CHD7 and SOX2 act in a common gene regulatory network during mammalian semicircular canal and cochlear development.

Inner ear morphogenesis requires tightly regulated epigenetic and transcriptional control of gene expression. CHD7, an ATP-dependent chromodomain helicase DNA-binding protein, and SOX2, an SRY-related HMG box pioneer transcription factor, are known to contribute to vestibular and auditory system development, but their genetic interactions in the ear have not been explored. Here, we analyzed inner ear development and the transcriptional regulatory landscapes in mice with variable dosages of Chd7 and/or Sox2. We show that combined haploinsufficiency for Chd7 and Sox2 results in reduced otic cell proliferation, severe malformations of semicircular canals, and shortened cochleae with ectopic hair cells. Examination of mice with conditional, inducible Chd7 loss by Sox2CreER reveals a critical period (~E9.5) of susceptibility in the inner ear to combined Chd7 and Sox2 loss. Data from genome-wide RNA-sequencing and CUT&Tag studies in the otocyst show that CHD7 regulates Sox2 expression and acts early in a gene regulatory network to control expression of key otic patterning genes, including Pax2 and Otx2. CHD7 and SOX2 directly bind independently and cooperatively at transcription start sites and enhancers to regulate otic progenitor cell gene expression. Together, our findings reveal essential roles for Chd7 and Sox2 in early inner ear development and may be applicable for syndromic and other forms of hearing or balance disorders.

Chromatin remodeler CHD7 is critical for cochlear morphogenesis and neurosensory patterning.

Epigenetic regulation of gene transcription by chromatin remodeling proteins has recently emerged as an important contributing factor in inner ear development. Pathogenic variants in CHD7, the gene encoding Chromodomain Helicase DNA binding protein 7, cause CHARGE syndrome, which presents with malformations in the developing ear. Chd7 is broadly expressed in the developing mouse otocyst and mature auditory epithelium, yet the pathogenic effects of Chd7 loss in the cochlea are not well understood. Here we characterized cochlear epithelial phenotypes in mice with deletion of Chd7 throughout the otocyst (using Foxg1Cre/+ and Pax2Cre), in the otic mesenchyme (using TCre), in hair cells (using Atoh1Cre), in developing neuroblasts (using NgnCre), or in spiral ganglion neurons (using ShhCre/+). Pan-otic deletion of Chd7 resulted in shortened cochleae with aberrant projections and axonal looping, disorganized, supernumerary hair cells at the apical turn and a narrowed epithelium with missing hair cells in the middle region. Deletion of Chd7 in the otic mesenchyme had no effect on overall cochlear morphology. Loss of Chd7 in hair cells did not disrupt their formation or organization of the auditory epithelium. Similarly, absence of Chd7 in spiral ganglion neurons had no effect on axonal projections. In contrast, deletion of Chd7 in developing neuroblasts led to smaller spiral ganglia and disorganized cochlear neurites. Together, these observations reveal dosage-, tissue-, and time-sensitive cell autonomous roles for Chd7 in cochlear elongation and cochlear neuron organization, with minimal functions for Chd7 in hair cells. These studies provide novel information about roles for Chd7 in development of auditory neurons.

How electrically evoked compound action potentials in chronically implanted guinea pigs relate to auditory nerve health and electrode impedance.

This study examined how multiple measures based on the electrically evoked compound action potential (ECAP) amplitude-growth functions (AGFs) were related to estimates of neural [spiral ganglion neuron (SGN) density and cell size] and electrode impedance measures in 34 specific pathogen free pigmented guinea pigs that were chronically implanted (4.9-15.4 months) with a cochlear implant electrode array. Two interphase gaps (IPGs) were used for the biphasic pulses and the effect of the IPG on each ECAP measure was measured ("IPG effect"). When using a stimulus with a constant IPG, SGN density was related to the across-subject variance in ECAP AGF linear slope, peak amplitude, and N1 latency. The SGN density values also help to explain a significant proportion of variance in the IPG effect for AGF linear slope and peak amplitude measures. Regression modeling revealed that SGN density was the primary dependent variable contributing to across-subject variance for ECAP measures; SGN cell size did not significantly improve the fitting of the model. Results showed that simple impedance measures were weakly related to most ECAP measures but did not typically improve the fit of the regression model.

Inner ear manifestations in CHARGE: Abnormalities, treatments, animal models, and progress toward treatments in auditory and vestibular structures.

The inner ear contains the sensory organs for hearing and balance. Both hearing and balance are commonly affected in individuals with CHARGE syndrome (CS), an autosomal dominant condition caused by heterozygous pathogenic variants in the CHD7 gene. Semicircular canal dysplasia or aplasia is the single most prevalent feature in individuals with CHARGE leading to deficient gross motor skills and ambulation. Identification of CHD7 as the major gene affected in CHARGE has enabled acceleration of research in this field. Great progress has been made in understanding the role of CHD7 in the development and function of the inner ear, as well as in related organs such as the middle ear and auditory and vestibular neural pathways. The goals of current research on CHD7 and CS are to (a) improve our understanding of the pathology caused by CHD7 pathogenic variants and (b) to provide better tools for prognosis and treatment. Current studies utilize cells and whole animals, from flies to mammals. The mouse is an excellent model for exploring mechanisms of Chd7 function in the ear, given the evolutionary conservation of ear structure, function, Chd7 expression, and similarity of mutant phenotypes between mice and humans. Newly recognized developmental functions for mouse Chd7 are shedding light on how abnormalities in CHD7 might lead to CS symptoms in humans. Here we review known human inner ear phenotypes associated with CHD7 pathogenic variants and CS, summarize progress toward diagnosis and treatment of inner ear-related pathologies, and explore new avenues for treatment based on basic science discoveries.

Vestibular dysfunction in the adult CBA/CaJ mouse after lead and cadmium treatment.

OBJECTIVES: The vestibular system allows the perception of position and motion and its dysfunction presents as motion impairment, vertigo and balance abnormalities, leading to debilitating psychological discomfort and difficulty performing daily tasks. Although declines and deficits in vestibular function have been noted in rats exposed to lead (Pb) and in humans exposed to Pb and cadmium (Cd), no studies have directly examined the pathological and pathophysiological effects upon the vestibular apparatus of the inner ear. METHODS: Eighteen young adult mice were exposed through their drinking water (3 mM Pb, 300 µM Cd, or a control treatment) for 10 weeks. Before and after treatment, they underwent a vestibular assessment, consisting of a rotarod performance test and a novel head stability test to measure the vestibulocolic reflex. At the conclusion of the study, the utricles were analyzed immunohistologically for condition of hair cells and nerve fibers. RESULTS: Increased levels of Pb exposure correlated with decreased head stability in space; no significant decline in performance on rotarod test was found. No damage to the hair cells or the nerve fibers of the utricle was observed in histology. CONCLUSIONS: The young adult CBA/CaJ mouse is able to tolerate occupationally-relevant Pb and Cd exposure well, but the correlation between Pb exposure and reduced head stability suggests that Pb exposure causes a decline in vestibular function. © 2016 Wiley Periodicals, Inc. Environ Toxicol, 2016. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 869-876, 2017.

microRNA-224 regulates Pentraxin 3, a component of the humoral arm of innate immunity, in inner ear inflammation.

microRNAs (miRNAs) are regulators of differentiation and development of inner ear cells. Mutations in miRNAs lead to deafness in humans and mice. Among inner ear pathologies, inflammation may lead to structural and neuronal defects and eventually to hearing loss and vestibular dysfunction. While the genetic factors of these pathways have not been defined, autoimmunity participates in these processes. We report that inflammatory stimuli in the inner ear induce activation of the innate immune system via miR-224 and pentraxin 3 (Ptx3). miR-224 is a transcriptional target of nuclear factor κB, a key mediator of innate immunity. Ptx3 is a regulator of the immune response. It is released in response to inflammation and regulated by nuclear factor κB. We show that miR-224 and Ptx3 are expressed in the inner ear and we demonstrate that miR-224 targets Ptx3. As a model of the innate immune response, we injected lipopolysaccharide into the scala tympani of mouse inner ears. This resulted in changes in the levels of miR-224 and Ptx3, in addition to activation of the complement system, as measured by immune cell infiltration and activated C3. This suggests that while miR-224 regulates Ptx3 under normal conditions, upon inflammation, both are recruited to offer a front line of defense in acting as responders to inflammation in the inner ear. miR-224 diminishes the innate immune response by down-regulating Ptx3 expression, while Ptx3 stimulates the innate immune response. An understanding of the molecular components of the inflammatory pathway may help develop therapeutics for reducing inflammation associated with inner ear injury.

CHD7 and retinoic acid signaling cooperate to regulate neural stem cell and inner ear development in mouse models of CHARGE syndrome.

CHARGE syndrome is a multiple congenital anomaly disorder that leads to life-threatening birth defects, such as choanal atresia and cardiac malformations as well as multiple sensory impairments, that affect hearing, vision, olfaction and balance. CHARGE is caused by heterozygous mutations in CHD7, which encodes an ATP-dependent chromatin remodeling enzyme. Identification of the mechanisms underlying neurological and sensory defects in CHARGE is a first step toward developing treatments for CHARGE individuals. Here, we used mouse models of Chd7 deficiency to explore the function of CHD7 in the development of the subventricular zone (SVZ) neural stem cell niche and inner ear, structures that are important for olfactory bulb neurogenesis and hearing and balance, respectively. We found that loss of Chd7 results in cell-autonomous proliferative, neurogenic and self-renewal defects in the perinatal and mature mouse SVZ stem cell niche. Modulation of retinoic acid (RA) signaling prevented in vivo inner ear and in vitro neural stem cell defects caused by Chd7 deficiency. Our findings demonstrate critical, cooperative roles for RA and CHD7 in SVZ neural stem cell function and inner ear development, suggesting that altered RA signaling may be an effective method for treating Chd7 deficiency.

Conditioning the cochlea to facilitate survival and integration of exogenous cells into the auditory epithelium.

The mammalian auditory epithelium (AE) cannot replace supporting cells and hair cells once they are lost. Therefore, sensorineural hearing loss associated with missing cells is permanent. This inability to regenerate critical cell types makes the AE a potential target for cell replacement therapies such as stem cell transplantation. Inserting stem cells into the AE of deaf ears is a complicated task due to the hostile, high potassium environment of the scala media in the cochlea, and the robust junctional complexes between cells in the AE that resist stem cell integration. Here, we evaluate whether temporarily reducing potassium levels in the scala media and disrupting the junctions in the AE make the cochlear environment more receptive and facilitate survival and integration of transplanted cells. We used sodium caprate to transiently disrupt the AE junctions, replaced endolymph with perilymph, and blocked stria vascularis pumps with furosemide. We determined that these three steps facilitated survival of HeLa cells in the scala media for at least 7 days and that some of the implanted cells formed a junctional contact with native AE cells. The data suggest that manipulation of the cochlear environment facilitates survival and integration of exogenously transplanted HeLa cells in the scala media.

siRNA targeting Hes5 augments hair cell regeneration in aminoglycoside-damaged mouse utricle.

Notch signaling is active during the development of mosaic epithelial sheets and during their turnover and regeneration. After the loss of hair cells in the mosaic sheet of the vestibular sensory epithelium, new hair cells can be spontaneously generated by transdifferentiation of supporting cells. This regenerative process involves downregulation of the Hes5 gene and is known to be limited and incomplete, especially when the lesion is severe. Here, we test whether further downregulation of Hes5 gene accomplished by the use of siRNA after a severe lesion induced by an aminoglycoside in the mouse utricle can enhance the transdifferentiation of supporting cells and lead to the increased production of new hair cells. We demonstrate that Hes5 levels in the utricle decreased after the application of siRNA and that the number of hair cells in these utricles was significantly larger than following control treatment. The data suggest that siRNA technology may be useful for inducing repair and regeneration in the inner ear and that the Notch signaling pathway is a potentially useful target for specific gene expression inhibition.

Combinatorial Atoh1, Gfi1, Pou4f3, and Six1 gene transfer induces hair cell regeneration in the flat epithelium of mature guinea pigs.

Flat epithelium (FE) is a condition characterized by the loss of both hair cells (HCs) and supporting cells and the transformation of the organ of Corti into a simple flat or cuboidal epithelium, which can occur after severe cochlear insults. The transcription factors Gfi1, Atoh1, Pou4f3, and Six1 (GAPS) play key roles in HC differentiation and survival in normal ears. Previous work using a single transcription factor, Atoh1, to induce HC regeneration in mature ears in vivo usually produced very few cells and failed to produce HCs in severely damaged organs of Corti, especially those with FE. Studies in vitro suggested combinations of transcription factors may be more effective than any single factor, thus the current study aims to examine the effect of co-overexpressing GAPS genes in deafened mature guinea pig cochleae with FE. Deafening was achieved through the infusion of neomycin into the perilymph, leading to the formation of FE and substantial degeneration of nerve fibers. Seven days post neomycin treatment, adenovirus vectors carrying GAPS were injected into the scala media and successfully expressed in the FE. One or two months following GAPS inoculation, cells expressing Myosin VIIa were observed in regions under the FE (located at the scala tympani side of the basilar membrane), rather than within the FE. The number of cells, which we define as induced HCs (iHCs), was not significantly different between one and two months, but the larger N at two months made it more apparent that there were significantly more iHCs in GAPS treated animals than in controls. Additionally, qualitative observations indicated that ears with GAPS gene expression in the FE had more nerve fibers than FE without the treatment. In summary, our results showed that co-overexpression of GAPS enhances the potential for HC regeneration in a severe lesion model of FE.

Mutations in Grxcr1 are the basis for inner ear dysfunction in the pirouette mouse.

Recessive mutations at the mouse pirouette (pi) locus result in hearing loss and vestibular dysfunction due to neuroepithelial defects in the inner ear. Using a positional cloning strategy, we have identified mutations in the gene Grxcr1 (glutaredoxin cysteine-rich 1) in five independent allelic strains of pirouette mice. We also provide sequence data of GRXCR1 from humans with profound hearing loss suggesting that pirouette is a model for studying the mechanism of nonsyndromic deafness DFNB25. Grxcr1 encodes a 290 amino acid protein that contains a region of similarity to glutaredoxin proteins and a cysteine-rich region at its C terminus. Grxcr1 is expressed in sensory epithelia of the inner ear, and its encoded protein is localized along the length of stereocilia, the actin-filament-rich mechanosensory structures at the apical surface of auditory and vestibular hair cells. The precise architecture of hair cell stereocilia is essential for normal hearing. Loss of function of Grxcr1 in homozygous pirouette mice results in abnormally thin and slightly shortened stereocilia. When overexpressed in transfected cells, GRXCR1 localizes along the length of actin-filament-rich structures at the dorsal-apical surface and induces structures with greater actin filament content and/or increased lengths in a subset of cells. Our results suggest that deafness in pirouette mutants is associated with loss of GRXCR1 function in modulating actin cytoskeletal architecture in the developing stereocilia of sensory hair cells.

The ATP-dependent chromatin remodeling enzyme CHD7 regulates pro-neural gene expression and neurogenesis in the inner ear.

Inner ear neurogenesis is positively regulated by the pro-neural bHLH transcription factors Ngn1 and NeuroD, but the factors that act upstream of this regulation are not well understood. Recent evidence in mouse and Drosophila suggests that neural development depends on proper chromatin remodeling, both for maintenance of neural stem cells and for proper neuronal differentiation. Here, we show that CHD7, an ATP-dependent chromatin remodeling enzyme mutated in human CHARGE syndrome, is necessary for proliferation of inner ear neuroblasts and inner ear morphogenesis. Conditional deletion of Chd7 in the developing otocyst using Foxg1-Cre resulted in cochlear hypoplasia and complete absence of the semicircular canals and cristae. Conditional knockout and null otocysts also had reductions in vestibulo-cochlear ganglion size and neuron number in combination with reduced expression of Ngn1, Otx2 and Fgf10, concurrent with expansion of the neural fate suppressor Tbx1 and reduced cellular proliferation. Heterozygosity for Chd7 mutations had no major effects on expression of otic patterning genes or on cell survival, but resulted in decreased proliferation within the neurogenic domain. These data indicate that epigenetic regulation of gene expression by CHD7 must be tightly coordinated for proper development of inner ear neuroblasts.

Nestin-expressing cells in the developing, mature and noise-exposed cochlear epithelium.

The auditory sensory epithelium in non-mammalian vertebrates can replace lost hair cells by transdifferentiation of supporting cells, but this regenerative ability is lost in the mammalian cochlea. Future cell-based treatment of hearing loss may depend on stem cell transplantation or on transdifferentiation of endogenous cells in the cochlea. For both approaches, identification of cells with stem cell features within the mature cochlea may be useful. Here we use a Nestin-ß-gal mouse to examine the presence of Nestin positive cells in the mature auditory epithelium, and determine how overstimulation of the ear impacts these cells. Nestin positive cells were found in the apical turn of the cochlea lateral to the outer hair cell area. This pattern of expression persisted into mature age. The area of Nestin positive cells was increased after the noise lesion. This increase in area coincided with an increase in expression of the Nestin mRNA. The data suggest that cells with potential stem cell features remain in the mature mammalian cochlea, restricted to the apical turn, and that an additional set of signals is necessary to trigger their contribution to cell replacement therapy in the ear. As such, this population of cells could serve to generate cochlear stem cells for research and potential therapy, and may be a target for treatments based on induced transdifferentiation of endogenous cochlear cells.

Cochlear Health and Cochlear-implant Function.

The cochlear implant (CI) is widely considered to be one of the most innovative and successful neuroprosthetic treatments developed to date. Although outcomes vary, CIs are able to effectively improve hearing in nearly all recipients and can substantially improve speech understanding and quality of life for patients with significant hearing loss. A wealth of research has focused on underlying factors that contribute to success with a CI, and recent evidence suggests that the overall health of the cochlea could potentially play a larger role than previously recognized. This article defines and reviews attributes of cochlear health and describes procedures to evaluate cochlear health in humans and animal models in order to examine the effects of cochlear health on performance with a CI. Lastly, we describe how future biologic approaches can be used to preserve and/or enhance cochlear health in order to maximize performance for individual CI recipients.

Role of auditory feedback for vocal production learning in the Egyptian fruit-bat.

Some species have evolved the ability to use the sense of hearing to modify existing vocalizations, or even create new ones. This ability corresponds to various forms of vocal production learning that are all possessed by humans, and independently displayed by distantly related vertebrates. Among mammals, a few species, including the Egyptian fruit-bat, would possess such vocal production learning abilities. Yet the necessity of an intact auditory system for the development of the Egyptian fruit-bat typical vocal repertoire has not been tested. Furthermore, a systematic causal examination of learned and innate aspects of the entire repertoire has never been performed in any vocal learner. Here we addressed these gaps by eliminating pups' sense of hearing at birth and assessing its effects on vocal production in adulthood. The deafening treatment enabled us to both causally test these bats vocal learning ability and discern learned from innate aspects of their vocalizations. Leveraging wireless individual audio recordings from freely interacting adults, we show that a subset of the Egyptian fruit-bat vocal repertoire necessitates auditory feedback. Intriguingly, these affected vocalizations belong to different acoustic groups in the vocal repertoire of males and females. These findings open the possibilities for targeted studies of the mammalian neural circuits that enable sexually dimorphic forms of vocal learning.

USH2A Gene Mutations in Rabbits Lead to Progressive Retinal Degeneration and Hearing Loss.

Purpose: Mutations in USH2A gene are responsible for the greatest proportion of the Usher Syndrome (USH) population, among which more than 30% are frameshift mutations on exon 13. A clinically relevant animal model has been absent for USH2A-related vision loss. Here we sought to establish a rabbit model carrying USH2A frameshift mutation on exon 12 (human exon 13 equivalent). Methods: CRISPR/Cas9 reagents targeting the rabbit USH2A exon 12 were delivered into rabbit embryos to produce an USH2A mutant rabbit line. The USH2A knockout animals were subjected to a series of functional and morphological analyses, including acoustic auditory brainstem responses, electroretinography, optical coherence tomography, fundus photography, fundus autofluorescence, histology, and immunohistochemistry. Results: The USH2A mutant rabbits exhibit hyper-autofluorescent signals on fundus autofluorescence and hyper-reflective signals on optical coherence tomography images as early as 4 months of age, which indicate retinal pigment epithelium damage. Auditory brainstem response measurement in these rabbits showed moderate to severe hearing loss. Electroretinography signals of both rod and cone function were decreased in the USH2A mutant rabbits starting from 7 months of age and further decreased at 15 to 22 months of age, indicating progressive photoreceptor degeneration, which is confirmed by histopathological examination. Conclusions: Disruption of USH2A gene in rabbits is sufficient to induce hearing loss and progressive photoreceptor degeneration, mimicking the USH2A clinical disease. Translational Relevance: To our knowledge, this study presents the first mammalian model of USH2 showing the phenotype of retinitis pigmentosa. This study supports the use of rabbits as a clinically relevant large animal model to understand the pathogenesis and to develop novel therapeutics for Usher syndrome.

Auditory hair cell replacement and hearing improvement by Atoh1 gene therapy in deaf mammals.

In the mammalian auditory system, sensory cell loss resulting from aging, ototoxic drugs, infections, overstimulation and other causes is irreversible and leads to permanent sensorineural hearing loss. To restore hearing, it is necessary to generate new functional hair cells. One potential way to regenerate hair cells is to induce a phenotypic transdifferentiation of nonsensory cells that remain in the deaf cochlea. Here we report that Atoh1, a gene also known as Math1 encoding a basic helix-loop-helix transcription factor and key regulator of hair cell development, induces regeneration of hair cells and substantially improves hearing thresholds in the mature deaf inner ear after delivery to nonsensory cells through adenovectors. This is the first demonstration of cellular and functional repair in the organ of Corti of a mature deaf mammal. The data suggest a new therapeutic approach based on expressing crucial developmental genes for cellular and functional restoration in the damaged auditory epithelium and other sensory systems.

Components of impedance in a cochlear implant animal model with TGFß1-accelerated fibrosis.

Outcomes of cochlear implantation are likely influenced by the biological state of the cochlea. Fibrosis is a pathological change frequently seen in implanted ears. The goal of this work was to investigate the relationship between fibrosis and impedance. To that end, we employed an animal model of extensive fibrosis and tested whether aspects of impedance differed from controls. Specifically, an adenovirus with a TGF-ß1 gene insert (Ad.TGF-ß1) was injected into guinea pig scala tympani to elicit rapid onset fibrosis and investigate the relation between fibrosis and impedance. We found a significant correlation between treatment and rate of impedance increase. A physical circuit model of impedance was used to separate the effect of fibrosis from other confounding factors. Supported by preliminary, yet nonconclusive, electron microscopy data, this modeling suggested that deposits on the electrode surface are an important contributor to impedance change over time.

Estimating health of the implanted cochlea using psychophysical strength-duration functions and electrode configuration.

It is generally believed that the efficacy of cochlear implants is partly dependent on the condition of the stimulated neural population. Cochlear pathology is likely to affect the manner in which neurons respond to electrical stimulation, potentially resulting in differences in perception of electrical stimuli across cochlear implant recipients and across the electrode array in individual cochlear implant users. Several psychophysical and electrophysiological measures have been shown to predict cochlear health in animals and were used to assess conditions near individual stimulation sites in humans. In this study, we examined the relationship between psychophysical strength-duration functions and spiral ganglion neuron density in two groups of guinea pigs with cochlear implants who had minimally-overlapping cochlear health profiles. One group was implanted in a hearing ear (N = 10) and the other group was deafened by cochlear perfusion of neomycin, inoculated with an adeno-associated viral vector with an Ntf3-gene insert (AAV.Ntf3) and implanted (N = 14). Psychophysically measured strength-duration functions for both monopolar and tripolar electrode configurations were then compared for the two treatment groups. Results were also compared to their histological outcomes. Overall, there were considerable differences between the two treatment groups in terms of their psychophysical performance as well as the relation between their functional performance and histological data. Animals in the neomycin-deafened, neurotrophin-treated, and implanted group (NNI) exhibited steeper strength-duration function slopes; slopes were positively correlated with SGN density (steeper slopes in animals that had higher SGN densities). In comparison, the implanted hearing (IH) group had shallower slopes and there was no relation between slopes and spiral ganglion density. Across all animals, slopes were negatively correlated with ensemble spontaneous activity levels (shallower slopes with higher ensemble spontaneous activity levels). We hypothesize that differences in strength-duration function slopes between the two treatment groups were related to the condition of the inner hair cells, which generate spontaneous activity that could affect the across-fiber synchrony and/or the size of the population of neural elements responding to electrical stimulation. In addition, it is likely that spiral ganglion neuron peripheral processes were present in the IH group, which could affect membrane properties of the stimulated neurons. Results suggest that the two treatment groups exhibited distinct patterns of variation in conditions near the stimulating electrodes that altered detection thresholds. Overall, the results of this study suggest a complex relationship between psychophysical detection thresholds for cochlear implant stimulation and nerve survival in the implanted cochlea. This relationship seems to depend on the characteristics of the electrical stimulus, the electrode configuration, and other biological features of the implanted cochlea such as the condition of the inner hair cells and the peripheral processes.