Decarboxylation of indomethacin induced by heat treatment.

As crystalline indomethacin is heated and subsequently cooled, it transforms into glassy indomethacin. While the original crystals are off-white in color, the glass becomes blackish-brown via a yellow intermediate stage. TLC of the components of the glass revealed three bands. The yellow component, which is generated either under hypoxic conditions or in the dark, was elucidated by NMR spectroscopy to be a decarboxylated fragment produced by thermal degradation. The colorless component is proposed to be formed by the opening of the indole ring of indomethacin; the structure of this degradation product was identified by EI-MS to be the same as the oxidative-cleavage product formed upon UV-irradiation, as previously reported. Another band was a blackish-brown pigment whose mobility placed it close to the TLC baseline. This oxidative-cleavage product and the blackish-brown pigment are not generated under hypoxic conditions. However, the extent of indomethacin decarboxylation under hypoxic conditions was found to be dependent on the heating temperature and time. Consequently, we prepared amorphous indomethacin through control of the heating temperature and time; heating at 160 °C for 30 min or less under hypoxic conditions is optimum for obtaining pure amorphous indomethacin.

Ultrastructural localization of cochlin in the rat cochlear duct.

Cochlin, a product of the COCH gene, is a major constituent of the inner ear extracellular matrix. Type II collagen, a protein that contributes to structural stability, is also a component of this extracellular matrix. In this study, using the postembedding immunogold method, we demonstrate the localization of cochlin and type II collagen in the cochlear duct at the ultrastructural level. The immunolabeling of cochlin was observed in the fibrillar substance in the spiral limbus, beneath the inner sulcus cells, and in the basilar membrane, the spiral prominence and the spiral ligament. Immunolabeling of type II collagen was observed in the same fibrillar substance in the extracellular matrix of the cochlear duct. This localization of cochlin is consistent with the expected localization of type II collagen. The localization of cochlin and type II collagen indicates the important roles played by these proteins in the hearing process.

Localization of megalin in rat vestibular dark cells and endolymphatic sac epithelial cells.

CONCLUSION: Megalin immunoreactivity was observed in kidney proximal tubule cells, vestibular dark cells, and epithelial cells of the endolymphatic sac. Endocytic mechanisms appear to differ between the endolymphatic sac and proximal tubule cells. We speculate that megalin is secreted by a certain type of cell into the endolymphatic space, and is then absorbed from the endolymphatic space by another type of cell to maintain endolymphatic sac homeostasis. OBJECTIVES: We previously detected megalin immunoreactivity in the rat cochlear duct. Megalin may be involved in endocytosis in the vestibular organ and endolymphatic sac. To examine this possibility, we extended our immunocytochemical investigation to the rat inner ear cells with special attention to vestibular dark cells and endolymphatic sac. MATERIALS AND METHODS: We observed immunoreactivity of megalin under light and electron microscopy. The primary antibody was rabbit polyclonal antibody that had been raised against rat immunoaffinity-purified megalin. RESULTS: The luminal membrane and subapical area of dark cells in the semicircular canal were immunolabeled. The stainable substance in the endolymphatic space was strongly stained. The cytoplasm of epithelial cells was also stained in various patterns.

Ultrastructural co-localization of cochlin and type II collagen in the rat semicircular canal.

Cochlin and type II collagen are major constituents of the inner ear extracellular matrix. To investigate the morphological relation of cochlin and type II collagen in the rat semicircular canal, immuno-electronmicroscopic analysis was performed using the post-embedding immunogold method. Immunolabeling for cochlin was detected in the fibrillar substance underlying the supporting epithelium of the sensory cells and beneath the epithelial cells facing the endolymph in the semicircular canals. Immunolabeling for type II collagen was observed in the same fibrillar substance in the subepithelial area. The co-localization of cochlin and type II collagen in the fibrillar substance in the subepithelial area indicate that cochlin may play a role in the structural homeostasis of the vestibule acting in concert with the fibrillar type II collagen bundles.

Pattern of cochlear damage caused by short-term kanamycin application using the round window microcatheter method.

CONCLUSION: Short-term local application of kanamycin by the microcatheter method can control the region and severity of drug effect on the cochlea by changing the concentration and administration time. OBJECTIVES: To investigate the effect of short-term round window administration of kanamycin by the microcatheter method on cochlear stereociliary bundle loss in a guinea pig model. MATERIALS AND METHODS: Two concentrations and two time periods were used for drug administration to the inner ear. In groups of six animals, a total dose of 172.5 mg/ml or 345 mg/ml of kanamycin was instilled at the rate of 0.1 ml/h into the round window niche over either a 1 h or 2 h period by the microcatheter method. One group of six animals received a saline infusion as a control. The animals were sacrificed after 4 days and stereociliary bundle loss was observed by SEM. Functional changes were evaluated by auditory brainstem response. RESULTS: With regard to the severity of the damage, higher drug concentrations were associated with more severe effects. The severity of damage was the same when the same total dose was used; however, it was found that when the dose was administered over a longer period, the damage region was wider. The functional changes also corresponded with the morphological changes.

[Comparison of audiological performance bewteen bone-anchored and conventional hearing aids].

The bone-anchored hearing aid (BAHA) is an implantable bone-conduction device that vibrates the skull directly via a surgically implanted titanium screw behind the ear. The BAHA has advantages for patients with aural atresia or chronic ear drainage, who cannot wear air-conduction hearing aids. We compared the function of BAHA to conventional bone and air-conduction hearing aids based on functional gain and speech discrimination tests in quiet and noisy environments in 2 patients with chronic ear problems. All expressed a clear preference for the BAHA over conventional bone-conduction hearing aids. The BAHA and air-conduction hearing aid provided similar audiological performance when the functional gain of each hearing aid coincided. As the air-bone gap widens, however, audiological performance with the BAHA showed an evident preference, indicating that the width of the air-bone gap is of some help in BAHA preference compared to air-conduction hearing aids.

Survey in to the prevalence of hearing loss in patients diagnosed with retinitis pigmentosa.

Sensorineural hearing loss is the most common disease associated with systemic retinitis pigmentosa (RP). We have conducted an epidemiological study to assess the correlation of age at onset of visual symptoms and hearing loss associated with RP. Epidemiological data was derived from a questionnaire-based study of patients who are registered members of the Japanese Retinitis Pigmentosa Society (n = 3200). The questionnaire was mailed to these patients in 2002, and information was requested regarding age at onset of visual disturbance, awareness of hearing loss and the presence of progressive hearing loss, age at onset of hearing loss, awareness of tinnitus, and history of audiometric examination and hearing aid usage. 26.1% of the questionnaires were returned, and data for 828 patients with RP diagnosed by an ophthalmologist were evaluated. Cochlear symptoms were reported by 356 patients (43.0% of the total population), with hearing loss in 29.5%, tinnitus in 31.5% and hearing loss and tinnitus in 39.3% of the 356 patients. Of these 356 patients, progressive hearing loss was reported by 44.9% and was independent of age at onset of cochlear symptoms. The mean age at onset of visual symptoms was higher for patients with progressive hearing loss, and a significant correlation was found between the age at onset of visual symptoms and hearing loss for patients who were older at onset of the symptoms (>30 years of age). Onset of hearing loss occurs later and hearing loss is also more progressive for patients with late onset of RP. This suggests that particular care regarding hearing loss is necessary for this patient population, and that cooperation between opthalmologists and otologists is required for diagnosis of RP-hearing impairment-associated syndromes in this group of patients.

A model of two-stage newborn hearing screening with automated auditory brainstem response.

Our purpose was to evaluate a two-stage newborn hearing screening program using automated auditory brainstem response (AABR) before discharge and to describe our follow-up program. This study used 4085 infants born in the Seirei-Hamamatsu and Mikatahara General Hospitals during a 2-year period. The initial screening test was performed 2 or 3 days after birth at an intensity of 35 dBnHL. For the infants who were referred from this test, the re-screening test was performed 5 or 6 days after birth. Diagnostic work-up with auditory brainstem response (ABR), otoacoustic emissions (OAE), and a conditioned orientation reflex audiometry (COR) test were performed by the age of 3-6 months. The referral rate was 1.20% (49/4085 infants) in the first test and 0.71% (29/4085 infants) in the two-stage screening. The two-stage screening procedure was able to reduce the false-positive rate from 0.83 to 0.34%. The incidence of bilateral and unilateral congenital hearing loss diagnosed by ABR was 8/4085 (0.20%) infants and 7/4085 (0.17%) infants, respectively. One infant with congenital cytomegalovirus infection, who passed the two-stage AABR tests, was diagnosed with hearing loss 1 month after birth, using ABR. The two-stage measurement of AABR is effective and time efficient due to significant decreases in the referral rate and the false-positive rate.