Membrane current possessing the properties of a mechano-electric transducer current in inner hair cells of guinea-pig cochlea.

We measured the membrane current possessing the properties of a mechano-electric transducer current in isolated inner hair cells of guinea-pig cochlea. In a free-standing hair bundle, depolarization to +80 mV evoked a stable outward current attributable to the opening of transducer channels, and repolarization to -80 mV evoked a transient inward current indicating adaptation. The time constant of adaptation increased as the membrane potential depolarized. Dihydrostreptomycin diminished both the outward and inward currents.

[Bilateral primary carcinoma of the external auditory meatus].

We report an extremely rare case of bilateral primary carcinoma of the external auditory meatus. A 50-year-old man suffered 1 month from left-ear discharge and otalgia. Examination revealed a tumor of the left ear canal and a biopsy showed well-differentiated squamous cell carcinoma. The patient underwent 60 Gy radiotherapy and left subtotal temporal bone resection. A tumor with irregular swelling of the right external canal was found 8 months after the first diagnosis. Biopsy of the right external canal confirmed the same squamous cell carcinoma. Genetic examination that the carcinoma of the right ear was probably not a metastatic from the carcinoma of the left ear.

Development of monovalent ions in the endolymph in mouse cochlea.

The present study was designed to clarify the chronological developmental process of monovalent ions (Na(+), K(+), Cl(-)) in the endolymph of the mouse in relation to the development of the endocochlear potential (EP). The EP and ionic concentrations were measured simultaneously with the ion-sensitive double-barreled microelectrodes from the scala media of the basal turn. The EP increased abruptly 7 days after birth (DAB) and reached approximately 80 mV 14 DAB. In the earliest postnatal days, the endolymphatic Na(+) concentration was significantly higher than that in adult mice, however, the K(+) and the Cl(-) concentrations were lower. The concentrations of all the monovalent ions in endolymph reached adult levels at 7 DAB when the EP was still under 20 mV. These data strongly suggest the presence of a different mechanism between the production of monovalent ions, especially of high K(+) in the endolymph and that of EP.

Induction of mucin gene expression in middle ear of rats by tumor necrosis factor-alpha: potential cause for mucoid otitis media.

Mucoid otitis media (MOM) is characterized by viscous fluid, high in mucin concentration, which accumulates in the middle ear cavity. Recent studies suggest that initial infection in the middle ear cleft may be key to the development of MOM. However, factors of the initial infection attributed to the stimulation of mucin production are not clearly understood. This study demonstrated that tumor necrosis factor (TNF)-alpha, a proinflammatory cytokine in mucoid effusion, markedly increased Muc2 mucin mRNA expression in middle ear epithelium, in a time- and dose-dependent manner. Parallel to this was a marked increase in mucin glycoprotein in middle ear fluid. Also, TNF-alpha demonstrated an autocrine and/or paracrine effect on the expression of endogenous TNF-alpha gene in the middle ear, which may contribute to the production of mucin in this study. These findings suggest that TNF-alpha plays an important role in the development of MOM by stimulating mucin metabolism.

A role of glutamate in drug-induced ototoxicity: in vivo microdialysis study combined with on-line enzyme fluorometric detection of glutamate in the guinea pig cochlea.

The time course of the changes in perilymphatic glutamate was determined during the application of kanamycin and ethacrynic acid, which are known to damage the hair cells in the inner ear. For the continuous recording of glutamate, the microdialysis technique combined with an enzyme-linked fluorometric assay was used. In guinea pigs receiving a loading dose of 800 mg/kg of kanamycin subcutaneously followed 3 h later by an i.v. injection of 40 mg/kg of ethacrynic acid, a marked glutamate release was clearly found about 2 h after the injection of ethacrynic acid. Injection of kanamycin or ethacrynic acid alone did not produce any change in the perilymphatic glutamate. The morphological changes induced by the administration of both drugs indicated that the collapsing hair cells might release glutamate into the perilymphatic space. The present findings provide additional evidence that glutamate acts as an aggravating factor in aminoglycoside-induced ototoxicity.

Management of cholesteatoma in the petrous apex.

Two cases of congenital and acquired cholesteatoma in the petrous apex were operated on via the translabyriathine approach. The cholesteatoma was removed, leaving a part of the matrix membrane in both cases as the matrix membrane was difficult to remove. Spaces including the mastoid and middle ear after removal of cholesteatoma were obliterated with only fibrin glue in order to pneumatize the operated space after surgery. Cerebrospinal fluid (CSF) leakage from the internal auditory meatus was prevented using a tiny fascia. The spaces occupied by the cholesteatoma were clearly pneumatized within 6 months after surgery in both patients. The advantage of creating pneumatized spaces after removal of cholesteatoms in the petrous apex was discussed.

Increase in glutamate-aspartate transporter (GLAST) mRNA during kanamycin-induced cochlear insult in rats.

Kanamycin (KM)-induced changes in expression of the gene for glutamate-aspartate transporter (GLAST) in the rat cochlea were analyzed by Northern blotting. With the administration of KM (600 mg/kg/day) once daily for 20 days, the expression of GLAST mRNA gradually increased and reached a peak on day 20. Although the expression of GLAST mRNA remained at a high level until 12 days after the completion of the KM treatment, it then fell to the normal level within 2 months. Such KM treatment resulted in loss of both inner and outer hair cells and a concomitant profound permanent threshold shift. The present findings suggest that during KM administration, high concentrations of extracellular glutamate released by collapsing hair cells induced GLAST mRNA expression. Increased GLAST mRNA might play an important role in the prevention of the secondary death of spiral ganglion neurons from glutamate neurotoxicity.

Pneumococcus activation of the 5-lipoxygenase pathway and production of glycoproteins in the middle ear of rats.

Pneumococcal otitis media is associated with the production of potent inflammatory mediators (leukotrienes), but the mechanism by which pneumococcus induces production of leukotrienes in the middle ear is poorly understood. In this study, up-regulation of 2 genes that govern the lipoxygenase pathway, cPLA2 and 5-LOX, was observed in rats following inoculation of pneumococcus into the middle ear cavity. Expression of cPLA2 was low, and 5-LOX gene expression was not detected in control animals. Up-regulation of cPLA2 and 5-LOX in middle ear epithelial cells was accompanied by an increase of high-molecular-weight glycoproteins in middle ear fluid and cells. These findings suggest that pneumococcus activates the lipoxygenase pathway by up-regulating expression of the cPLA2 and 5-LOX genes. This, in turn, may stimulate synthesis and secretion of high-molecular-weight glycoproteins that facilitate production of fluid in the middle ear cleft.

High potassium-induced glutamate release in the cochlea: in vivo microdialysis study combined with on-line enzyme fluorometric detection of glutamate.

The time course of changes in perilymphatic glutamate release and their Ca2+-dependency were studied in the guinea pig cochlea during high K+-evoked depolarization. The glutamate concentration was analyzed continuously by an enzyme-linked fluorometric assay combined with microdialysis. Two peaks of glutamate increase were found in response to perfusion for 10 min. In the absence of Ca2+, the first peak was diminished, whereas the inhibition of the second peak was minimal.

Effects of trimetaphan-induced deliberate hypotension on human cochlear blood flow.

In order to observe the reaction of cochlear blood flow (CBF) to trimetaphan (TMP)-induced hypotension, CBF was measured with laser-Doppler flowmetry in 7 human subjects during general anaesthesia for middle ear surgery. All subjects showed a decrease in mean arterial pressure (MAP) during intravenous infusion of TMP, followed by a gradual return to the baseline level after termination of the infusion. The CBF generally followed the MAP changes with the same pattern. Three of the seven subjects demonstrated a CBF change larger than the maximum MAP change, indicating the lack of a local autoregulatory mechanism in CBF. On the other hand, CBF changes were smaller in magnitude than the maximum change in MAP for the rest of the subjects, suggesting an autoregulatory mechanism in CBF. However, since the audiograms from these subjects indicated profound damage along the cochlear basal turn probably due to middle ear inflammation, concomitant vascular damage in this region offers another possible explanation for the inappropriate CBF changes. The present observations may also suggest that deliberately TMP-induced hypotension has a potentially harmful effect on CBF during otological surgery that attempts to preserve or improve hearing.

Changes of perilymphatic glutamate and cochlear blood flow following ischemia.

Dynamic changes of perilymphatic glutamate and cochlear blood flow were measured simultaneously in the guinea pig following cochlear ischemia. Glutamate was measured by the microdialysis technique with a probe inserted into the scala tympani at the basal turn. Cochlear blood flow was monitored with a laser-Doppler probe on the lateral wall of the second cochlear turn. Both parameters were measured before and after electrocauterization of the anterior inferior cerebellar artery and other vessels supplying the internal auditory canal. In four animals, glutamate increased with a decline of cochlear blood flow and then decreased with recovery of blood flow. No normalization of glutamate was observed in seven animals with a persistent decrease of blood flow. The results of this study indicate that the glutamate regulating system in the cochlea is dependent on cochlear blood flow.

Quantitative immunogold cytochemistry reveals sources of glutamate release in inner ear ischemia.

Glutamate is thought to be a major neurotransmitter between hair cells and afferent dendrites in the inner ear. However, excessive glutamate is known to be excitotoxic, and may be involved in ischemic neuronal damage in the central nervous system. The glutamate concentration in the perilymph has been reported to increase during ischemia, but the source of glutamate is still unclear. In the present study, we have used post-embedding immunogold cytochemistry to analyse changes in the cellular distribution of glutamate in the guinea pig organ of Corti during ischemia. The areal gold particle densities in the inner hair cells of the ischemic side were lower than those of the control side, indicating that glutamate may be released from the hair cells during ischemia. Adjacent supporting cells (border cells) also showed a decrease in particle density, suggesting that they constitute an additional source of glutamate.

Mode of interaction of loop diuretics with human serum albumin and characterization of binding site.

PURPOSE: The purpose of this study was to investigate the binding mechanism of loop diuretics with HSA and to characterize the binding site on HSA. METHODS: Quantitative analysis of potential interaction between ligands bound to HSA was performed by equilibrium dialysis and data for binding of the two ligands to HSA were analyzed on the basis of a theoretical model of simultaneous binding of two ligands. RESULTS: The binding of loop diuretics is dependent upon the N-B transition, conformational change of albumin. Furthermore, from the results of binding of the drugs to modified HSA, the lysine residue seems to be involved in the binding of loop diuretics to HSA. CONCLUSIONS: Analysis using models describing independent, competitive, cooperative and anti-cooperative binding led to the conclusion that loop diuretics bind to site I, particularly to the warfarin region on HSA.

Effects of experimental round window membrane laceration on the free amino acid profile of perilymph.

To evaluate biochemical changes of inner ear fluid following perilymphatic fistula (PLF), free amino acid (FAA) profiles of perilymph in experimental PLF were determined using high performance liquid chromatography (HPLC). Thirty-five guinea pigs were anesthetized and prepared as PLF models by perforating the round window membrane (RWM) of the left ear. Right ears served as controls. Samples (2 microliters) were aspirated from scala tympani through a RWM perforation. Animals were divided into two groups according to time of sampling following PLF induction: 2-week group (n = 17) and 4-week group (n = 18). Compound action potential (CAP) evoked by 1, 2, 4 and 8 kHz tone bursts were measured using a round window electrode from the left ear before PLF induction and from both ears before final sampling. RWM perforations were completely closed at the time of the final sampling in 8 of 17 animals from the 2-week group, and 15 of 18 animals from the 4-week group. In comparison with that in control ears, concentrations of FAA throughout all profiles was dramatically elevated in the PLF ears with a healed RWM perforation. Most PLF ears with persistent RWM perforation showed minimal differences between 2-week and 4-week groups. No remarkable CAP threshold changes were found at any frequencies tested following PLF induction in both the 2-week and 4-week groups. The unchanged 8 kHz threshold suggests that FAA concentration increases only at the basal end of the cochlea. FAAs accumulate within the basal end of scala tympani in ears with a healed RWM.(ABSTRACT TRUNCATED AT 250 WORDS)

[A study of preoperative administration of 5'-DFUR in patients with colo-rectal cancer].

For the purpose to assess tumor-selectivity of doxifluridine (5'-DFUR), an anticancer drug of fluorinated pyrimidine class, the authors conducted a clinical-pharmacological study with the drug. The subjects in the present study, 26 patients with colorectal cancer received surgeries, were preoperatively administered 5'-DFUR orally at either doses of 800 or 1,200 mg/day for 3 days followed by oral administration of the drug at a dose of 400 mg 4-6 hrs before operation. The tissues were collected from specimens removed at operations and we measured both the activities of pyrimidine nucleoside phosphorylase (PyNPase), an activating enzyme for 5'-DFUR, and 5-FU concentrations in the tissue samples. The PyNPase activities showed higher levels in tumor tissues and regional lymph nodes than in normal tissues adjacent to the tumors. The 5-FU concentrations were: 102.7 ng/g in tumor tissues, 12.5 ng/g in normal tissues adjacent to the tumors, 97.6 ng/g in metastatic lymph nodes, and 7.3 ng/g in normal lymph nodes. In other words, the 5-FU concentrations were significantly (p < 0.01) higher in either of tumor tissues and metastatic lymph nodes than in the rests. The 5-FU concentrations in the sera were also extremely low. The results above clearly indicate that 5'-DFUR has a high selectivity for tumors.

Conjugation of Cu,Zn-superoxide dismutase with succinylated gelatin: pharmacological activity and cell-lubricating function.

Superoxide dismutase (SOD) and succinylated gelatin (succinyl gelatin) were conjugated to improve in vivo pharmacological activity of SOD. Lysyl residues of human recombinant Cu,Zn-SOD were cross-linked with carboxyl residues of succinyl gelatin using 1-ethyl-3-[3- (dimethylamino)propyl]carbodiimide. Various chemical and pharmacokinetic parameters of the conjugate were determined. Analysis by atomic absorption spectrometry and amino acid composition revealed that the conjugate was composed of about 2.9 mol of succinyl gelatin (with a mean molecular weight of 23,000) to 1 mol of SOD and exhibited an apparent mean molecular weight of 98,000. The conjugate retained almost 100% of its original activity on a molar basis. When the succinyl gelatin-conjugated Cu,Zn-SOD (Suc-gel-SOD) was administered intravenously to mice, its plasma half-life was prolonged to 29.7 min compared with 4.5 min for native SOD. Tissue distribution analysis revealed that intravenously administered Suc-gel-SOD showed a much greater accumulation than native SOD in the liver followed by in decreasing order the kidney, the lung, and the spleen; native SOD was excreted more rapidly into urine before it accumulated in tissues. Furthermore, Suc-gel-SOD exhibited lower antigenicity and immunogenicity than native SOD, and it had a better therapeutic effect against ischemic edema of the foot pad in mice. The conjugate was found to accumulate more than native SOD in the ischemic foot pad. A newly added property of the conjugate is cell-lubricating activity, which facilitated cell passage through micropores and reduced hemolysis during cell passage in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)