Non-linear response to amplitude-modulated waves in the apical turn of the guinea pig cochlea

Mechanical vibrations of the Hensen's cells were measured in the apical turn of the cochlea in living guinea pigs, in response to amplitude-modulated (AM) sound. The FFT of the input wave consisted of spectral components at the carrier frequency C and two sidebands (C+/-M) separated from the carrier by the modulation frequency M. The FFT of the velocity response consisted of components at: (i) the modulation frequency M, and harmonics n M; (ii) Carrier frequency C and sidebands (C+/-n M); (iii) harmonics of the carrier frequency and their side bands (2C+/-n M); (3C+/-n M); (4C+/-n M); em leader n=1,2,3, em leader,10. The carrier and the first pair of side bands were broadly tuned and nearly linear. Other components were sharply tuned and highly non-linear, suggesting a different origin. Evidence is presented that these components are generated in the non-linear stereocilia dynamics. An important function of this non-linearity is to demodulate the AM wave to extract information contained in the modulation.     

Vibrations of the guinea pig organ of Corti in the apical turn

Vibrations of the organ of Corti were measured in response to sound applied to the ear in the apical turn of a living guinea pig. Measurements were made at 29 points on the Reissner's membrane (RM) at 10 micro spacing along a radial track. Measurements also included 22 points on the reticular lamina (RL), Claudius' cells and osseous spiral lamina. Our goal was to characterize the vibration of the RM and the RL with high spatial resolution along a radial axis. The tuning and spatial patterns of the RM are compared in the radial direction with those for the RL at the fundamental frequency and at the second harmonic. The shape of the RM tuning curve changes with radial position, and differ significantly from those observed at the RL. These results support our earlier findings (Hao and Khanna, Hear. Res. 99 (1996) 176-189).     

Mechanical nonlinearity in the apical turn of the guinea pig organ of Corti

Confocal microscopy was used to view the sealed apical turn of the cochlea in a living guinea pig, and to identify the cochlear structures through the intact Reissner's membrane. X, Y and Z coordinates for each point of interest were recorded. A confocal laser heterodyne interferometer measured the cellular vibration in response to acoustical signals applied to the ear. Velocity time waveforms were recorded at 32 frequencies between 25 and 2500 Hz at each point of measurement. To characterize the vibration pattern of the organ of Corti, vibrations at multiple locations along a radial track of the reticular lamina were measured before and after sacrificing the animal. Amplitude and phase tuning curves of the fundamental and the second harmonic, velocity time waveforms, and FFTs of time waveforms are compared before and after sacrifice. The results show that a sharply tuned nonlinear part of the response disappears shortly after sacrifice.     

Displacement of the reticular lamina with and without the tectorial membrane in the guinea pig cochlea

OBJECTIVE: In an attempt to establish the mechanical relationships between the reticular lamina and tectorial membrane, we studied the morphological changes of the reticular lamina on a micrometer scale when an in vitro preparation of guinea pig cochlea with and without tectorial membrane was exposed to a potassium-rich medium. MATERIAL AND METHODS: Using video-enhanced differential interference contrast microscopy, the radial displacement of the inner hair cells (IHCs) and outer hair cells (OHCs) in the reticular lamina was measured in real time after exposure to the potassium-rich medium for 3 min. RESULTS: The amplitude of the displacement of the OHCs in preparations with an intact tectorial membrane was half of that observed in those in which the tectorial membrane had been removed. A similar displacement response was also observed for the IHCs, although it was smaller than that for the OHCs. There was no significant difference in the amplitude of the displacement among the three rows of OHCs. CONCLUSIONS: These results suggest that the structure linking the OHCs to the pillar cells is very elastic and that the movement of the OHCs in situ is weakly mechanically coupled to the IHCs. The tectorial membrane provides increased compliance in the motion of the IHCs and OHCs.     

Amplification in the apical turn of the cochlea with negative feedback

The apical turn of the anesthetized guinea pig cochlea was opened to examine the basilar membrane optically through the intact Reissner's membrane. Vibrations of the outer Hensen's cell and the basilar membrane (BM) adjacent to and about 130 microm below the level of the Hensen's cell were measured. Outer Hensen's cell vibration at the characteristic frequency was up to 900 times higher compared to the BM amplitude. After sacrifice BM vibration increased while Hensen's cell vibration decreased. The magnitude and sequence of change after sacrifice can best be explained by the presence of negative feedback between reticular lamina and BM. In other experiments using ototoxic drugs that damage outer hair cells, similar changes in Hensen's cell and BM vibration were observed. These results show that the apical turn behavior is different from that observed by other investigators in the basal turn. The potential benefits of the negative feedback are discussed. The presence of negative feedback would explain the linearity at the fundamental frequency observed in the apical turn of cochlea.     

Effects of a dopaminergic agonist in the guinea pig cochlea

This study investigates the role of dopamine, a putative lateral efferent neurotransmitter/modulator, in cochlear physiology and physiopathology. Cochlear potentials were recorded in guinea pigs after intracochlear perfusion of increasing doses (0.1–1 mM) of piribedil, an agonist of the D2/D3 receptors. A dose-dependent reduction in the amplitude of auditory nerve compound action potential (CAP) was observed, predominantly at high-intensity tone-burst stimulations, and without significant effect on CAP threshold. There was no variation of cochlear microphonic and summating potential.

When 1 mM piribedil was perfused into the cochlea during continuous 130 dB SPL pure tone exposure (6 kHz, 15 min), CAP threshold shifts were significantly less than in control animals with artificial perilymph-perfused cochleas. No dendritic damage was observed, although there was evident hair cell damage. Similarly, radial dendrites were clearly protected against ischemia-induced damage when 1 mM piribedil was applied prior to a 10-min ischemia.

These results suggests that dopamine modulates the activity of radial afferent fibers via D2/D3 receptors. The protective effect of piribedil during acoustic trauman or ischemia suggests that this modulation corresponds to a prevention of excitotoxicity due to dysfunction of inner hair cell neurotransmission.     

Cyclic GMP-dependent protein kinase-I in the guinea pig cochlea.

Recent studies have begun to characterize the nitric oxide/cyclic GMP/protein kinase G pathway in the mammalian cochlea by demonstrating the presence of both the enzyme that produces nitric oxide (NO), nitric oxide synthase, and the NO receptor, soluble guanylate cyclase. The present study investigated protein kinase G (cyclic GMP-dependent protein kinase-I, cGK-I), the downstream enzyme of this pathway that frequently mediates its physiological effects. A commercial antibody to a human cGK-I sequence recognized a protein of appropriate molecular weight in Western blots of guinea pig aorta. Immunostaining of guinea pig aorta was consistent with the expected distribution of cGK-I. In lateral wall tissues of the cochlea, pericytes lining the blood vessels of the spiral ligament were strongly immunoreactive. In the organ of Corti, cGK-I was detected in Hensen's, Deiters', and pillar cells, but not in inner and outer hair cells. This distribution coincides with the localization of soluble guanylate cyclase activity and suggests that cGK-I mediates the effects of the NO/cyclic GMP pathway in the cochlea. It reinforces the hypothesis that the NO/cyclic GMP/cGK-I pathway is involved in regulation of cochlear blood flow and supporting cell physiology.     

Electrically evoked otoacoustic emissions from apical and basal perilymphatic electrode positions in the guinea pig cochlea

Stimulation of the cochlea with sinusoidal current results in the production of an otoacoustic emission at the primary frequency of the stimulus current. In this study we test the hypothesis that the wide frequency response from round window (RW) stimulation is due to the involvement of a relatively large spatial segment of the organ of Corti. Tonotopically organized group delays would be evident from perilymphatic electrode locations that restrict the spatial extent of hair cell stimulation. Monopolar and bipolar-paired stimulus electrodes were placed in perilymphatic areas of the first or third cochlear turns and the electrically evoked otoacoustic emissions (EEOAE) produced by these electrodes were compared to that from the RW monopolar electrode in the anesthetized guinea pig. Current stimuli of 35 microA RMS were swept across the frequency range between 60 Hz and 100 kHz. The EEOAE was measured using a microphone coupled to the ear canal. It was found that the bandwidth of EEOAEs from RW stimulation extended to at least 40 kHz and was a relatively insensitive to electrode location on the RW. The group delay of the EEOAE from stimulation at the RW membrane (corrected to stapes motion) was about 53 micros. First and third turn stimulations from electrode placements in perilymph near the bony wall of cochlea yielded narrower band EEOAE magnitude spectra but which had the same short group delays as for RW stimulation. A confined current (from a bipolar electrode pair) applied close to the basilar membrane (BM) in the first turn produced the narrowest frequency-band magnitude emissions and a mean corrected group delay of 176 micros for a location approximately 3 mm from the high frequency end of the BM (corresponding to about the 18 kHz best frequency location). Bipolar electrodes in the third turn scala tympani produced low pass EEOAE magnitude functions with corrected group delays ranging between approximately 0.3 and 1 ms. The average phase slopes did not change with altered cochlear sensitivity and postmortem. These data indicate that the EEOAE from RW stimulation is the summed response from a wide-tonotopic distribution of outer hair cells. A preliminary model study indicates that short time delayed emissions are the result of a large spatial distribution of current applied to perilymphatic locations possibly giving rise to "wave-fixed" emissions.     

Melanocytes in the modiolus of the guinea pig cochlea

The cochleae of pigmented and albinotic guinea pigs aged 6-8 weeks were examined by electron microscopy for melanocytes in the modiolus. Two different types of melanocyte were observed: i) The first (Type A) was characterized by spherical, uniformly pigmented stage IV melanosomes. No signs of melanogenesis were seen in this perivascular pigment cell form, which is found mainly in the region of the cochlear plexus. ii) The second, very rarely observed type of melanocyte (Type B) was characterized by fusiform to oval melanosomes in various stages of maturation. Connective tissue cells containing fusiform to oval melanosomes enclosed in membrane-bound vacuoles were frequently found. These two types can be regarded as corresponding to uveal (Type A) and dermal (Type B) melanocytes. The connective tissue of the cochlear plexus seems to be derived from the leptomeninx.     

Immunocytochemical detection of peptides in the guinea pig cochlea

The cochleae of juvenile guinea pigs were investigated for the presence of several neuropeptides. Glucagon, insulin, CCK and beta-endorphin immunoreactive neurons and nerve fibers as well as hair cells were demonstrated by the peroxidase antiperoxidase technique. Small amounts of substance P were also found in different sites in the inner ear. In contrast, prolactin-like material could not be found at all. These findings have significance with regard to the putative role of neuropeptides in neuromodulation.     

Potassium circulation in the perilymph of guinea pig cochlea

To determine whether any differences exist in potassium circulation between the scala vestibuli and scala tympani, we recorded the change in K⁺ activity in both scalae of the guinea pig cochlea at the basal and third turns, using a double-barrelled, K⁺-sensitive microelectrode after perfusion with artificial perilymph containing 20 mM KCl and 130 mM NaCl. K⁺ activity increased immediately after the start of perfusion and decreased after its completion. The rates of decrease of K⁺ activities were approximately 1.0 mEq/l per min in the scala vestibuli of the basal and third turns, also 1.0 mEq/l per min in the scala tympani of the basal turn, and approximately 0.5 mEq/l per min in the scala tympani of the third turn. The rate of decrease of K⁺ activity in the scala tympani was significantly slower in the third turn than in the basal turn. Blockage of the cochlear aqueduct depressed the rate of decrease of K⁺ activity in the scala tympani more in the basal turn than in the third turn. These results suggest that there is a difference in potassium circulation between the scala vestibuli and scala tympani, and that the cochlear aqueduct plays an important role in potassium circulation in the perilymph of the scala tympani.     

Localization of laminin isoforms in the guinea pig cochlea

OBJECTIVE: To elucidate the pathogenesis of inner ear disorders by examining the distribution of a major component of basement membranes, the glycoprotein laminin. STUDY DESIGN: Animal model. METHODS: Adult guinea pig cochleae were fixed with methanol and sectioned. Sections were examined for the presence of laminins alpha2, alpha5, beta1, and gamma1 using immunohistochemistry. RESULTS: Laminins alpha2, alpha5, and beta1 were not detected. Laminin gamma1 was found in the limbus, spiral ligament, and stria vascularis. CONCLUSIONS: Anti-laminin antibodies have been found in patients with inner ear disorders. Laminin gamma1 demonstrates specific staining at multiple sites in the guinea pig cochlea. These structures may be targets of antibodies causing sensorineural hearing loss.     

Immunocytochemical detection of peptides in the guinea pig cochlea

Summary The cochleae of juvenile guinea pigs were investigated for the presence of several neuropeptides. Glucagon, insulin, CCK and -endorphin immunoreactive neurons and nerve fibers as well as hair cells were demonstrated by the peroxidase antiperoxidase technique. Small amounts of substance P were also found in different sites in the inner ear. In contrast, prolactin-like material could not be found at all. These findings have significance with regard to the putative role of neuropeptides in neuromodulation.     

Ultrastructure of cultured marginal cells of the guinea pig cochlea.

Explants of stria vascularis and spiral ligament of guinea pig cochlea were kept in primary culture. On the explant, proliferating marginal cells advanced by 15 microns/day, suggesting that in vivo defects of the strial epithelium can be covered by new marginal cells. The marginal cells growing in the cell culture dish had a diameter of 12.8 +/- 0.7 microns and formed an epithelial monolayer. Adjacent cells were connected by desmosomes and tight junctions. The cells were uniformly polarized. The apical membrane had small invaginations and numerous microvillus-like extensions, and the convoluted lateral membrane interdigitated with adjacent cells. The basal infoldings were smaller in cultured cells than in vivo; mitochondria were dispersed in the entire cytoplasm rather than concentrated in basolateral infoldings. The basal membrane infoldings of cultured marginal cells did not interdigitate with underlying fibroblast-like cells. Marginal cells were separated from underlying fibroblast-like cells by a fluid-filled space which was sometimes enlarged, leading to the formation of "domes" in the otherwise planar epithelium.     

Effects of bacterial endotoxin applied to the guinea pig cochlea.

The ototoxic potential of bacterial endotoxin present in middle ear effusions was evaluated by inoculating endotoxin solutions of 10 micrograms/mL or 1000 micrograms/mL directly into the scala tympani of the guinea pig cochlea. Median auditory brainstem response threshold shifts of 12.5 and 20 dB were observed in the experimental ears exposed to the higher dose at days 2 and 4, respectively, with gradual improvement to near baseline by day 28. Histologic examination revealed inflammatory infiltrates in the scala tympani and scala vestibuli of several of these cochleas. The lower concentration, closer to that found in human effusions, elicited no significant loss of hearing and a milder inflammatory response. It is concluded that brief exposure of the cochlea to endotoxin from middle ear effusions is insufficient to cause hearing loss. Under chronic conditions, however, persistent inflammation within the cochlea may result in a sensorineural deficit.     

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.