Transient receptor potential channels in the inner ear: presence of transient receptor potential channel subfamily 1 and 4 in the guinea pig inner ear

CONCLUSION: The results of this study indicate that transient receptor potential subfamily 1 (TRPV1) may play a functional role in sensory cell physiology and that TRPV4 may be important for fluid homeostasis in the inner ear. OBJECTIVE: To analyze the expression of TRPV1 and -4 in the normal guinea pig inner ear. MATERIAL AND METHODS: Albino guinea pigs were used. The location of TRPV1 and -4 in the inner ear, i.e. cochlea, vestibular end organs and endolymphatic sac, was investigated by means of immunohistochemistry. RESULTS: Immunohistochemistry revealed the presence of TRPV1 in the hair cells and supporting cells of the organ of Corti, in spiral ganglion cells, sensory cells of the vestibular end organs and vestibular ganglion cells. TRPV4 was found in the hair cells and supporting cells of the organ of Corti, in marginal cells of the stria vascularis, spiral ganglion cells, sensory cells, transitional cells, dark cells in the vestibular end organs, vestibular ganglion cells and epithelial cells of the endolymphatic sac.     

Expressions of aquaporin-2, vasopressin type 2 receptor, transient receptor potential channel vanilloid (TRPV)1, and TRPV4 in the human endolymphatic sac

OBJECTIVE: To localize aquaporin (AQP)2, vasopressin type 2 receptor (V2-R), and transient receptor potential channel vanilloid subfamily 1, 4 (TRPV1, TRPV4) in the human endolymphatic sac (ES). METHODS: Three samples of human ES were sampled during the removal of vestibular schwannoma by way of the translabyrinthine approach. The samples were immediately fixed in 4% paraformaldehyde and embedded in OCT compound; immunohistochemistry was performed with AQP2, V2-R, TRPV1, and TRPV4 polyclonal antibodies. RESULTS: AQP2, V2-R, TRPV1, and TRPV4 proteins were detected in the epithelial layer of the ES but were not observed in connective tissue around the ES. TRPV1 was also expressed in blood vascular endothelial cells of the connective tissue of ES. CONCLUSIONS: AQP2, V2-R, and TRPV4 were expressed in the luminal epithelium of human ES. The same characteristic distribution of water and ion channels is seen in the kidney, where a significant amount of fluid is filtrated and resorbed. ES probably plays an active role in the homeostasis of the endolymph.     

The influence of transient asphyxia on receptor potentials in inner hair cells of the guinea pig cochlea

Intracellular recordings were made from inner hair cells in the basal turn of the guinea pig cochlea during transient asphyxia. During these periods the endocochlear potential was reversibly reduced, the hair cell resting membrane potential was slightly hyperpolarized and the cochlear microphonic potential was decreased, all with similar time courses. The eighth nerve compound action potential and DC and AC components of the inner hair cell receptor potential were strongly attenuated and with similar slow time courses. The asymmetrical low frequency receptor potentials became symmetrical, and this change was attributed to alterations in the mechanical properties of the organ of Corti. The desensitization of the cochlea during transient asphyxia was associated with the loss of asymmetry of the inner hair cell receptor potential.     

Dynamic aspects of guinea pig inner hair cell receptor potentials with transient asphyxia

DC and AC receptor potentials of cochlear inner hair cells in response to tone bursts of various frequencies and intensities were continuously measured during and following periods of transient asphyxia. The effects of asphyxia were most pronounced for low sound pressure level (SPL) acoustic stimuli near the characteristic frequency (CF) of the inner hair cell, leading to vulnerability of the 'tip' of the cell's frequency tuning curve (FTC). The resulting changes in the shape of the FTC are, first, a reduction in tip criterion sensitivity of 10-20 dB without significant loss in sharpness of tuning. Later, when the full effect of 30-45 s asphyxia occurs, tip sensitivity loss between 30 and 65 dB is accompanied by greatly broadened tuning and a shift downward in frequency of the CF by greater than 1/4 octave. The CF shift is due to a progressive loss of high frequency sensitivity. The linear segment of the input-output (intensity) function, plotted as log DC receptor potential versus SPL (at the original CF), becomes longer during the early phase asphyxia, and the slope of the segment declines by 50%. At high SPLs, for all frequencies, the time course of the receptor potential change was similar in shape to that exhibited by the endocochlear potential (EP). In particular, for high sound levels, the recovery of response matches the EP while for low level tip frequency sounds recovery is protracted. No difference between the decline of the AC and DC receptor potentials at CF was observed. Inner hair cell resting membrane potential (Em) hyperpolarized during asphyxia by 2-6 mV, correlating with the change in EP according to a ratio of 1/10 (Em/EP).     

Distribution of gentamicin by immunofluorescence in the guinea pig inner ear

Summary We studied the distribution of gentamicin in the inner ear, brain and kidney of the guinea pig following intraperitoneal administration or perfusion of gentamicin through the perilymphatic space. The resulting histopathologcial changes were examined by immunofluorescence using antigentamicin antiserum. After perfusion of gentamicin through the perilymphatic space, specific fluorescence was found in the cochlea, and was especially prominent in the outer hair cells, basilar membrane and basilar crest. Although no fluorescence was observed in the cochlea following intraperitoneal administration of high doses of gentamicin, type I hair cells in the vestibule were seen to be selectively stained with the antibody. Furthermore, some of the vestibular ganglion cells, Purkinje cells and unidentified nuclei in the brain stem were also stained. In particular, fine granules showing relatively intense fluorescence were recognized in the cytoplasm of the stained cells. In the cortex of kidney, only proximal tubular cells were stained with intense fluorescence. Our results suggest that the aminoglycoside antibiotics have two sites of action: one is the cell membrane of the sensory hair cells and the other is the cytoplasm.This study was supported in part by a grant from the Ministry of Education, Science and Culture of Japan, and by a Research Grant for Specific Diseases from the Ministry of Health and Welfare to the Acute Profound Deafness Research Committee of Japan     

Localization of melatonin and its receptors (melatonin 1a and 1b receptors) in the mouse inner ear

Abstract

Background: In the inner ear, evidence has been gathered indicating that melatonin plays important roles in inner ear physiology and pathophysiology. However, no attempt has been made previously to investigate the localization or expression of melatonin and its receptors in the whole inner ear.

Aims/objectives: To analyze the presence of melatonin and its receptors in the normal mouse inner ear.

Material and methods: C57BL6/J mice were used in this study. The localizations of melatonin, MT1a and MT1b in the inner ear, i.e. cochlea, vestibular end organs, vestibular ganglion and endolymphatic sac (ES), were studied by immunohistochemistry.

Results: The organ of Corti, spiral ganglion, vestibular ganglion, vestibular sensory cells, vestibular dark and transitional cells, and ES epithelial cells showed an immunofluorescence reaction to melatonin, MT1a and MT1b.

Conclusion and significance: The present findings show that melatonin and its receptors (MT1a and MT1b) are present in the inner ear, thus supporting the hypothesis that melatonin plays a physiological role in the inner ear.     

Vasopressin and isoproterenol activate adenylate cyclase in the guinea pig inner ear

Summary Cochleas from guinea pigs were perfused by isotonic buffer after punction of the carotid artery. The cochlea tissue was removed from the bony capsule and separated from the mediolus as band with a sharp needle under the microscope. Cell membranes were prepared subsequently from whole tissue. Purified membranes from the inner ear of guinea pigs contain adenylate cyclase which functionally is coupled with membrane receptors for Vasopressin and- receptors for isoproterenol (epinephrine), respectively. Both hormones stimulate production of cyclic AMP at 37° C.Furthermore, cyclase activity is increased by addition of Gpp (NH)p, a GTP analog. Possible relationships of these molecular events to cochlear events such as glycogenolysis, ionfluxes, transport and secretion mechanisms, and synaptic transmission are discussed.Part of this work was presented at the 15th Workshop on Inner Ear Biology in Seefeld/Innsbruck, Austria, September 3–6, 1978     

Postnatal development of substance P in the inner ear of the guinea pig

Summary Appreciable amounts of substance P (SP) were found in guinea pig cochleas. The highest values were found in the postnatal period. Data presented favor the assumption of SP acting as a neuromodulator or neurotransmitter in the inner ear.     

The preparation of acetic acid for use in otic drops and its effect on endocochlear potential and pH in inner ear fluid

The ototoxicity of an otic drop preparation containing 2% acetic acid and 3% propylene glycol (VoSol, Denver Chemical Co., Humacao, PR) was investigated according to measurements of endocochlear potential (EP) and inner ear fluid pH. The application of this preparation to the round window membrane for 30 minutes caused a depression in EP from 80.5 +/- 2.5 mV (mean +/- SD; n = 6) to 11.7 +/- 7.7 mV, and lowered inner ear fluid pH from 7.55 +/- 0.09 to 5.06 +/- 0.19 (n = 6) in perilymph and from 7.52 +/- 0.07 to 5.88 +/- 0.63 (n = 6) in endolymph. Two percent acetic acid produced similar changes after 30 minutes: EP was reduced from 83.0 +/- 2.2 mV to 34.0 +/- 2.9 mV and endolymphatic pH from 7.49 +/- 0.04 to 6.83 +/- 0.21 (n = 4). However, the application of artificial perilymph of pH 4 titrated with HCl induced no significant changes in either EP or endolymphatic pH. We suggest that the mechanisms of ototoxicity in the otic drop preparation are Na+ and K+-ATPase inhibition, and that such inhibition is due to the intracellular acidification of strial cells resulting from the penetration of acetic acid across the cell membrane, and to the direct and synergistic actions of propylene glycol.     

Localization of histamine (H1, H2, H3 and H4) receptors in mouse inner ear

Conclusion The present findings show that all four types of histamine receptors (H1R, H2R, H3R, and H4R) are present in the inner ear, thus supporting the hypothesis that histamine plays a physiological role in the inner ear. Objective To analyse the presence of histamine receptors in the normal mouse inner ear. Methods CBA/J mice were used in this study. The localization of H1R, H2R, H3R, and H4R in the inner ear, i.e. cochlea, vestibular end organs, vestibular ganglion, and endolymphatic sac, was studied by real-time PCR and immunohistochemistry. Results The mRNA for each receptor sub-type was detected in the inner ear. In the immunohistochemical study, the organ of Corti, spiral ganglion, vestibular ganglion, vestibular sensory epithelium, and endolymphatic sac cells showed an immunofluorescent reaction to all histamine receptors.     

A comparison of acidic glycosaminoglycans in the inner ear and other organs: Electrophoretic microanalysis

Summary Acidic glycosaminoglycans in the ground substance of the membranous cochlear lateral wall, kidney, brain and liver of the guinea pigs were quantitatively and qualitatively analyzed, using electrophoretic microanalysis. The lateral wall of the cochlea showed highest content (0.46±0.08%/D.W.) of acidic glycosmainoglycans, which were chondroitin sulfate-B (75%), chondroitin sulfate-A (12%) and hyaluronic acid (13%). However, the pattern of these acidic glycosaminoglycans in the inner ear differed from the other organs. A possible role of acidic glycosaminoglycans in the inner ear was discussed.     

颞骨高分辨率CT在中、内耳手术中的应用

目的分析颞骨高分辨率CT在中、内耳手术中的应用。方法回顾分析近3年的600例中、内耳手术前颞骨CT和术中病变情况。结果术前CT诊断慢性化脓性中耳炎、中耳胆脂瘤、中耳炎后遗症(包括鼓膜穿孔、粘连性中耳炎、鼓室硬化症)、耳硬化症和中耳畸形及恶性肿瘤的符合率分别为98.7%(221/224耳)、94.4%(319/302耳)、94.1%(32/38耳)、66.7%(16/24耳)和100%(12/12耳)。结论颞骨CT提供了大量关于中、内耳的信息,临床医师术前应该认真阅读分析颞骨CT片,充分利用CT提供的信息,做好手术的个体化设计,提高治疗质量。     

肿瘤坏死因子受体1在中耳胆脂瘤上皮中的表达

目的 研究肿瘤坏死因子受体1（TNFR1）在中耳继发性胆脂瘤上皮的表达，探讨其在胆脂瘤型中耳炎的发病机制中的作用。方法 分别采用免疫组化法和Western blot技术检测TNFR1在36例中耳胆脂瘤组织及20例正常外耳道皮肤组织中的表达情况。结果 TNFR1在36例胆脂瘤上皮各层细胞中均有强表达，定位于胞膜和胞质，而外耳道皮肤表达较弱甚至没有表达，TNFR1在胆脂瘤上皮中的蛋白表达水平较外耳道皮肤显著增高，差异有统计学意义（P〈0．01）。结论 胆脂瘤上皮中TNFR1表达较外耳道皮肤显著增高，肿瘤坏死因子可能通过与TNFR1相结合而导致胆脂瘤上皮细胞的过度增殖及凋亡。     

Effect of acute inner ear pressure changes on low-level distortion product otoacoustic emissions in the guinea pig

Objective—To determine a relation between acute inner ear pressure changes and cochlear function as measured by low-level 2f₁–f₂ distortion product otoacoustic emissions (DPOAEs).

Material and Methods—During and after a change in inner ear pressure induced by injection or aspiration of perilymph, the 2f₁–f₂ DPOAE at 4.5 kHz generated by low-level primaries was recorded in the guinea pig.

Results—Large changes in overall inner ear pressure produced only small changes in the 2f₁–f₂ amplitude and phase. During injection of 0.5 μl of artificial perilymph into the scala tympani over a 10-s period, the mean inner ear pressure increased by ≈500 Pa, with an accompanying mean increase in the 2f₁–f₂ amplitude of 0.7 dB. During aspiration of 0.5 μl of perilymph over a 10-s period, the mean inner ear pressure decreased by ≈700 Pa, with an accompanying mean decrease in the 2f₁–f₂ amplitude of 0.9 dB. Changes in DPOAE amplitude followed inner ear pressure changes with a delay of 1–2 s. The magnitude and sign of the amplitude changes can (partly) be explained by a change in oval window stiffness. No explanation was found for the measured delay.

Conclusion—Clinically, these experiments can be of value in gaining insight into the pathophysiological mechanisms of pathological pressure changes as seen in Ménière's disease and perilymphatic fistulae.     

Expression of epidermal growth factor, epidermal growth factor receptor and transforming growth factor-alpha in the human fetal inner ear

The expression of epidermal growth factor (EGF), EGF receptor and transforming growth factor (TGF)-alpha was analyzed in the human fetal inner ear using immuno-histochemical methods. EGF receptor was observed only in 9.5-week-old fetal vestibular epithelia. In 14- and 16-week-old fetuses, EGF receptor could not be detected. TGF-alpha was observed strongly in the 9- and 11-week-old vestibular epithelia, whereas only trace amounts were detectable in the 14- and 16-week-old vestibular epithelia. These findings suggest that EGF and TGF-alpha probably have a mitogenic effect in the sensory epithelia of the fetal inner ear, especially at early stages of development.     

Localization of sirtuins (SIRT1-7) in the aged mouse inner ear

Conclusion: The expression of sirtuin in vestibular end organs and cochlea responds differently to age-related changes. Down-regulation of SIRT1, 3, and 5 in the cochlea may weaken the protective activity regarding degeneration of the organ of Corti as well as of spiral ganglion cells, resulting in the development of age-related hearing loss. An increase in SIRT 1, 4, or 5 in vestibular tissue could indicate an increased need of detoxification of reactive oxygen species and an increased anti-ageing potential. Objective: To analyse the expression of sirtuins (SIRT1-7) in the normal young and old mouse inner ears. Methods: Young (8 weeks) and old (22 months) CBA/J mice were used in this study. Localization of SIRT1-7 in the inner ear, i.e. cochlea, vestibular end organs, and vestibular ganglion, was investigated using real-time PCR and immunohistochemistry. Results: In the vestibular end organs, the expression of SIRT1, 2, 4, 5 (both mRNA and protein), SIRT6, and 7 (only mRNA) was found to be increased, while a slightly decreased immunoreactivity was observed in SIRT3. In the cochlea, the expression of SIRT1, 3, and 5 (both mRNA and protein) was decreased in the old mice, whereas no noticeable difference was observed regarding SIRT2, 4, 6, or 7.     

The effects of low-frequency ultrasound on the inner ear: an electrophysiological study using the guinea pig cochlea

Summary By examining 218 albino guinea pigs, electrophysiological methods were used to investigate the effects of low frequency ultrasound at moderate sound pressure levels after long-term exposure to the inner ear. From 10 kHz to 28 kHz, low frequency ultrasound below 100 dB SPL induced significant changes in cochlear microphonics, elevated thresholds and decreased maximum output voltage of action potentials and decreased absolute values of negative potentials of the endocochlear potentials.     

The use of immunofluorescence in the non-decalcified frozen guinea pig cochlea to detect autoantibodies in inner ear disorders

Summary Autoantibodies in the serum of patients with inner ear disorders have been detected using the immunofluorescence technique. In the present study the results of the use of frozen sections of non-decalcified guinea pig cochlea in the indirect immunofluorescence test are described. Human sera with positive anti-mitochondrial (AMA), anti-nuclear (ANA^S) and anti-nucleolar (ANA^N) antibodies were utilized, as well as a negative serum control and sera from three patients suffering from chronic progressive sensorineural hearing loss. The positive reactions obtained were clear and specific. Autofluorescence was minimal and non-specific reactions were negligible. The fine structure of the inner ear tissue and the cellular antigenicity present were well preserved.