Hair cell loss in the aged guinea pig cochlea

The various effects of ageing on the auditory system, collectively termed presbycusis, are being studied across a wide range of animal species, including humans. One contributing factor to presbycusis is thought to be losses of the sensory hair cells in the cochlea. In this study, hair cell counts were obtained from cochleas of pigmented guinea pigs (Cavia porcellus) at ages ranging from 11 days to 4 years 7 months, using scanning electron microscopy to visualize the organ of Corti. Representative samples of the basal, middle and apical turn of the cochlea were photographed for analysis. Hair cell loss was observed, even in young animals. However, the loss was greater in the aged animals, but was not distributed evenly throughout the length of the cochlea. No significant loss of hair cells was seen in the basal (high frequency) or middle turn of the cochlea of the aged animals. In the apical (low frequency) turn, there was a significant loss of hair cells in all rows of outer hair cells (up to around 20%), and was most severe in the third row. There was no loss of apical inner hair cells in the aged animals.     

Hair cell damage produced by acoustic trauma in the chick cochlea

Examination of pure-tone acoustic damage in the chick basilar papilla revealed that the location and extent of hair cell damage was a function of both the stimulus intensity and the age at which the chicks were exposed. Scanning electron microscopic evaluation of noise-exposed cochleae at post-hatching days 1, 10 and 30 permitted the identification of discrete regions of damage, including hair cells with stereocilia injuries as well as those lost from the epithelium. The hair cell damage was tonotopically distributed along the cochlea according to frequency. However, for each exposure frequency two distinct sites of damage were often produced, and their locations were correlated with stimulus intensity. At low intensities, a longitudinal strip of hair cell damage ran along the superior edge of the basilar papilla. As exposure intensity increased, a second damage site developed along the inferior edge of the basilar papilla, distal to the longitudinal strip. This second type of damage initially took the form of a series of laterally-oriented wedges, but at higher intensities, the wedges coalesced to form a large crescent-shaped patch of damage. The location of the damage sites for each frequency did not shift with age. However, there were differences in the extent and position of the damage which could be correlated with stimulus intensity and with changes in middle ear admittance during development [(1983) Development of Auditory and Vestibular Systems, pp. 3-25. Editor: R. Romand. Academic Press, New York]. These results suggest that developmental changes in the location and extent of hair cell damage depend on the effective stimulus intensity reaching the cochlea, rather than on alterations in the frequency coding of the hair cells.     

Hair cell and supporting cell response to acoustic trauma in the chick cochlea

Damage to the chick cochlea in response to progressively increased periods of noise exposure was studied with scanning electron microscopy. Ten day-old chick hatchlings were exposed to a 1500 Hz pure tone at 120 dB SPL for 4, 8, 12, 24, and 48 h. Measurements of hair cell and supporting cell surface areas within a defined region of the cochlea showed that the average hair cell surface area decreased over the first 12 h of exposure. Between 12 h and 48 h there was no significant change in hair cell surface area. Supporting cells showed a corresponding increase in surface area over the same period. Noise damage first appeared after 4 h of exposure as a localized expansion of supporting cell surfaces near the inferior edge of the basilar papilla (BP). Between 8 and 12 h of exposure the supporting cell surface area increased dramatically and was visible throughout the noise damaged region. Hair cell expulsion was first seen after 12 h. Exposure to noise for 24-48 h resulted in further expansion of supporting cells, extensive expulsion of hair cells from the BP, and the appearance of a strip of noise damage along the superior, edge of the BP.     

Hair cell regeneration in the chicken cochlea following aminoglycoside toxicity.

Hair cell loss in the avian cochlea partially recovers following both acoustic trauma and aminoglycoside intoxication. DNA labeling with tritiated thymidine has shown that the restoration of cell number following acoustic trauma results from the production of new hair cells by mitotic division. The purpose of the present study was to determine if mitosis also contributes to the recovery of hair cell number which occurs following aminoglycoside intoxication. Chickens received daily injections of either gentamicin sulfate or distilled water for 10 consecutive days. During the latter 7 days of this period, all birds were also injected with [3H]thymidine. Following postinjection survival periods of 3 or 6 days, one papilla from each bird was processed for autoradiography and the other for scanning electron microscopy (SEM). Incorporation of [3H]thymidine was seen over hair cells and support cells in experimental papillae in regions of hair cell loss. No labeling was seen outside of damaged regions or in the papillae of control birds. SEM showed that damaged regions in experimental birds contained cells similar in appearance to developing auditory hair cells in avian embryos. These results show that the restoration of hair cell number following aminoglycoside toxicity results from the production of new cells by mitosis.     

Expression of glycoconjugates in the human fetal cochlea.

The distribution of glycoconjugates in the human fetal cochlea was analyzed using six biotinylated lectins: wheat germ agglutinin (WGA), abrus precatorius agglutinin (APA), ulex europaeus agglutinin I (UEA-I), ricinus communis agglutinin 120 (RCA120), helix pomatia agglutinin (HPA), concanavalin A (ConA). The tectorial membrane (TM) in the 11- and 15-week-old human fetuses was labelled with WGA, APA, RCA120 and ConA, but not with UEA-I and HPA. In the 19-week-old fetuses, the reaction of the TM decreased. In the 11-, 15- and 19-week-old fetuses, the surface of the greater and the lesser epithelial ridges were respectively labelled with WGA, APA and RCA120. Reissner's membrane was labelled with WGA, ConA, APA, RCA120 and HPA. WGA, RCA120 and APA strongly stained the stria vascularis, especially in the 15-week-old fetuses. HPA did not stain the 11-week-old fetal cochleas at all, while it stained the apical surface of the hair cells, Reissner's membrane, the cells within the stria vascularis and spiral osseous lamina in the 15-week-old fetuses. In the 19-week-old fetuses, the fluorescent reaction with HPA became decreased and the apical surface of the hair cells was not labelled with HPA at all. This result suggests that HPA reactive glycoconjugates may be related to the molecule responsible for stereociliary adhesion only during development.     

The efferent innervation of the avian cochlea

The efferent auditory innervation of the budgerigar (Melopsittacus undulatus) was investigated by means of acetylcholinesterase staining. The course of the efferent fibres is described: the single terminal fibre regularly branches out to innervate several hair cells. Three types of efferent, axosomatic synapses could be distinguished: small outer hair cells bear large cup-like efferent synapses, intermediate hair cells with cone-shaped synapses, and tall inner hair cells with small, knob-like synapses. In the basal part of the cochlea the small hair cells with heavy efferent endings predominate, while in the apex only tall inner hair cells with small efferent terminals exist. There are more intermediate hair cells in the apical area than in the basal area.     

Hair cell damage and recovery following chronic application of kanamycin in the chick cochlea

Three-day old chicks were given kanamycin at a dose of 200 mg/kg/day for 10 days and their cochleae were processed for scanning electron microscopy at 1, 3, 7 and 14 days following the last injection. Both hair cells and supporting cells were damaged by kanamycin in the basal 35% of the basilar papilla. By 14 days post-treatment, however, most of the damaged region had been replaced with regenerating hair cells and supporting cells. The base-to-apex gradient of morphological development along the cochlea was observed in the process of regeneration. Kinocilium and microvilli were observed on the apical surfaces of the regenerating hair cells.     

新生鼠耳蜗毛细胞再生

各种疾病、噪声、耳毒药等所致感音神经性聋,其实质均在于内耳毛细胞的变性和坏死。以往研究表明在生后正常哺乳类动物内耳耳蜗内、外毛细胞是不能再生的。但是,科学家经过听觉系统基因调控研究,发现了毛细胞分化、再生的特有的调控基因,文章已发表在“科学”和“自然(神经科学)”、“发育”、“神经科学”、“发育机制”等杂志上,引起国际上的关注。现将其结果和意义短评如下。     

OCP2 immunoreactivity in the human fetal cochlea at weeks 11, 17, 20, and 28, and the human adult cochlea

The two most abundant proteins of the organ of Corti, OCP1 and OCP2, are acidic, cytosolic, low molecular weight proteins diffusely distributed within the cytoplasm of supporting cells. A recent study by Henzl et al. (2001) found first, that these two proteins co-localize with connexin 26 along the epithelial gap junction system and second, that OCP2 could participate with OCP1 in an organ of Corti-specific SCF complex (Skp1, cul1in, and Fbp), a ubiquitin ligase complex. Previous study has also implicated OCP2 in the recycling and regulation of intracellular K(+) efflux as well as pH homeostatic mechanisms. In the present study, we document the emergence and distribution features of OCP2 through various stages (weeks 11-28) of gestation in human fetal cochleae. Four fetal cochleae, the cochleae of a normal hearing human adult and a mature rat for positive control were fixed in 4% formalin within 2 h post mortem. Immunohistochemical studies were performed using a rabbit polyclonal antibody raised against a synthetic peptide corresponding to amino acids 3-16. Specimens were mounted in paraffin sections. Results show that OCP2 immunoreactivity is evident at a prenatal age of 11 weeks, peaks in expression at the onset of cochlear function at 20 weeks and achieves adult-like patterns of distribution just prior to histological maturation at 28 weeks. Though this protein could be associated with the development, maturation, and electrochemical maintenance of the cochlear gap junction system, the nature of this protein's function in the developing and mature human cochlea remains unclear.     

Hair cell regeneration after local application of gentamicin at the round window of the cochlea in the pigeon

Hair cells in the basilar papilla of birds have the capacity to regenerate after injury. Methods commonly used to induce cochlear damage are systemic application of ototoxic substances such as aminoglycoside antibiotics or loud sound. Both methods have disadvantages. The systemic application of antibiotics results in damage restricted to the basal 50% of the papilla and has severe side effects on the kidneys. Loud sound damages only small parts of the papilla and is restricted to the short hair cells. The present study was undertaken to determine the effect of local aminoglycoside application on the physiology and morphology of the avian basilar papilla. Collagen sponges loaded with gentamicin were placed at the round window of the cochlea in adult pigeons. The time course of hearing thresholds was determined from auditory brain stem responses elicited with pure tone bursts within a frequency range of 0.35–5.565 kHz. The condition of the basilar papilla was determined from scanning electron micrographs. Five days after application of the collagen sponges loaded with gentamicin severe hearing loss, except for the lowest frequency tested, was observed. Only at the apical 20% of the basilar papilla hair cells were left intact, all other hair cells were missing or damaged. At all frequencies there was little functional recovery until day 13 after implantation. At frequencies above 1 kHz functional recovery occurred at a rate of up to 4 dB/day until day 21, beyond that day recovery continued at a rate below 1 dB/day until day 48 at the 5.6 kHz. Below 1 kHz recovery occurred up to day 22, the recovery rate was below 2 dB/day. A residual hearing loss of about 15–25 dB remained at all frequencies, except for the lowest frequency tested. At day 20 new hair cells were seen on the basilar papilla. At day 48 the hair cells appeared to have recovered fully, except for the orientation of the hair cell bundles. The advantage of the local application of the aminoglycoside drug over systemic application is that it damages almost all hair cells in the basilar papilla and it has no toxic side effects. The damage is more extensive than with systemic application.     

Hair cell regeneration in the avian cochlea: if it works in birds, why not in man?

Hearing loss caused by cochlear hair cell loss is the most common process afflicting the hearing impaired. Recent studies in the avian cochlea following ototoxic drug and noise damage have demonstrated a remarkable capacity for anatomical and functional recovery. Hair cell regeneration has been shown to play a major role in this recovery process. Future studies may one day make hair cell regeneration or transplantation possible in man.     

Glyoxylic acid in the study of autonomic innervation in the gerbil cochlea.

The autonomic innervation of the inner ear has been investigated earlier, yet questions concerning the origin, function, extent, and distribution of sympathetic nerves in the cochlea still remain unanswered. This study investigates the extent and distribution of adrenergic nerves in the cochlea. Our procedure combines the glyoxylic acid method of catecholamine fluorophore identification with rapid dissection, decalcification, and whole-mount slide preparation techniques to topographically trace the adrenergic innervation of the cochlear infrastructure. We have demonstrated that perivascular adrenergic innervation extends beyond the immediate branches of the modiolar artery and reaches into radiating arterioles. These findings also suggest the possibility of segmental regulation of cochlear blood flow.     

Expression of carbonic anhydrase isoenzyme-like immunoreactivity in the limbus spiralis of the human fetal cochlea.

The distribution of carbonic anhydrase isoenzymes (CA I, II, III, V) was analyzed the 14-, 15- and 16-week-old human fetal cochleae using immunohistochemical methods. The interdental cells in the limbus spiralis were strongly labelled with antibodies against CA I and III and the spiral ligament was also positive for CA I and III. The stria vascularis and organ of Corti were however negative for CA I, II, III and V. These results suggest that the interdental cells in the human fetus may be related to fluid and ion transport of endolymph especially in the early stages of development.     

Synaptophysin immunohistochemistry in the human cochlea

Light microscopy and immunohistochemical analyses of a freshly prepared human cochlea, removed at meningioma skull base surgery, were performed with particular emphasis on synaptophysin (SY) reactivity. Synaptophysin, a 38-kDa glycoprotein, is one of the most abundant integral membrane proteins of small presynaptic vesicles and is a useful marker for sites of synaptic transmission of the efferent olivocochlear system in the cochlea. Following fixation and decalcification, cryosections of 30 microm were prepared. To introduce immunostaining, free-floating sections were exposed to monoclonal SY antibody. Positive SY immunostaining was solely restricted to the neural and sensory structures and did not include supporting cells of the organ of Corti. Dense reaction products were noted around the hair cells, especially at the basal portion of the inner and outer hair cells and their neural poles, as well as around the inner spiral bundle, tunnel spiral bundle, outer spiral bundle and upper tunnel crossing fibers. The majority of spiral ganglion cells stained positively. An intermingling network of thin unmyelinated nerve fibers stained densely, especially at the basal portions of the cochlea. The spiral limbus, inner and outer sulcus cells, basilar membrane, myelinated nerve fibers, spiral ligament and the stria vascularis were unstained. Human cochlea obtained during surgery offers excellent conditions for immunohistochemical analysis. In the basal cochlea in the organ of Corti, outer hair cell area, there may be alterations due to noise trauma from the drilling procedure.     

大上皮嵴与哺乳类耳蜗毛细胞的分化和再生

位于感觉上皮外的大上皮嵴细胞经诱导能产生大量异位毛细胞,这是毛细胞再生的一条新的途径,将来也许有助于感音神经性聋的治疗。本文概述了大上皮嵴中异位毛细胞的产生及其分子水平机制,最后介绍目前存在的问题及将来的发展趋势。     

Comparison of the vascular innervation of the rat cochlea and vestibular system

In order to gain a better understanding of the neuronal and local control of inner ear blood flow, the vascular innervation to the rat cochlea and vestibular system was examined. Specimens were removed in toto beginning at the basilar artery extending to the anterior inferior cerebellar artery, labyrinthine artery, common cochlear artery, modiolar artery and anterior vestibular artery. When possible the vessels were dissected in continuity through the cribrose area. The vestibular endorgans were also removed. Specimens were examined using immunohistochemical techniques for the presence of vasoactive intestinal peptide, neuronal nitric oxide synthase, neuropeptide-Y, substance P and calcitonin gene related peptide. Results show that the vasculature to the cochlea and vestibular portion of the inner ear receive similar types of nonadrenergic innervation, that within the vestibular endorgans, only CGRP and SP were found in the neuroepithelium or in association with vessels, and that within the vestibular system, the majority of the vascular innervation appears to stop at or near the cribrose area. In the cochlea however, it extends to include the radiating arterioles. These findings suggest that cochlear blood flow is under finer control and that neuronally induced changes in blood flow may have a more global effect in the vestibular periphery.