Some electrical circuit properties of the organ of Corti. I. Analysis without reactive elements

A simplified network model of the organ of Corti is analyzed with the assumption of parametric excitation via resistance changes in the hair cells' apical membrane. Pertinent network variables (intracellular resting and receptor potentials, cellular input resistance, extracellular potentials) depend on the ratios of basal (perilymphatic face) and apical (endolymphatic face) receptor cell resistances, denoted as shape factors. In the Appendix two methods are suggested for the computation of shape factors; both are based on the geometrical properties of hair cells. Various electrical quantities computed on the basis of shape factors are consistent with recent recordings from third turn inner and outer hair cells (Dallos et al. (1982): Science 218, 582-584). The model provides a plausible explanation for the experimentally observed discrepancy between inner and outer hair cell resting and receptor potentials. One potentially significant result of the analysis is the demonstration that since shape factors for outer hair cells are probably longitudinally graded, so must be all cellular electrical characteristics. Another interesting finding is that electrical interaction among neighboring hair cells is unlikely. A large-signal analysis of the circuit demonstrates that even in the absence of a non-linear input, the parametrically excited circuit itself generates pronounced distortion. The most significant consequence of this nonlinearity is a response asymmetry in which the depolarizing phase is greater than the hyperpolarizing one. Thus the circuit nonlinearity may, at least in part, account for the large positive d.c. response seen in both types of receptor cell (Dallos et al. (1982): Science 218, 582-584; Russell and Sellick (1978): J. Physiol. Lond. 284, 261-290).     

Cochlear transduction: an integrative model and review

A model for cochlear transduction is presented that is based on considerations of the cell biology of its receptor cells, particularly the mechanisms of transmitter release at recepto-neural synapses. Two new interrelated hypotheses on the functional organization of the organ of Corti result from these considerations, one dealing with the possibility of electrotonic interaction between inner and outer hair cells and the other with a possible contributing source to acoustic emissions of cochlear origin that results from vesicular membrane turnover.     

Serial Section Reconstruction of the Neural Poles of Hair Cells in the Human Organ Of Corti. I. Inner Hair Cells

Study of the anatomy of the cochlea, and in particular the morphology of synaptic relationships between hair cells and cochlear neurons, is essential for elucidation of the mechanisms of transduction of mechanical acoustic signals into electrical neural events. Because considerable gaps remain in our understanding of the microscopic anatomy of these synapses, particularly in the human, a reconstruction of the neural pole of inner hair cells of the human organ of Corti was performed. The data are based on 526 serial sections from the basal turn (10 mm region) and 356 serial sections from the middle turn (26 mm region). This provided complete data on 3 and partial data on 5 inner hair cells. Afferent terminals on inner hair cells were variable in size, ranging 1 to 20 µm in diameter. Branching of large fibers to produce multiple terminals innervating from 1 to 3 inner hair cells was common. Each inner hair cell received approximately 6 to 8 different nerve terminals. In addition, each terminal possessed a variable number of synaptic contacts. Junctional membrane specialization consisted of synapses, desmosomes, coated vesicles and arrays of microtubules and membrane cisternae. Specialization at synapses consisted of asymmetrical membrane thickening. At inner hair cells the postsynaptic membrane was thicker than the presynaptic membrane. Eighty-three percent of synapses had presynaptic bodies. Vesiculated efferent terminals synapsed on afferent fibers at the base of inner hair cells, but never directly on the inner hair cell. These anatomical data demonstrate distinct differences between the human and animal inner ear, which are important in the interpretation of neurophysiological data in animals and the formulation of hypotheses that involve assumptions crossing species.     

蛋氨酸对离体培养小鼠耳蜗毛细胞顺铂耳毒性的拮抗作用

目的 在培养基中加入顺铂,建立离体培养小鼠耳蜗毛细胞的顺铂损害模型,探讨蛋氨酸对毛细胞的保护作用.方法 32只出生后2 d昆明小鼠,取出耳蜗基底膜,分为4组,每组16个样本,分别用无血清培养液、含顺铂的无血清培养液、含顺铂+蛋氨酸的无血清培养液以及含蛋氨酸的无血清培养液进行离体培养.采用肌球蛋白Ⅵ(myosinⅥ)免疫荧光染色,通过激光共聚焦显微镜、光镜、毛细胞计数等方法,观察蛋氨酸对顺铂引起的耳蜗毛细胞损伤的保护作用.结果 对照组和蛋氨酸组基底膜内、外毛细胞未见损伤缺失;顺铂组耳蜗基底膜内、外毛细胞均有不同程度的损伤,与对照组和蛋氨酸组比较,差异均具有统计学意义(P值均＜0.05);顺铂+蛋氨酸组基底膜内、外毛细胞的损伤程度与顺铂组相比明显减轻,差异具有有统计学意义(t内毛细胞=3.929、t外毛细胞=8.582,P值均＜0.05).结论 顺铂能损伤离体培养的小鼠耳蜗基底膜毛细胞,蛋氨酸可以在一定程度上拮抗顺铂对毛细胞的损伤.

Abstract：

Objective To establish an in vitro model of mouse cochlear basilar membrane impairment using cisplatin, and observe the protective effect of methionine on the hair cells. Methods The cochlear basilar membrane samples of thirty two Kunming mice were harvested on the 2nd day after birth and randomly divided into four groups. Each group had 16 samples. Overnight preincubation the cochlear organ followed by appropriate treatment respectively as follows: the serum-free culture medium, the serum-free culture medium with methionine and cisplatin, the cisplatinum-containing serum-free culture medium, and the methionine-containing serum-free culture medium. The protective effect of methionine for injury of cochlea hair cells induced by cisplatin was observed by myosin-Ⅵ immunofluorescence, lightmicroscop,laser confocal scanning microscope and hair cells counting. Results The outer hair cells(OHC) and inner hair cells(IHC) of control group and methionine group were not damaged. The outer and inner hair cells of cisplatin group were damaged in various degree, and had remarkable difference compared with control group and methionine group(P ＜ 0. 05). The outer hair cells and inner hair cells of cisplatin + methionine group were damaged less than the cisplatin group with remarkable difference (tIHC = 3. 929, tOHC = 8. 582, P ＜0. 05). Conclusions Cisplatinum could damage the cochlear hair cells of the basal membrane in Kunming mice. Methionine might protect against cisplatin's damage on the cochlear hair cells.     

Serial section reconstruction of the neural poles of hair cells in the human organ of corti. II. outer hair cells

Study of the anatomy of the cochlea, and in particular the morphology of synaptic relationships between hair cells and cochlear neurons, is essential for elucidation of the mechanisms of transduction of mechanical acoustic signals into electrical neural events. Because considerable gaps remain in our understanding of the microscopic anatomy of these synapses, particularly in the human, a reconstruction of the neural pole of outer hair cells of the human organ of Corti was performed. The data are based on 577 serial sections from the basal turn and 368 sections from the middle turn. This provided complete data on 11 and partial data on 9 outer hair cells. Terminal size of afferent fibers on outer hair cells was much more uniform than that found at the base of inner hair cells. Only small bouton-like terminals were found. Branching of afferent fibers was also seen at the base of outer hair cells. Each outer hair cell received approximately two to eight afferent nerve terminals. Multiple synaptic contacts between a single afferent terminal and an outer hair cell were common. Junctional membrane specialization consisted of synapses, desmosomes, coated vesicles and arrays of microtubules and membrane cisternae. Specialization at synapses consisted of asymmetrical membrane thickening. At outer hair cells the presynaptic membrane was thicker than the postsynaptic membrane. At inner hair cells the converse was true. At outer hair cells 35% of synapses had presynaptic bodies, compared to 83% of synapses at inner hair cells. Reciprocal synapses, with both hair cell to neuron and neuron to hair cell polarities, were found only on outer hair cells. Vesiculated efferent terminals were common at the base of outer hair cells. Both axosomatic and axodendritic efferent synapses were found. In addition, the same efferent fibers were found to synapse both on an outer hair cell and on an afferent dendrite. One example of a probable dendro-dendritic synapse in the outer spiral bundle is presented.     

Anatomical correlates of impulse noise-induced mechanical damage in the cochlea

Changes in the surface morphology of the organ of Corti in the chinchilla were studied following exposure to blast waves at 160 dB peak SPL. The generation and development of a mechanically induced lesion on the organ of Corti was followed over a period of 30 days using scanning electron microscopy. The most prominent feature of the lesion was the complete separation of a 5–7 mm strip of the sensory epithelia consisting of outer hair cells, Deiter cells and Hensen cells from the reticular lamina and the basilar membrane. The inner hair cells in this same area survived for several days in a remarkably normal condition. A spectrum of ciliary changes was observed on the inner and outer hair cells that differ somewhat from those commonly reported following continuous noise exposure. Some of the observed changes in morphology can be related to a variety of inferred mechanical events on the basilar membrane.     

Some electrical circuit properties of the organ of Corti. II. Analysis including reactive elements

The simplified network model of the organ of Corti-hair cell complex that was studied in previous publications (Dallos, P. (1983): In: Hearing - Physiological Bases and Biophysics, pp. 32-37, editors: R. Klinke and R. Hartmann; Dallos, P. (1983): Hearing Res. 12, 89-119) is extended to include the capacitive reactances of hair cell 12, 89-119) is extended to include the capacitive reactances of hair cell membranes. With the assumption that the circuit is parametrically excited by resistance changes of the apical hair cell membrane, intracellular responses for outer (eO) and inner (eI) hair cells and the organ of Corti potential (eOC) are computed for the linear case. In order that frequency-dependent effects be primarily determined by the network properties of the system, the eO/eOC and eI/eOC ratios are considered. The amplitude and phase dependence of these computed functions is compared to corresponding experimental results from the third turn of the guinea pig's cochlea. The eO/eOC function is predicted to be a simple low-pass filter whose cutoff frequency (1250 Hz) is determined by the parallel resistance-capacitance circuit of the cell's baso-lateral membrane. This circuit prediction is in excellent agreement with the data. The eI/eOC function is again derived as a low-pass filter. The cutoff frequency is obtained as 472 Hz, which corresponds to an RC membrane time constant of 0.34 ms. Experimental data are widely divergent from this predicted pattern. In order to bring the theoretical and experimental results in agreement, two steps are necessary. First, a differentiation (i.e., multiplication by j omega) needs to be included in the input to inner hair cells to account for their velocity dependence [4,10]. Second, an underdamped complex pole, located above the cell's best frequency must be introduced. The velocity dependence and the resonance represented by the underdamped pole-pair indicate that the electrical response pattern of inner hair cells is more complex than that of outer hair cells.     

Degeneration of the organ of Corti following intravenous administration of cisplatin.

The development of acute morphological changes in the cochlea was studied in guinea pigs given one intravenous high-dose injection of cisplatin. In the light microscope three major stages of degeneration in the organ of Corti could be recognized: 1) an initial swelling of the Hensen's cells and protrusion of the Deiters' cells into the space of Nuel enclosing the outer hair cells, 2) a gradual degeneration of the outer hair cells together with a vacuolization in the region of the base of the inner hair cells, 3) a collapse of the Reissner's membrane and the entire organ of Corti with different degrees of damage to the inner hair cells. Sporadic bulging of the marginal cells of the stria vascularis into the endolymphatic space could be observed 4 days after injection.     

The receptor and neuron distribution in the cochlea of the bat, Taphozous kachhensis

Cochlear receptors and spiral ganglion neurons in the inner ear of the bat, Taphozous kachhensis of the family Emballonuridae, were quantitatively analyzed on the basis of surface specimen technique, histological sections, and three-dimensional computer reconstruction. The length of the basilar membrane was 12.1 (+/- 0.4) mm. There were about 23,000 neurons in the spiral ganglion and about 1430 inner hair cells and 4635 outer hair cells in the organ of Corti. Cochlear neurons had two density maxima, one basal and one apical. Inner hair cells had basically the same distribution but less pronounced. Outer hair cells had only one density maximum in the apical region corresponding to the respective maxima of inner hair cells and cochlear neurons. Basal density maxima of inner hair cells and neurons are most probably involved in the analysis of echolocation signals, and presumably correlated with maximum sensitivity of the auditory system. A comparison with the bat, Nyctalus noctula of the family Vespertilionidae revealed a conspicuous similarity in some quantitative aspects of the cochlear structure. These particular similarities may be looked upon as convergent adaptations given by similar flight, hunting, and echolocation strategies in both bat species.     

Measurement of low-frequency receptor potentials in inner hair cells: a theoretical analysis

The measurement of the low-frequency responses of inner hair cells is complicated by the fact that extracellular cochlear microphonic may be larger than the intracellular inner hair cell potential. A subtraction technique is proposed in which the extracellular and intracellular potentials are recorded sequentially and then subtracted to give an estimate of the true membrane potential. A theoretical analysis is presented which predicts that this calculated inner hair cell membrane potential is a more accurate measure of inner hair cell receptor current than the membrane potential measured with respect to a remote indifferent electrode.     

耳蜗图的绘制及应用进展

位于耳蜗基底膜上的Corti器是接受声振动刺激的初级感受器，作为Corti器中的感觉细胞，毛细胞的损害将引起不同程度及频率范围的听力损失。对耳蜗毛细胞在基底膜不同部位的丢失情况进行定量分析是内耳组织病理学研究中最常用的手段，将全耳蜗毛细胞损害的定量定位分析结果与听功能相结合将有益于对疾病机制的探索和对治疗方法的评价。目前对全耳蜗毛细胞定位定量分析常采用耳蜗图来表示，但在听力学研究中耳蜗图的绘制方法并不完全相同，尚缺乏统一的绘制标准及原则，使得各个实验结果之间缺乏可比性。为更精准地对全耳蜗毛细胞的损伤情况进行定位定量分析，需要统一耳蜗图的绘制方法及原则。本文就目前文献报导中常用的耳蜗图绘制方法及绘制原则进行了综述。     

Strategies to regenerate hair cells: identification of progenitors and critical genes

Deafness commonly results from a lesion of the sensory cells and/or of the neurons of the auditory part of the inner ear. There are currently no treatments designed to halt or reverse the progression of hearing loss. A key goal in developing therapy for sensorineural deafness is the identification of strategies to replace lost hair cells. In amphibians and birds, a spontaneous post-injury regeneration of all inner ear sensory hair cells occurs. In contrast, in the mammalian cochlea, hair cells are only produced during embryogenesis. Many studies have been carried out in order to demonstrate the persistence of endogenous progenitors. The present review is first focused on the occurrence of spontaneous supernumerary hair cells and on nestin positive precursors found in the organ of Corti. A second approach to regenerating hair cells would be to find genes essential for their differentiation. This review will also focus on critical genes for embryonic hair cell formation such as the cell cycle related proteins, the Atoh1 gene and the Notch signaling pathway. Understanding mechanisms that underlie hair cell production is an essential prerequisite to defining therapeutic strategies to regenerate hair cells in the mature inner ear.     

Tiron对抗外源性过氧化氢对离体培养小鼠耳蜗Corti器损伤研究

目的：探讨Tiron能否有效地对抗过氧化氢（H2O2）对离体培养小鼠Corti器内、外毛细胞的损伤。方法：采用新生小鼠Corti器体外培养技术，建立外源性H2O2损伤内外毛细胞模型，给予不同浓度的Tiron，观察对H2O2损伤的保护作用。结果：H2O2浓度大于1.0mmol/L时，基底膜顶回、中回和底回毛细胞损伤程度不同，底回较重，预回和中回损伤程度在统计学上无差别。在低于0.5mmol/L时毛细胞损伤与位置无关。在培养液中加入Tiron(10mmol/L)可以明显减轻H2O2对毛细胞的损伤程度。当H2O2浓度在0.01-1.0mmol/L之间时，Tiron几乎可以完全抑制毛细胞的损伤和缺失。结论：Tiron对于小鼠离体培养的Corti器具有明显的保护作用，其对抗H2O2损伤的机制可能通过清除超氧阴离子O2^-和螯合铁离子共同发挥作用。     

Cochlear cultures as a model system for studying aminoglycoside induced ototoxicity

Light and electron microscopy have been used to evaluate the effects of treating mouse cochlear cultures with the ototoxic aminoglycoside antibiotic neomycin sulphate at concentrations of 0.2 mM and greater for periods of up to 1 hour. Neomycin rapidly induces the formation of numerous, membrane filled blisters on the apical surfaces of the sensory hair cells. Such morphological damage is restricted to the hair cells, and is not observed on the surfaces of supporting cells within the organ of Corti. Hair cells in apical-coil cultures are less sensitive than those in basal-coil cultures, and, at any given point along the cochlea, outer hair cells appear to be more extensively damaged by neomycin than inner hair cells. These morphological effects of neomycin are considerably more severe when the drug is applied in calcium/magnesium free saline, and can be blocked by elevating the saline concentration of either calcium or magnesium. The effects can also be blocked by lowering the temperature to 4 degrees C, but not by either K+ depolarization or the lectin Concanavalin A. The potential value of this culture system as a model for studying aminoglycoside induced ototoxicity is discussed.     

Absence of Voltage-Dependent Compliance in High-Frequency Cochlear Outer Hair Cells

Cochlear outer hair cells are the key element in a mechanical amplification process that enhances auditory sensitivity and tuning in the mammalian inner ear. The electromotility of outer hair cells, that is, their ability to extend or contract at acoustic frequencies, is proposed to be the source of the mechanical amplification. For amplification to take place, some stiffness is required for outer hair cells to communicate force to the organ of Corti, the sensory epithelium of the inner ear. Modulation of this stiffness would be expected to have a significant effect on inner ear function. Outer hair cell compressive stiffness has recently been shown to be dependent on membrane potential, but this has only been demonstrated for cells originating in the apical, low-frequency segment of the cochlea, whereas cochlear amplification is arguably more important in the more basal high-frequency segment. The voltage-dependent compliance (the reciprocal of stiffness) of high-frequency outer hair cells was investigated by two methods in cells isolated from the basal turns of the guinea pig cochlea. In contrast to previous findings, no evidence was found for voltage-dependent changes in compliance. The results call into question the importance of outer hair cell voltage-dependent compliance as a component of cochlear amplification.     

Ultrahistochemical demonstration of glucose-6-phosphatase in hair cells of the guinea pig organ of corti (author's transl)]

This study is concerned with the ultrahistochemical demonstration of glucose-6-phosphatase activity and its localization in inner and outer hair cells of the guinea pig organ of Corti. The enzyme activity has been demonstrated by a method according to Hugon et al. (1970). In hair cells of the organ of Corti a characteristic distribution pattern of reaction products has been registered. Subsurface cisterns and the Hensen's bodies of outer hair cells were heavily loaded with reaction products. In addition, the endoplasmic reticulum and the nuclear membrane as well as postsynaptic cisterns were rich in precipitates. With regard to their morphologic pecularities the inner hair cells show a more homogeneous distribution of enzyme activity. The findings corroborate the former assumption of a genetic relationship of either subsurface cisterns and Hensen's body to the endoplasmic reticulum of outer hair cells. Furthermore, the high glucose-6-phosphatase activity of both subsurface cisterns and Hensen's bodies are considered indicative of their participitation in the energy metabolism of outer hair cells. Referring to biochemical studies of Thalmann and associates (1973), the narrow spatial relationship of glucose-6-phosphatase positive ER membranes to mitochondria presumably represents a morphologic correlation of aerobic and anaerobic metabolic pathways in the guinea pig organ of Corti.     

Tubulin expression in the developing and adult gerbil organ of Corti

In the late stages of inner ear development, the relatively undifferentiated cells of Kollicker's organ are transformed into the elaborately specialized cell types of the organ of Corti. Microtubules are prominent features of adult cells in the organ of Corti, particularly supporting cells. To test the possible role of microtubules in organ of Corti development, the microtubule organization in the organ of Corti has been examined using indirect immunofluorescence to beta-tubulin in the developing gerbil cochlea. Tubulin first appears at post-natal day 0 (P0) as filamentous asters in inner hair cells and by P2, asters are also seen in outer hair cells. Tubulin appears at P3 in inner pillar cells in a tooth crown-like figure. By P6, tubulin expression is also evident in outer pillar cells and by P9, it is seen in Deiters cells. Elaboration of microtubules in pillar cells was observed to proceed from the reticular lamina towards the basilar membrane. The pattern of tubulin expression in the apical organ of Corti lags the base by about 3 days until P6, but by P9, apical and basal organ of Corti appear substantially the same.     

Actin-binding and microtubule-associated proteins in the organ of Corti.

Actin-binding and microtubule-associated proteins regulate microfilament and microtubule number, length, organization and location in cells. In freeze-dried preparations of the guinea pig cochlea, both actin and tubulin are found in the sensory and supporting cells of the organ of Corti. Fodrin (brain spectrin) co-localized with actin in the cuticular plates of both inner and outer hair cells and along the lateral wall of the outer hair cells. Alpha-actinin co-localized with actin in the cuticular plates of the hair cells and in the head and foot plates of the supporting cells. It was also found in the junctional regions between hair cells and supporting cells. Profilin co-localized with actin in the cuticular plates of the sensory hair cells. Myosin was detected only in the cuticular plates of the outer hair cells and in the supporting cells in the region facing endolymph. Gelsolin was found in the region of the nerve fibers. Tubulin is found in microtubules in all cells of the organ of Corti. In supporting cells, microtubules are bundled together with actin microfilaments and tropomyosin, as well as being present as individual microtubules arranged in networks. An intensely stained network of microtubules is found in both outer and inner sensory hair cells. The microtubules in the outer hair cells appear to course throughout the entire length of the cells, and based on their staining with antibodies to the tyrosinated form of tubulin they appear to be more dynamic structures than the microtubules in the supporting cells. The microtubule-associated protein MAP-2 is present only in outer hair cells within the organ of Corti and co-localizes with tubulin in these cells. No other MAPs (1,3,4,5) are present. Tau is found in the nerve fibers below both inner and outer hair cells and in the osseous spiral lamina. It is clear that the actin-binding and microtubule-associated proteins present in the cochlea co-localize with actin and tubulin and that they modulate microfilament and microtubule structure and function in a manner similar to that seen in other cell types. The location of some of these proteins in outer hair cells suggests a role for microfilaments and microtubules in outer hair cell motility.     

Canalicular reticulum in vestibular hair cells

A membrane limited system referred to as canalicular reticulum (CR) has been demonstrated in the apical cytosol of the cochlea's inner and outer hair cells. Similarities between cochlear and vestibular hair cells prompted investigation of the presence of CR in hair cells of the gerbil vestibular labyrinth. A method of fixation with glutaraldehyde followed by an osmium-ferrocyanide mixture demonstrated abundant CR in the apex of both type I and type II hair cells. The CR was closely associated with numerous Golgi zones in the apex of the vestibular hair cells, indicating its genesis from Golgi cisternae. Also preserved in upper cytosol were discrete complexes of mitochondria with granular reticulum. These complexes offered a possible site for generating the membrane in Golgi zones and CR. Single and parallel cisternae of granular reticulum were observed in the basal half of the hair cells together with numerous synaptic-like vesicles. These cisternae with their terminal blebbing and accompanying canaliculi were interpreted as novel structures mediating synaptic vesicle genesis in vestibular hair cells in a manner comparable to that postulated for cochlear inner hair cells.     

Comparative anatomy of the cochlea and auditory nerve in mammals

The numbers and structure of hair cells; afferent, efferent, and reciprocal synapses as seen at the base of hair cells; innervation patterns of first order cochlear neurons; and number and morphology of spiral ganglion cells will be discussed and compared in the guinea pig, rat, cat, monkey and man. Despite many similarities both in the organ of Corti and the spiral ganglion in these species, there are a number of differences which may have important physiologic implications. In the organ of Corti, the major differences among species are the length and width of the basilar membrane, the number of inner and outer hair cells, and the length of hairs on both inner and outer hair cells. Significant differences in the innervation pattern of the inner hair cell among these species include the number of afferent nerve terminals per inner hair cell, the degree of branching of afferent fibers, and the number of synapses per afferent nerve terminal. Among outer hair cells, the number of afferent nerve terminals per outer hair cell, presence or absence of a pre-synaptic body, presence or absence of reciprocal synapses, the number of efferent terminals per outer hair cell, and the presence of dendodendritic synapses in outer spiral bundles may be differences important physiologically. In the spiral ganglion, there are significant differences in the number of spiral ganglion cells, the number of cochlear nerve fibers, the percentage of spiral ganglion cells which are myelinated, and the presence of synapses on spiral ganglion cells.