic intervention had a lasting effect, which is promising for future clinical application of neurotrophic factors in implanted human cochleae.

Background Developmental exposure to polychlorinated biphenyls (PCBs) or methylmercury (MeHg) can result in a variety of neurotoxic effects, including long-term auditory deficits. However, little is known about the effects of combined exposure to PCBs and MeHg on auditory function. Objective We developmentally exposed rats to PCBs and/or MeHg and assessed auditory function in adulthood to determine the effects of exposure to these contaminants individually and in combination. Methods We exposed female Long-Evans rats to 1 or 3 mg/kg PCB in corn oil, 1.5 or 4.5 ppm MeHg in drinking water, or combined exposure to 1 mg/kg PCB + 1.5 ppm MeHg or 3 mg/kg PCB + 4.5 ppm MeHg. Controls received corn oil vehicle and unadulterated water. Dosing began 28 days before breeding and continued until weaning at postnatal day (PND) 21. Auditory function of the offspring was assessed at approximately PND 200 by measuring distortion product otoacoustic emissions (DPOAEs) and auditory brainstem responses (ABRs). Results Groups exposed to PCBs alone had attenuated DPOAE amplitudes, elevated DPOAE thresholds, and elevated ABR thresholds compared with controls. Groups exposed to MeHg alone did not differ from controls. Unexpectedly, the effects of PCB exposure appeared to be attenuated by coexposure to MeHg. Conclusion Developmental exposure to PCBs can result in permanent hearing deficits, and the changes in DPOAE amplitudes and thresholds suggest a cochlear site of action. Coexposure to MeHg appeared to attenuate the PCB-related deficits, but the mechanism for this unexpected interaction remains to be determined.

This thesis is divided into six chapters (following Introduction). Each chapter was intended to be self-contained, so they do not have to be read in the order they are presented. Second chapter (Sec. 2) contains a detailed description of experimental techniques: surgery, recording, and training techniques we used in awake head-fixed rats. We have also included a detailed description of all sets of stimuli we used to probe neurons, analytical methods used to analyze data, and description of computational models used in other parts of the thesis.Third chapter (Sec. 3) focuses on description of single-neuron responses in primary auditory cortex of awake head-fixed rats. The primary emphasis of this part is on the sparse representation of various auditory stimuli we used to probe neurons, and the heterogeneity of responses of single neurons. To characterize population responses to sound in the auditory cortex we asked the question "What is the typical response to acoustic stimuli?" instead of what is usually asked "What is the stimulus that evokes a response?" We found that the population response was sparse, with many unresponsive neurons. In addition, the responsive neurons showed a great variety of responses. This heterogeneity of neuronal responses ("response zoo," courtesy of Anthony M. Zádor) was, however, surprisingly well characterized by lognormal distribution of firing rates. The observation that firing rates in awake auditory cortex were lognormally distributed was even more interesting given the observation of lognormal distribution of synaptic weights in the cerebral cortex.The fourth chapter (Sec. 4) focuses on mechanisms which could give rise to lognormal distribution of firing rates, as well as synaptic weights. We proposed specific types of correlations among synaptic connections, and formulated a multiplicative learning rule which led to the observed distributions. We were also able to characterize intracellular activity of neurons in awake auditory cortex. The fifth chapter (Sec. 5) contains analysis of so-called up and down states in awake auditory cortex. We show that up and down states—the "signature" subthreshold dynamics so often described in various cortical areas of anesthetized animals—were rare in the primary auditory cortex of awake rats, instead, subthreshold dynamics was consisted of brief, infrequent fluctuations of membrane potential.The experiments described and analyzed in chapters 2—-4 were conducted in naïve awake rats. As behavior or attention can influence neuronal activity even in primary sensory areas, we developed a setup for head-fixed behavior. In the sixth chapter (Sec. 6) we describe the sound discrimination task we have used to study behavior in head-fixed rats. We present a comparison of basic behavioral parameters between restrained and unrestrained rats, as well as evidence of nonauditory modulations of single neuron activity in auditory cortex.

In this study, we investigate the ability of the mammalian auditory pathway to adapt its strategy for temporal processing under natural stimulus conditions. We derive temporal receptive fields from the responses of neurons in the inferior colliculus to vocalization stimuli with and without additional ambient noise. We find that the onset of ambient noise evokes a change in receptive field dynamics that corresponds to a change from bandpass to lowpass temporal filtering. We show that these changes occur within a few hundred milliseconds of the onset of the noise and are evident across a range of overall stimulus intensities. Using a simple model, we illustrate how these changes in temporal processing exploit differences in the statistical properties of vocalizations and ambient noises to increase the information in the neural response in a manner consistent with the principles of efficient coding.

How do neuronal populations in the auditory cortex represent acoustic stimuli? Although sound-evoked neural responses in the anesthetized auditory cortex are mainly transient, recent experiments in the unanesthetized preparation have emphasized subpopulations with other response properties. To quantify the relative contributions of these different subpopulations in the awake preparation, we have estimated the representation of sounds across the neuronal population using a representative ensemble of stimuli. We used cell-attached recording with a glass electrode, a method for which single-unit isolation does not depend on neuronal activity, to quantify the fraction of neurons engaged by acoustic stimuli (tones, frequency modulated sweeps, white-noise bursts, and natural stimuli) in the primary auditory cortex of awake head-fixed rats. We find that the population response is sparse, with stimuli typically eliciting high firing rates (>20 spikes/second) in less than 5% of neurons at any instant. Some neurons had very low spontaneous firing rates (<0.01 spikes/second). At the other extreme, some neurons had driven rates in excess of 50 spikes/second. Interestingly, the overall population response was well described by a lognormal distribution, rather than the exponential distribution that is often reported. Our results represent, to our knowledge, the first quantitative evidence for sparse representations of sounds in the unanesthetized auditory cortex. Our results are compatible with a model in which most neurons are silent much of the time, and in which representations are composed of small dynamic subsets of highly active neurons.

Background: Mucopolysaccharidosis (MPS) IIIB (Sanfilippo Syndrome type B) is caused by a deficiency in the lysosomal enzyme N-acetyl-glucosaminidase (Naglu). Children with MPS IIIB develop disturbances of sleep, activity levels, coordination, vision, hearing, and mental functioning culminating in early death. The murine model of MPS IIIB demonstrates lysosomal distention in multiple tissues, a shortened life span, and behavioral changes. Principal findings: To more thoroughly assess MPS IIIB in mice, alterations in circadian rhythm, activity level, motor function, vision, and hearing were tested. The suprachiasmatic nucleus (SCN) developed pathologic changes and locomotor analysis showed that MPS IIIB mice start their daily activity later and have a lower proportion of activity during the night than wild-type controls. Rotarod assessment of motor function revealed a progressive inability to coordinate movement in a rocking paradigm. Purkinje cell counts were significantly reduced in the MPS IIIB animals compared to age matched controls. By electroretinography (ERG), MPS IIIB mice had a progressive decrease in the amplitude of the dark-adapted b-wave response. Corresponding pathology revealed shortening of the outer segments, thinning of the outer nuclear layer, and inclusions in the retinal pigmented epithelium. Auditory-evoked brainstem responses (ABR) demonstrated progressive hearing deficits consistent with the observed loss of hair cells in the inner ear and histologic abnormalities in the middle ear. Conclusions/significance: The mouse model of MPS IIIB has several quantifiable phenotypic alterations and is similar to the human disease. These physiologic and histologic changes provide insights into the progression of this disease and will serve as important parameters when evaluating various therapies.

Plasticity studies suggest that behavioral relevance can change the cortical processing of trained or conditioned sensory stimuli. However, whether this occurs in the context of natural communication, where stimulus significance is acquired through social interaction, has not been well investigated, perhaps because neural responses to species-specific vocalizations can be difficult to interpret within a systematic framework. The ultrasonic communication system between isolated mouse pups and adult females that either do or do not recognize the calls' significance provides an opportunity to explore this issue. We applied an information-based analysis to multi- and single unit data collected from anesthetized mothers and pup-naïve females to quantify how the communicative significance of pup calls affects their encoding in the auditory cortex. The timing and magnitude of information that cortical responses convey (at a 2-ms resolution) for pup call detection and discrimination was significantly improved in mothers compared to naïve females, most likely because of changes in call frequency encoding. This was not the case for a non-natural sound ensemble outside the mouse vocalization repertoire. The results demonstrate that a sensory cortical change in the timing code for communication sounds is correlated with the vocalizations' behavioral relevance, potentially enhancing functional processing by improving its signal to noise ratio.