Hair cell recovery in the vestibular sensory epithelia of mature guinea pigs

The progression of recovery of the vestibular sensory epithelia of guinea pigs after gentamicin-induced hair cell injury was assessed quantitatively and qualitatively. Evaluations were made of the number of cells bearing hair bundles by using scanning electron microscopy (SEM) and of identifiable hair cells in thin sections. Both assessment procedures showed that an initial loss of hair cells in utricular maculae is followed by significant recovery in the number of hair cells present. SEM also showed recovery in saccules comparable to that in utricles. During the recovery, progressive maturation of hair bundles, which exhibited features similar to those seen during normal ontogenetic development of hair cells, could be identified. The pattern and extent of hair cell loss and subsequent reappearance revealed by SEM corresponded with that derived from analysis of thin sections. This suggests that repair of nonlethally damaged hair cells is unlikely but, rather, that new hair cells are produced. An apparent decrease in supporting cell numbers was observed coincident with the increase in hair cell numbers. This complements previous morphological observations, which have suggested new hair cells arise from direct, nonmitotic transdifferentiation of supporting cells. The quantitative analyses indicate that more than half of the hair cells that are lost are replaced, but the recovery process does not result in complete restoration of the epithelium. Eight months after the end of drug treatment, the number of hair cells present was still significantly less than normal, and several other abnormalities persisted. There was also no evidence of any hair cell recovery in the organ of Corti. Thus, there appear to be limitations on the capacity for spontaneous replacement of lost hair cells in the mammalian inner ear.     

Evaluation and application of ribotyping for epidemiological studies of Actinobacillus pleuropneumoniae in Denmark

Hair cell regeneration is well documented in the inner ear sensory epithelia of lower vertebrates and birds and may occur in the vestibular organs of mammals. By contrast, hair cell loss in the mature mammalian cochlea is considered irreversible. However, recent reports have suggested that an attempt at hair cell regeneration could occur in vivo in aminoglycoside-lesioned cochleas from neonatal rats. After amikacin treatment, atypical cells with apical specialization reminiscent of early differentiating stereocilia are transiently present at the apex of the intoxicated cochleas but fail to differentiate as hair cells in later stages. In the present study, we used electronic microscopy, histochemistry, and confocal microscopy to investigate the cellular rearrangements in the amikacin-lesioned organ of Corti of rat pups. In addition, we used 5-bromo-2'-deoxyuridine immunocytochemistry to determine whether mitotic processes are involved in the formation of the atypical cells. The morphologic and molecular data suggest that atypical cells are not recovering hair cells, but share characteristics of immature hair cells and supporting cells. Proliferative cells were absent from the region occupied by atypical cells, suggesting that the latter did not arise through mitotic processes. Altogether, the present results support the hypothesis that atypical cells arise through direct transformation of some of the supporting cells that reorganize during hair cell degeneration.     

Hair bundle morphology on surviving hair cells of the chick basilar papilla exposed to intense sound

Exposure to intense sound produces a well-defined "patch" lesion on the chick basilar papilla in which 30-35% of the short hair cells are lost. The present study compares various aspects of sensory hair bundle morphology on surviving hair cells in the patch lesion with hair bundles from matched locations on nonexposed control papilla immediately after removal from the exposure and 12-days post exposure. The height and thickness of the hairs, the total number of hairs in the bundle, the width of the bundle, and the area and perimeter of the apical surface of the hair cell were quantified from scanning electron microscope photomicrographs. An attempt was also made to determine if there was a consistent microstructure to the pattern of hair cell loss within the lesion area. Similar observations in 12-day recovered ears are also presented. The results indicated that stereocilia height increased and width decreased on surviving hair cells in the exposed ear. The width of the hair bundle, the hair cell surface area, and perimeter also decreased. However, the number of hairs per cell remained unchanged, and there was no evidence of any consistent organization to the hair cell loss within the patch across a number of specimens. These observations indicated that the hair bundles on short hair cells underwent changes as a consequence of intense sound exposure. The results after 12 days of recovery were complicated by developmental changes on the papilla and incomplete maturation of the newly regenerated hair cells. It remains to be seen whether these changes were the result of cell sampling in the sound-damaged ear or were due to true structural alterations within the sensory hairs themselves.     

The cuticular plate of the hair cell in relation to morphological gradients of the chicken basilar papilla

The aim of the present study was to provide details on the diversity and morphological gradients in the anatomy of the cuticular plate of hair cells in the chicken basilar papilla. The structure of the cuticular plate, which is mainly made up of a network of actin filaments, may be related to differences in the mechanical demands on the anchorage of the stereovillar bundle. We describe the morphological gradients in the cuticular plates as seen in transverse section for four positions along the basilar papilla. Three different shapes of cuticular plate could be distinguished. In general, cuticular plates in neurally-lying hair cells have their main mass on the neural side of the cells; for abneural cells, the converse is true. The shape of the plates changes gradually across the papilla; symmetrical forms exist. The hair-cell bundle orientation (and thus the preferred direction of stimulation of the bundle), as measured using scanning EM preparations, does not correlate with the shape of the plate in transverse section. The present data confirm the notion developed from other studies that (1) there are no distinct populations of hair cells, (2) there are no linear or monotonic morphological gradients, and (3) the gradients on the papilla are species- and position-specific.     

Neurofilament proteins form an annular superstructure in guinea-pig type I vestibular hair cells

Neurofilaments, the neuron-specific intermediate filaments, are composed of three immunochemically distinct subunits: NF-L, NF-M and NF-H that can be either phosphorylated or unphosphorylated. In mammals, the distribution of these subunits has been described in vestibular ganglion neurons, but there are no reports on the presence of neurofilaments in vestibular hair cells. We investigated, by immunocytochemistry, neurofilaments in vestibular hair cells from rat and guinea-pig using antibodies against the three subunits and to dephosphorylated NF-H (clone SMI 32, recognizes also NF-M on immunoblots), on Vibratome sections of the vestibular end-organs and on isolated hair cells. Various immunostaining protocols were used, as appropriate for the method of observation: laser scanning confocal microscopy (immunofluorescence) and transmission electron microscopy (immunoperoxidase, pre-embedding technique). In rat and guinea-pig cristae and utricles, neurofilament immunoreactivity was observed in axons inside and below the sensory epithelia. In guinea-pig, in addition to this staining, intensely immunoreactive annular structures were found in the basal regions of hair cells. These rings were detected with anti-NF-L, -NF-M and -dephosphorylated NF-H/M antibodies, but not with anti-phosphorylation-independent NF-H. Ring-containing hair cells were present in all regions of the sensory epithelia but were more abundant in the peripheral areas. All levels of observation (Vibratome and thin sections, and isolated hair cells) showed that only the guinea-pig type I hair cells contained a neurofilament ring. High-resolution observations showed that the ring was located below the nucleus, often close to smooth endoplasmic reticulum and the cell membrane.     

Localization of Merkel cells at hairless and hairy human skin sites using keratin 18

Merkel cells are neurosecretory cells of the skin with epithelial features such as desmosomes and expression of keratins 8, 18, 19, and 20. Merkel cells are scarcely distributed in adult human skin. Although they are present in hair follicles, their density is higher at hairless anatomic sites such as palms and soles. These cells are often innervated by sensory nerve fibers and are thought to be specialized mechanosensory skin receptor cells. However, their precise origin and function are not clearly established. The aim of this study was to localize Merkel cells in human hairless and hairy skin by immunohistochemistry with antibodies Ks18.174 and Ks19.1 directed against keratins 18 and 19, respectively. In glabrous skin of palm and sole, Merkel cells have been localized at the bottom of the rete ridges, in the epidermal basal layer. To study Merkel cell distribution at hairy anatomic sites, we have chosen breast skin, a tissue containing small hair follicles typical of those covering most of the body's surface. Merkel cells were present in the interfollicular epidermis. In hair follicles, they have been identified in the isthmus region.     

Mechanism of mitochondrial dysfunction and cytotoxicity induced by tropolones in isolated rat hepatocytes

The hair cell orientation patterns present on the saccules of fishes may be important for encoding the direction of a sound source. This study was conducted to determine whether primary afferent projections to the medulla are organized by the best directions for the hair cells they innervate. The toadfish saccule has hair cells oriented primarily in the vertical plane: both the rostral and the caudal saccule have hair cell orientations sweeping from 0 degrees to 45 degrees, and the middle saccule has hair cells oriented at 90 degrees. Fluorescent dextran amines were used singly and in combination to label regions of the saccular nerve innervating rostral, middle, and caudal saccule. The projections of those afferents were examined in detail in the anterior and descending octaval nuclei, which are auditory nuclei in this species. There was no evidence of topographic projections based on location along the length of the saccule or based on hair cell orientation. There was some evidence that parallel inputs are present from each region of the saccule examined, which may be based on the 180 degrees opposition of hair cells found throughout the saccule; however, afferents from the rostral, middle, and caudal saccule appear to have overlapping projections to the anterior and descending octaval nuclei. These data suggest that in toadfish, calculations of the direction of the sound source may begin in either of these primary auditory nuclei by comparing afferent input from along the saccule.     

Differential glycosylation of auditory and vestibular hair bundle proteins revealed by peanut agglutinin

Up to four morphologically distinct types of cross-link are found between the stereocilia in the hair bundles of avian hair cells. These links are involved in mechanotransduction, force transmission across the bundle, and maintenance of hair bundle structure. They appear to be specialisations of the cell coat, but very little is known about their molecular composition. Chick inner ear tissues were therefore screened with a number of different lectins to find markers for specialisations of the hair bundle surface. One lectin, peanut agglutinin (PNA), which recognises the dissacharide Gal beta 1-3GalNAc, was found to be a fairly selective marker for vestibular hair bundles, but it does not stain the stereocilia of auditory hair cells. The staining patterns observed with PNA in the vestibular system closely resemble those seen with a monoclonal antibody (mab) directed against a 275 kD component of the hair cell's apical surface known as the hair-cell antigen (HCA). However, unlike PNA, the mab recognises both vestibular and auditory hair cells. A detailed comparison of the fluorescence staining patterns observed with PNA and the anti-HCA mab indicates that binding sites for both ligands spatially codistribute on the surface of vestibular hair cells. The lectin and the anti-HCA mab binding sites are both sensitive to trypsin treatment, and, with sections of the vestibular system, PNA pretreatment blocks subsequent anti-HCA mab staining. Immunoelectron microscopy of vestibular hair bundles shows that PNA and the anti-HCA mab both label a type of cross-link known as the shaft connector. This link type is present on both auditory and vestibular hair bundles but reacts with PNA only in the vestibular system. The lectin jacalin, which has greater specificity for Gal beta 1-3GalNAc than does PNA, also only labels vestibular and not auditory hair bundles. Although terminal sialic acid residues can block both PNA and jacalin binding, neuraminidase treatment does not unmask cryptic binding sites for these lectins on auditory hair cells but does reveal PNA and jacalin staining at a number of other locations in the inner ear. The results obtained with the lectins PNA and jacalin indicate that either the HCA or other components of the shaft links are differentially glycosylated in the vestibular and auditory epithelia of the bird. The functional significance for such a difference in glycosylation remains to be determined, but auditory and vestibular hair cells operate over different frequency ranges, and variations in glycosylation might confer different micromechanical properties on the hair bundles in these two systems.     

Motility of isolated vestibular hair cells induced by potassium

Many studies of the outer hair cells in cochlea have demonstrated active motility. However, very few studies have been done on vestibular hair cells. This study was designed to demonstrate the motile responses of isolated vestibular hair cells of the chick, induced by potassium promoting contraction. Reversible cell shape changes were observed in 4 of 6 type I hair cells and 3 of 5 type II hair cells by applying the contraction solution. The cell shape changes were revealed mainly in the cuticular plate and infracuticular region. It was suggested that contraction in the cuticular plate of the isolated hair cells might be converted into tension which increases the stiffness of the sensory hairs and restricts their motions, based on the results of the present study, and the structure of contractile proteins and hair behaviors reported by previous investigators.     

Myosin Ibeta is located at tip link anchors in vestibular hair bundles

Recent studies have suggested that myosin Ibeta mediates the adaptation of mechanoelectrical transduction in vestibular hair cells. An important prediction of this hypothesis is that myosin Ibeta should be found in the side insertional plaque, an osmiophilic hair bundle structure that anchors tip links and is thought to house the adaptation motor. To determine whether myosin Ibeta was situated properly to perform adaptation, we used immunofluorescence and immunoelectron microscopy with the monoclonal antibody mT2 to examine the distribution of myosin Ibeta in hair bundles of the bullfrog utricle. Although utricular hair cells differ in their rates and extent of adaptation [Baird RA (1994) Comparative transduction mechanisms of hair cells in the bullfrog utriculus. II. Sensitivity and response dynamics to hair bundle displacement. J Neurophysiol 71:685-705.], myosin Ibeta was present in all hair bundles, regardless of adaptation kinetics. Confirming that, nevertheless, it was positioned properly to mediate adaptation, myosin Ibeta was found at significantly higher levels in the side insertional plaque. Myosin Ibeta was also present at elevated levels at the second tip link anchor of a hair bundle, the tip insertional plaque, found at the tip of a stereocilium. These data support myosin Ibeta as the adaptation motor and are consistent with the suggestion that the motor serves to restore tension applied to transduction channels to an optimal level, albeit with different kinetics in different cell types.     

Touch-plate and statolith formation in graviceptors of ephyrae which developed while weightless in space

Ultrastructural studies of the statocysts and touch-plates of graviceptors (rhopalia) of Aurelia ephyrae revealed that (1) touch-plate hair cells are present; and (2) cytoplasmic strands from the hair cell bases extend from the neurite plexus to touch similar strands from the lithocytes. This close association of hair cell neurites and statocysts may have important implications regarding the transmitting and processing of positional information with respect to the gravity vector. Graviceptors of ephyrae which developed while weightless in microgravity were compared with controls at the ultrastructural level. We found that hair cells of ephyrae which developed in microgravity had fewer lipid droplets in the large spaces near their bases as compared with 1 g controls. In the ephyrae from the first microgravity experiment, hair cells had more large apical vacuoles with filamentous content than were found in hair cells of ephyrae from the second experiment and controls. The neurite plexus and the network of cytoplasmic strands extending to the statocysts were not different in microgravity-developed ephyrae from controls. Behavioral differences in swimming and orienting in ephyrae in microgravity and controls (reported earlier) were not explained by morphological differences in the hair cells of the touch-plates or the statocysts, although functional differences apparently occurred.     

Fasciola hepatica development in the experimentally infected black rat Rattus rattus

Coat colour changes in polar animals are related to seasonal variation in photic inputs. The present work was performed to study the photoresponses of hair follicular melanocytes in human skin. The melanocytes, being photosensitive cells, can function as UV biosensors, since dendrites extend towards the source of UV light. Fifty-one skin biopsies from the margin of vitiligo were subjected to whole skin organ cultures. These were exposed to a pulse of UV light to study hair bulb melanocytes in vitiligo. It is observed that the melanocytes are seen within the anagen matrix. These melanocytes are poorly dendritic in control and dark-incubated cultures. On UV exposure, they become highly dendritic, the dendrites extending towards the hair shaft in 93.5%. They show prominent catechol oxidase and noradrenaline positivity, all features of UV responsiveness. The melanocytes within the hair follicle are not directly exposed to UV light. The melanocyte dendricity and the alignment of dendrites towards the shaft on UV exposure indicate that the columns of the cells in the hair shaft act as an efficient fibre-optic system, transmitting UV light. Morphologically, the keratinocytes in the hair shaft are arranged in compressed linear columns which resemble the coaxial bundles of commercial fibre-optic strands as is observed in plants. Keratinocytes in the inner and outer sheaths do not show this arrangement. Thus the hair follicle functions as a specialised UV receptor in the skin responding to nuances of photic inputs in human skin. This is reflected in coat colour changes in animals exposed to large variations in day-night cycles.     

Mechanotransduction of hair bundles arising from multicellular complexes in anemones

Sea anemones are among the simplest animals to use hair bundles to detect vibrations. Although we previously found anemone bundles to be morphologically similar to vertebrate hair bundles, only indirect evidence implicated anemone bundles in mechanotransduction. Here, we test mechanotransduction of these bundles using loose-patch current recording from apical membranes of cells at the base of deflected bundles. Step bundle deflection results in graded membrane currents that are inward in some cells (positive) and outward in other cells (negative). Positive responses range from 5 to 30 pA, abruptly saturate with stronger stimuli, and increase in duration with prolonged deflections. Negative responses range from 10 to 150 pA, show a logarithmic relation to stimulus strength, and attenuate with prolonged deflections. Additionally, responses are reversibly inhibited by streptomycin. We present a model for anemone bundle mechanotransduction modified from the gating spring model for vertebrate mechanotransduction. Because anemone bundles comprise stereocilia arising from a multicellular complex, we propose that supporting cells on opposite sides of a bundle function as oppositely polarized hair cells. Thus, deflection induces ion channels to open in cells on one side of the complex, while allowing channels to close in cells on the opposite side of the complex.     

Ultrastructural, physiological, and molecular defects in the inner ear of a gene-knockout mouse model for autosomal Alport syndrome

It is well documented that damage to the chick cochlea caused by acoustic overstimulation or ototoxic drugs is reversible. Second-order auditory neurons in nucleus magnocellularis (NM) are sensitive to changes in input from the cochlea. However, few experiments studying changes in NM during cochlear hair cell loss and regeneration have been reported. Chicks were given a single systemic dose of gentamicin, which results in maximal hair cell loss in the base of the cochlea after 5 days. Many new hair cells are present by 9 days. These new hair cells are mature but not completely recovered in organization by 70 days. We counted neurons in Nissl-stained sections of the brainstem within specific tonotopic regions of NM, comparing absolute cell number between gentamicin- and saline-treated animals at both short and long survival times. Our data suggest that neuronal number in rostral NM parallels hair cell number in the base of the cochlea. That is, after a single dose of gentamicin, we see a loss of both cochlear hair cells and NM neurons early, followed by a recovery of both cochlear hair cells and NM neurons later. These results suggest that neurons, like cochlear hair cells, can recover following gentamicin-induced damage.     

Regulation of human dermal papilla cell production of insulin-like growth factor binding protein-3 by retinoic acid, glucocorticoids, and insulin-like growth factor-1

Insulin-like growth factor-1 (IGF1) has been reported to stimulate hair elongation and to facilitate maintenance of the hair follicle in anagen phase. However, little is known about IGF1 signaling in the hair follicle. In this study we investigate the effects of IGF1, glucocorticoids, and retinoids on dermal papilla (DP) cell production of insulin-like growth factor binding proteins (IGFBPs). IGFBPs comprise a family of IGF binding proteins that are produced and released by most cell types. They bind to IGFs to either enhance or inhibit IGF activity. In the present report we identify IGFBP-3 as being produced and released by cultured human dermal papilla (DP) cells. IGFBP-3 levels are increased fivefold by retinoic acid, eightfold by dexamethasone, and tenfold by IGF1. DP cells are known to produce IGF1, and so the observed stimulation of DP cell IGFBP-3 production by IGF1 is consistent with the idea that DP cells possess the IGF transmembrane receptor kinase and are autoregulated by IGFs. The level of another IGFBP, tentatively identified as IGFBP-2, is, in contrast, not regulated by these agents. IGFBP-3 has been shown to inhibit the activity of IGFs in a variety of systems. Our results are consistent with a model in which retinoids and glucocorticoids inhibit IGF action on DP cells and surrounding matrix cells by stimulating increased DP cell production of IGFBP-3. The IGFBP-3, in turn, forms a complex with free IGF1 to reduce the concentration of IGF1 available to stimulate hair elongation and maintenance of anagen phase.     

Sound damage and gentamicin treatment produce different patterns of damage to the efferent innervation of the chick cochlea

Both sound exposure and gentamicin treatment cause damage to sensory hair cells in the peripheral chick auditory organ, the basilar papilla. This induces a regeneration response which replaces hair cells and restores auditory function. Since functional recovery requires the re-establishment of connections between regenerated hair cells and the central nervous system, we have investigated the effects of sound damage and gentamicin treatment on the neuronal elements within the cochlea. Whole-mount preparations of basilar papillae were labeled with phalloidin to label the actin cytoskeleton and antibodies to neurofilaments, choline acetyltransferase, and synapsin to label neurons; and examined by confocal laser scanning microscopy. When chicks are treated with gentamicin or exposed to acoustic overstimulation, the transverse nerve fibers show no changes from normal cochleae assayed in parallel. Efferent nerve terminals, however, disappear from areas depleted of hair cells following acoustic trauma. In contrast, efferent nerve endings are still present in the areas of hair cell loss following gentamicin treatment, although their morphological appearance is greatly altered. These differences in the response of efferent nerve terminals to sound exposure versus gentamicin treatment may account, at least in part, for the discrepancies reported in the time of recovery of auditory function.     

Hair cell formation in cultures of dissociated cells from the vestibular sensory epithelium of the bullfrog

HYPOTHESIS: Bullfrog vestibular hair cells are capable of regenerating in vitro. BACKGROUND: Recent studies have established that sensory organs in the inner ear of vertebrates continue to produce hair cells after birth. However, the mechanisms responsible for the regulation of this process are not well understood. The current study reports the development of a novel method for the culture of dispersed cells from the bullfrog inner ear. METHODS: New hair cell formation in this in vitro preparation was shown by sequential photomicroscopy. Studies with the selective marker for mitotic activity 5-bromo-2-deoxyuridine (BrdU) were done to estimate the level of cell proliferation and to quantify postmitotic hair cell formation. Finally, confirmation of cell type was obtained by scanning electron microscopy and by the use of specific markers for hair cells. RESULTS: Once the optimal culture conditions were established in the initial experiments, the formation of new hair cells was directly visualized in all unstained live cultures and fixed preparations without exception. Asymmetric division of progenitor cells, with subsequent differentiation of one of the daughter cells into new hair cells, also was documented by photomicroscopy. Approximately 12% of the cells were labeled with BrdU, of which 6% were hair cells, showing that new hair cell formation was subsequent to mitotic division in vitro. The identity of newly formed hair cells was verified as follows: 1) morphologically by scanning electron microscopy; 2) by positive labeling with phalloidin-rhodamine, a marker for actin; and 3) by positive calmodulin immunocytochemistry. CONCLUSIONS: This study reports the development of an in vitro culture preparation in which undifferentiated epithelial cells proliferate to become new hair cells. Evidence is provided of division of hair cell progenitors and subsequent differentiation of the daughter cells as one of the mechanisms involved in new hair cell formation in the culture preparation. This newly developed cell culture technique provides a powerful tool for further study of the process of hair cell formation in the vestibular end organ.     

Mechanisms and effects of transepithelial polarization in the isolated semicircular canal

Previous studies have shown that galvanic stimulation of semicircular canal organs can modulate their afferent discharge. However, it has not been resolved whether this modulation derived from direct stimulation of hair cells, afferent nerve fibers, some combination of the two, or some as yet unknown path. This problem is addressed in the present study. Experiments were designed first to determine the gross current path necessary for the DC current to modulate afferent firing. These led to the conclusion that the current path had to flow between endolymph and perilymph across the neuroepithelium. Next, the various components in this established path were considered: the afferents, the hair cells, between the hair cells, or some combination of the three. These experiments led to the conclusion that the current pathway was across the hair cells causing transmitter release and thus affecting afferent activity.     

Macrophage and microglia-like cells in the avian inner ear

Recent studies suggest that macrophages may influence early stages of the process of hair cell regeneration in lateral line neuromasts; numbers of macrophages were observed to increase prior to increases in hair cell progenitor proliferation, and macrophages have the potential to secrete mitogenic growth factors. We examined whether increases in the number of leukocytes present in the in vivo avian inner ear precede the proliferation of hair cell precursors following aminoglycoside insult. Bromodeoxyuridine (BrdU) immunohistochemistry was used to identify proliferating cells in chicken auditory and vestibular sensory receptor epithelia. LT40, an antibody to the avian homologue of common leukocyte antigen CD45, was used to label leukocytes within the receptor epithelia. Macrophages and, surprisingly, microglia-like cells are present in normal auditory and vestibular sensory epithelia. After hair cell loss caused by treatment with aminoglycosides, numbers of macrophage and microglia-like cells increase in the sensory epithelium. The increase in macrophage and microglia-like cell numbers precedes a significant increase in sensory epithelial cell proliferation. The results suggest that macrophage and microglia-like cells may play a role in releasing early signals for cell cycle progression in damaged inner ear sensory epithelium.     

Electrophysiological properties of vestibular sensory and supporting cells in the labyrinth slice before and during regeneration

The whole cell patch-clamp technique in combination with the slice preparation was used to investigate the electrophysiological properties of pigeon semicircular canal sensory and supporting cells. These properties were also characterized in regenerating neuroepithelia of pigeons preinjected with streptomycin to kill the hair cells. Type II hair cells from each of the three semicircular canals showed similar, topographically related patterns of passive and active membrane properties. Hair cells located in the peripheral regions (zone I, near the planum semilunatum) had less negative resting potentials [0-current voltage in current-clamp mode (Vz) = -62.8 +/- 8.7 mV, mean +/- SD; n = 13] and smaller membrane capacitances (Cm = 5.0 +/- 0.9 pF, n = 14) than cells of the intermediate (zone II; Vz = -79.3 +/- 7.5 mV, n = 3; Cm = 5.9 +/- 1.2 pF, n = 4) and central (zone III; Vz = -68.0 +/- 9.6 mV, n = 17; Cm = 7.1 +/- 1.5 pF, n = 18) regions. In peripheral hair cells, ionic currents were dominated by a rapidly activating/inactivating outward K+ current, presumably an A-type K+ current (IKA). Little or no inwardly rectifying current was present in these cells. Conversely, ionic currents of central hair cells were dominated by a slowly activating/inactivating outward K+ current resembling a delayed rectifier K+ current (IKD). Moreover, an inward rectifying current at voltages negative to -80 mV was present in all central cells. This current was composed of two components: a slowly activating, noninactivating component (Ih), described in photoreceptors and saccular hair cells, and a faster-activating, partially inactivating component (IK1) also described in saccular hair cells in some species. Ih and IK1 were sometimes independently expressed by hair cells. Hair cells located in the intermediate region (zone II) had ionic currents more similar to those of central hair cells than peripheral hair cells. Outward currents in intermediate hair cells activated only slightly more quickly than those of the cells of the central region, but much more slowly than those of the peripheral cells. Additionally, intermediate hair cells, like central hair cells, always expressed an inward rectifying current. The regional distribution of outward rectifying potassium conductances resulted in macroscopic currents differing in peak-to-steady state ratio. We quantified this by measuring the peak (Gp) and steady-state (Gs) slope conductance in the linear region of the current-voltage relationship (-40 to 0 mV) for the hair cells located in the different zones. Gp/Gs average values (4.1 +/- 2.1, n = 15) from currents in peripheral hair cells were higher than those from intermediate hair cells (2.3 +/- 0.8, n = 4) and central hair cells(1.9 +/- 0.8, n = 21). The statistically significant differences (P < 0.001) in Gp/Gs ratios could be accounted for by KA channels being preferentially expressed in peripheral hair cells. Hair cell electrophysiological properties in animals pretreated with streptomycin were investigated at approximately 3 wk and approximately 9-10 wk post injection sequence (PIS). At 3 wk PIS, hair cells (all zones combined) had a statistically significantly (P < 0.001) lower Cm (4.6 +/- 1.1 pF, n = 24) and a statistically significantly (P < 0.01) lower Gp(48.4 +/- 20.8 nS, n = 26) than control animals (Cm = 6.2 +/- 1.6 pF, n = 36; Gp = 66 +/- 38.9 nS, n = 40). Regional differences in values of Vz, as well as the distribution of outward and inward rectifying currents, seen in control animals, were still obvious. But, differences in the relative contribution of the expression of the different ionic current components changed. This result could be explained by a relative decrease in IKA compared with IKD during that interval of regeneration, which was particularly evident in peripheral hair cells. (ABSTRACT TRUNCATED)