Electrical impedance measurements of cochlear structures using the four-electrode reflection-coefficient technique

In individuals with severe-to-profound hearing loss, cochlear implants (CIs) bypass normal inner ear function by applying electrical current directly into the cochlea, thereby stimulating surviving auditory nerve fibers. Although cochlear implants are able to restore some auditory sensation, they are far from providing normal hearing. It has been estimated that up to 75% of the current injected via a CI is shunted along scala tympani and is not available to stimulate auditory neurons. The path of the injected current and the consequent population of stimulated spiral ganglion cells are dependent upon the positions of the electrode contacts within the cochlea and the impedances of cochlear structures. However, characterization of the current path remains one of the most critical, yet least understood, aspects of cochlear implantation. In particular, the impedances of cochlear structures, including the modiolus, are either unknown or based upon estimates derived from circuit models. Impedance values for many cochlear structures have never been measured. By combining the hemicochlea preparation, a cochlea cut in half along its mid-modiolar plane, and the four-electrode reflection-coefficient technique, impedances can be measured for cochlear tissues in a cochlear cross section including the modiolus. Advantages and disadvantages of the method are discussed in detail and electrical impedance measurements obtained in the gerbil hemicochlea are presented. The resistivity values for the cochlear wall in Ωcm are, 528 (range: 432–708) for scala media 3rd turn, 502 (range: 421–616) for scala tympani 3rd turn and scala vestibuli 2nd turn, 627 (range: 531–759) for scala media 2nd turn, 434 (range: 353–555) for scala tympani 2nd turn and scala vestibuli basal turn, 434 (range: 373–514) for scala media basal turn, and 590 (range: 546–643) for scala tympani basal turn. The resistivity was 455 Ωcm (range: 426–487) for the modiolus.     

Regenerated Synapses Between Postnatal Hair Cells and Auditory Neurons

Regeneration of synaptic connections between hair cells and spiral ganglion neurons would be required to restore hearing after neural loss. Here we demonstrate by immunohistochemistry the appearance of afferent-like cochlear synapses in vitro after co-culture of de-afferented organ of Corti with spiral ganglion neurons from newborn mice. The glutamatergic synaptic complexes at the ribbon synapse of the inner hair cell contain markers for presynaptic ribbons and postsynaptic densities. We found postsynaptic density protein PSD-95 at the contacts between hair cells and spiral ganglion neurons in newly formed synapses in vitro. The postsynaptic proteins were directly facing the CtBP2-positive presynaptic ribbons of the hair cells. BDNF and NT-3 promoted afferent synaptogenesis in vitro. Direct juxtaposition of the postsynaptic densities with the components of the preexisting ribbon synapse indicated that growing fibers recognized components of the presynaptic sites. Initiation of cochlear synaptogenesis appeared to be influenced by glutamate release from the hair cell ribbons at the presynaptic site since the synaptic regeneration was impaired in glutamate vesicular transporter 3 mutant mice. These insights into cochlear synaptogenesis could be relevant to regenerative approaches for neural loss in the cochlea.     

新霉素致发育中大鼠毛细胞损伤的电镜观察

目的应用扫描电镜研究氨基甙类抗生素导致出生后发育过程中大鼠耳蜗毛细胞的损伤病理变化。方法７天龄ＳＤ大鼠肌肉注射８０ｍｇｋｇ－１ｄ－１新霉素连续８天,停药后７天处死动物制备样本进行扫描电镜观察。结果新霉素可造成出生后发育过程中的耳蜗毛细胞严重损伤,表现为底回和钩回三排外毛细胞全部损伤缺失,损伤病变累及顶回的外毛细胞,底回和钩回的内毛细胞出现纤毛脱落和细胞损伤丢失;外毛细胞损伤缺失的部位由顶部具有微绒毛的多边形细胞取代,该细胞形态与胚胎发育早期的毛细胞相似;毛细胞损伤区域未发现再生的毛细胞。结论出生后发育过程中大鼠耳蜗毛细胞对氨基甙类抗生素的敏感性较高,表面有微绒毛的多边形细胞在毛细胞损伤区域出现提示听觉感觉上皮细胞有再生的倾向。     

Cochlear hair cell loss in ears with cholesteatomas. Scanning electron microscopy study

Cochleas from 16 Mongolian gerbils with spontaneous aural cholesteatomas, and four of similar age without cholesteatomas, were examined by scanning electron microscopy to quantify cochlear hair cell loss. Loss of hair cell stereocilia was found in all ears with cholesteatomas and was significantly increased when compared with uninvolved ears from animals of similar age. The hair cell loss associated with gerbilline cholesteatomas appeared to be most marked in the middle turn of the cochlea and increased in severity with increasing size of the cholesteatomas. Outer hair cells were affected more than inner hair cells. Inner and outer hair cell loss was not significantly different in infected cholesteatomas versus sterile cholesteatomas. The greater damage to hair cells at the middle turn compared to the basal turn suggests that these losses may be the result of some agent acting through the cochlear wall rather than through the round window.     

Inner ear changes in the ferret model for Reye's syndrome

The acute effects of influenza B, aspirin, and hyperammonemia on the inner ear were examined using the ferret model for Reye's syndrome. Histopathologic examination revealed varying degrees of vacuolization in both the sensory and nonsensory endolymphatic tissues of the cochlear and vestibular membranous labyrinth. The secretory epithelial cells of the stria vascularis and the vestibular dark cells appeared to be more severely affected, as demonstrated by the presence of an extensive number of intracytoplasmic vacuoles. The cells of the perilymphatic tissues appeared to be unaltered. Although all neuroepithelial regions of the inner ear appeared to be altered, an unexpected observation was that only the inner hair cells of the organ of Corti were pathologically affected. These results suggest that metabolic and virus-induced derangements may alter inner ear microhomeostasis in patients with Reye's syndrome and may potentially result in loss of hearing in such patients.     

Aspartate aminotransferase immunoreactivity in cochlea of guinea pig

The distribution of aspartate aminotransferase-like immunoreactivity in the cochlea of the guinea pig was studied at the light microscopy level. Indirect immunofluorescence histochemistry using antisera against cytoplasmic aspartate aminotransferase prepared from pig heart was applied to surface preparations of the organ of Corti and cryostat sections of the cochlea. In the modiolus, immunofluorescence was localized to spiral ganglion cells and myelinated fibers of the auditory nerve and intraganglionic spiral bundles. In the organ of Corti, immunofluorescence was seen in upper tunnel crossing fibers and at the base of outer hair cells, following a distribution similar to that of the efferent innervation of the outer hair cells. Weak immunofluorescence was seen in the inner spiral bundle and tunnel spiral bundle, but was not present in all preparations. Immunofluorescence was not seen in inner hair cells, nor at the base of inner hair cells, and may have been absent from outer hair cells.It is concluded that spiral ganglion cells and myelinated auditory nerve axons contain aspartate aminotransferase-like immunoreactivity; such immunoreactivity has previously been determined in auditory nerve endings in the cochlear nucleus. Olivocochlear neurons that innervate outer hair cells also contain such immunoreactivity while other cochlear efferents contain little or none.     

耳蜗毛细胞和螺旋神经节及其神经纤维的联合定量观察

目的：将耳蜗基底膜取材技术与耳蜗切片技术相结合以观察同一耳蜗中几个主要结构在病理发展中的相互关系。方法：耳蜗铺片用于定量观察毛细胞，骨性螺旋板切片用于定量观察缰孔内的神经纤维，中轴切片用于定量观察螺旋神经节。结果：将卡铂耳中毒灰鼠的耳蜗标本与正常灰鼠进行比较，显示内毛细胞，神经纤维和螺旋神经节的损失百分比基本一致。结论：卡铂对内毛细胞，神经纤维和螺旋神经节都有破坏作用，至于在卡铂病变早期，究竟哪一     

The effects of kainic acid on the cochlear ganglion of the rat

The effects of locally applied kainic acid on cells and fibers in the rat cochlea were examined in a quantitative and ultrastructural study. Doses of 5 nM per microliter of artificial perilymph destroyed part of the spiral ganglion type I cell population, with no ototoxic effects on cochlear hair cells or supporting cells. Type II cells also appeared unaffected. A quantitative evaluation of the cell loss with the 5 nM dosage showed that 34% of spiral ganglion neurons were lost 10 days after treatment. Doses of 20 nM per microliters and 40 nM per microliters did not result in increasing neuronal loss. This differential toxicity could reflect the presence of a sub-population of spiral ganglion cells with an increased number of KA receptors.     

Spiral Ligament Pathology: A Major Aspect of Age-Related Cochlear Degeneration in C57BL/6 Mice

Data from systematic, light microscopic examination of cochlear histopathology in an age-graded series of C57BL/6 mice (1.5-15 months) were compared with threshold elevations (measured by auditory brain stem response) to elucidate the functionally important structural changes underlying age-related hearing loss in this inbred strain. In addition to quantifying the degree and extent of hair cell and neuronal loss, all structures of the cochlear duct were qualitatively evaluated and any degenerative changes were quantified. Hair cell and neuronal loss patterns suggested two degenerative processes. In the basal half of the cochlea, inner and outer hair cell loss proceeded from base to apex with increasing age, and loss of cochlear neurons was consistent with degeneration occurring secondary to inner hair cell loss. In the apical half of the cochlea with advancing age, there was selective loss of outer hair cells which increased from the middle to the extreme apex. A similar gradient of ganglion cell loss was noted, characterized by widespread somatic aggregation and demyelination. In addition to these changes in hair cells and their innervation, there was widespread degeneration of fibrocytes in the spiral ligament, especially among the type IV cell class. The cell loss in the ligament preceded the loss of hair cells and/or neurons in both space and time suggesting that fibrocyte pathology may be a primary cause of the hearing loss and ultimate sensory cell degeneration in this mouse strain.     

Tuberculous meningitis-induced unilateral sensorineural hearing loss: a temporal bone study

The relationship between meningitis and sensorineural hearing loss (SNHL) has long been studied. Many histopathological studies of animal models and human temporal bones with respect to bacterial meningitis have been carried out. However, the relationship between SNHL and tuberculous meningitis was seldom addressed and the pathophysiology remains unclear. We carried out temporal bone studies on material from a 22-year-old patient who developed a right unilateral SNHL before dying from tuberculous meningitis. The histopathological findings for the right temporal bone were as follows: (1) inflammation mainly appeared in the internal auditory canal, modiolus and Rosenthal's canal and extended to the osseous spiral ligament, whereas the perilymphatic spaces were less involved; (2) the organ of Corti, cochlear nerve fibres and spiral ganglion cells were severely degenerated, particularly in the basal and middle turns; (3) the contralateral side (for which the patient had no complaints) showed an inner space free from inflammation, but some granulomatous formations were observed in the middle ear cavity. We conclude that the modiolus and cochlear aqueduct are the main routes for the spread of infection from the meninges to the inner ear. The progression of hearing loss resembles that of bacterial meningitis and shares attributes of retrocochlear SNHL.     

Aminoglycoside ototoxicity: a human temporal bone study.

OBJECTIVE: Hearing loss after aminoglycoside administration has been thought to result primarily from hair cell injury. The purpose of the study was to determine the potential for direct injury of spiral ganglion cells and hair cells in cases of documented human aminoglycoside ototoxicity. STUDY DESIGN: Retrospective case review. METHODS: The clinical course of two individuals with aminoglycoside ototoxicity are documented, including the details of administration of tobramycin and other ototoxic medication and serial audiograms. The temporal bones were processed, and the cochlear elements quantified. RESULTS: Histopathological study of the temporal bones from the individuals in the study demonstrated reduction of both ganglion cell and hair cell populations. Spiral ganglion cell loss was not necessarily subadjacent to areas of hair cell loss in cases of aminoglycoside ototoxicity. Instead, spiral ganglion cell reduction may be present in segments of the cochlea with normal-appearing hair cells. CONCLUSIONS: The study suggests that aminoglycoside antibiotics can injure spiral ganglion cells directly, as well as hair cells. Thus, the characteristic hearing loss of ototoxicity can result from degeneration of either cochlear element.     

Expression and localization of K channels KCNQ2 and KCNQ3 in the mammalian cochlea

KCNQ1 and KCNQ4 voltage-gated potassium channel subunits play key roles in hearing. Other members of the KCNQ family also encode slow, low voltage-activated K(+) M currents. We have previously reported the presence of M-like K(+) currents in sensory hair cells, and expression of Kcnq family genes in the cochlea. Here, we describe Kcnq2/3 gene expression and distribution of M channel subunits KCNQ2 and 3 in the cochlea. By using RT-PCR, we found expression of Kcnq2 in the modiolus and organ of Corti, while Kcnq3 expression was also detected in the cochlear lateral wall. Five alternative splice variants of the Kcnq2 gene, one of which has not been reported previously, were identified in the rat cochlea. KCNQ2 and KCNQ3 immunoreactivities were observed in spiral ganglion auditory neurons. In addition, the unmyelinated parts of the nerve fibers innervating hair cells and synaptic regions under hair cells showed KCNQ2 immunoreactivity. KCNQ3 immunoreactivity was also prominent in spiral ganglion satellite cells. These findings suggest that cochlear M channels play important roles in regulation of cellular excitability and maintenance of cochlear K(+) homeostasis in the auditory system.     

Spiral ganglion cell density in young and old gerbils

The Mongolian gerbil, like other mammalian species, has a decreased number of spiral ganglion cells as a function of age. This loss of cells was first seen in 24- to 30-month old animals in the basal end of the ganglion. In the oldest individuals the apical end of the ganglion was also affected. There were approximately 15-25% fewer cells in the affected areas in the 36- to 42-month old animals. In the oldest animals degeneration of the stria vascularis was seen in the apical turn and some degenerative changes in the organ of Corti were seen throughout the length of the cochlear duct. The aging pattern in the gerbil cochlea, is similar to that described for other species. Vacuoles, previously described in the gerbil cochlear nucleus, were also seen in the auditory nerve within the modiolus, but central to the Schwann-glial border in all animals. Vacuoles were not present within the spiral ganglion or the peripheral processes of the ganglion cells. Because the ganglion cell axons should be similar on either side of the Schwann-glial border, but the vacuoles were confined to the central nervous system, it is concluded that the degenerative process affects glial cells as opposed to neurons.     

The effect of organ of corti loss on ganglion cell survival in humans.

HYPOTHESIS: Severe spiral ganglion cell loss does not necessarily follow loss of hair cells or supporting cells in humans. BACKGROUND: Despite some publications to the contrary, statements that loss of hair cells and/or supporting cells of the organ of Corti results in a severe loss of spiral ganglion cells in humans still appear in the literature, especially in respect to cochlear implants. This assumption is apparently based on studies in animals or cell culture and not from studies of human temporal bones. METHODS: Morphological analysis of archival temporal bones with microscopic and statistical analysis of ganglion cell, hair cell, and supporting cell populations was performed in 33 ears with total hearing losses of varying causes and durations of deafness. None of the ears had remaining hair cells. Six ears had had cochlear implants. RESULTS: Ganglion cell counts ranging from 2,889 to 34,299 and the corresponding percentage of remaining ganglion cells based on age-normative data were not significantly related to the duration of hearing loss (r = -0.13 and 0.02, respectively, p > 0.05) or to remaining supporting cell populations (r's from 0.15 to 0.27, p > 0.05). More than half of ears (51.5%) had ganglion cell counts within two standard deviations of age-normative means. Mean ganglion cell counts and percentage of remaining ganglion cells of ears with surviving peripheral processes (dendrites) did not differ significantly from those of ears with no peripheral processes. CONCLUSION: The loss of hair and supporting cells in the organ of Corti in humans does not necessarily result in as significant a loss of spiral ganglion cells as has been reported animals. In fact, our results suggest that ganglion cell loss may be a primary concomitant loss due to the disease process.     

Kanamycin and bumetanide ototoxicity: Anatomical,physiological and behavioral correlates

Severe hair-cell degeneration and cochlear dysfunction was observed in chinchillas examined at 60 days (or longer) after administration of a single injection of 150 mg/kg kanamycin, followed 2 h later by a single injection of 20 mg/kg bumetanide. Outer hair cells in the cochlear base were most severely affected. While inner and outer hair-cell loss was common, some animals showed large regions along the basilar membrane where almost all inner hair cells were present and almost all outer hair cells were absent. Wherever areas of complete degeneration of the organ of Corti occurred, a small, diffuse population of nerve fibers within the spiral lamina was always present. Single-unit tuning curves correlated best with anatomical observations, compared with the other functional measures of auditory sensitivity that were obtained (behavioral audiogram and compound action potential thresholds). Results indicated that behavioral detection of auditory stimuli is relatively independent of innervation density as long as a few inner hair cells are present. Thus, the cross-fiber threshold envelope of the single-unit tuning curves appeared very similar to the behavioral audiogram.     

Histopathologic features of the temporal bone in usher syndrome type I

Temporal bones of 2 patients with Usher syndrome type I were examined using light microscopy. In both patients, findings from histopathologic examination of the cochlea were characterized by degeneration of the organ of Corti, which was most marked in the basal turn, atrophy of the stria vascularis, and a decrease in the number of spiral ganglion cells. The cochlear nerve appeared to be diminished. The sensory epithelium of the saccular and utricular maculae of patient 1 was normal for age. The left temporal bone of patient 2, classified as Usher syndrome genetic subtype USH1D or USH1F, demonstrated the typical signs of severe cochleosaccular degeneration. Present cases and cases from the literature were reviewed in search of an explanation for the above-described differences in histologic findings.     

Reduction in Sharpness of Frequency Tuning but not Endocochlear Potential in Aging and Noise-Exposed BALB/cJ Mice

Schuknecht proposed categories for human age-related hearing loss (ARHL) based upon whether the primary degeneration involves the organ of Corti (sensory ARHL), spiral ganglion cells (neural), stria vascularis (strial), or a combination of these (mixed). Genetically standardized mouse ARHL models can help validate Schuknecht's framework and clarify the underlying cellular processes. Much recent work has focused on the mouse Ahl locus, which promotes both ARHL and noise-induced hearing loss. On the C57BL/6 inbred background, Ahl has been associated with degeneration of organ of Corti, afferent neurons, and stria vascularis/spiral ligament, suggesting that it promotes mixed (sensory/neural/strial) ARHL. Some cochlear degeneration in C57BL/6 mice could be caused by genes other than Ahl, however. The question of what constitutes Ahl-related pathology can be addressed by comparing C57BL/6 mice with other strains that carry the same allele, including BALB/c substrains. We examined the effects of aging and broadband noise exposure in inbred BALB/cJ mice (1.5–13.0 mos) using measures of frequency tuning (compound action potential tuning curves) (CAPTCs), strial function (endocochlear potential recording, EP), and light microscopy. Aging and noise led to generally similar physiological and anatomical changes. Reductions in sensitivity and sharpness of frequency tuning were not consistently linked to hair cell loss, reduction in the EP, or changes in the lateral wall. Instead they appeared best explained by alterations in supporting cells in the basal half of the cochlear and in the spiral limbus in the apex. These results emphasize the importance of cell types other than hair cells in cochlear pathology. They also indicate that Ahl does not necessarily promote a strial form of ARHL.     

Distribution and significance of norepinephrine in the lateral cochlear wall of pigmented and albino rats

Picogram quantities of norepinephrine were found in cochlear regions of pigmented and non-pigmented rats. These regions of the cochlea were the modiolus, organ of Corti-osseous spiral lamina and the lateral cochlear wall. The content of norepinephrine in the modiolus and lateral cochlear wall of the pigmented rat was significantly greater than that in areas of the non-pigmented rat. In contrast, there was no statistical difference between the norepinephrine content of the organ of Corti-osseous spiral lamina region of the pigmented rat and that of the albino rat. Since a major difference between the pigmented and albino rats is the presence of melanin-containing melanocytes in the modiolus and lateral cochlear wall region of the pigmented animals, it is possible that norepinephrine is stored in cochlear melanocytes.     

In Vitro and In Vivo Assessment of the Ability of Adeno‐Associated Virus–Brain‐Derived Neurotrophic Factor to Enhance Spiral Ganglion Cell Survival Following Ototoxic Insult

Objectives/Hypothesis Auditory dysfunction following ototoxic insult results from loss of cochlear hair cells. Secondary degeneration of auditory neurons ensues from withdrawal of neurotrophic support from hair cells and can be prevented with administration of neurotrophins. Administration of adeno‐associated virus containing the gene for brain‐derived neurotrophic factor will promote spiral ganglion neuron survival after the destruction of hair cells. Methods Prevention of aminoglycoside‐induced spiral ganglion neuron loss through the expression of brain‐derived neurotrophic factor mediated by means of the adeno‐associated virus was tested in vitro in cochlear explants and in vivo in mammalian cochlea. Results Neuronal survival was significantly enhanced in adeno‐associated virus–brain‐derived neurotrophic factor transfected rat cochlear explants compared with control samples (30% vs. 19%, P <.05) following exposure to aminoglycoside. Following deafening with aminoglycoside and loop diuretic and introduction of adeno‐associated virus–brain‐derived neurotrophic factor through osmotic minipump, the experimental group of animals infused with adeno‐associated virus–brain‐derived neurotrophic factor displayed enhanced spiral ganglion neuron survival in the basal turn of the cochlea when compared with the control group infused with adeno‐associated virus containing green fluorescent protein reporter gene. Conclusions Administration of adeno‐associated virus–brain‐derived neurotrophic factor enhances spiral ganglion neuron survival following ototoxic exposure in vitro and in vivo. These studies lay the groundwork for further exploration of its application as an adjunct therapy for patients undergoing cochlear implantation because the success of implantation depends directly on the population of neurons available for electrical stimulation.