Concerted action of the high affinity calcium binding sites in skeletal muscle troponin C.

Mutants of each of the four divalent cation binding sites of chicken skeletal muscle troponin C (TnC) were constructed using site-directed mutagenesis to convert Asp to Ala at the first coordinating position in each site. With a view to evaluating the importance of site-site interactions both within and between the N- and C-terminal domains, in this study the mutants are examined for their ability to associate with other components of the troponin-tropomyosin regulatory complex and to regulate thin filaments. The functional effects of each mutation in reconstitution assays are largely confined to the domain in which it occurs, where the unmutated site is unable to compensate for the defect. Thus the mutants of sites I and II bind to the regulatory complex but are impaired in ability to regulate tension and actomyosin ATPase activity, whereas the mutants of sites III and IV regulate activity but are unable to remain bound to thin filaments unless Ca2+ is present. When all four sites are intact, free Mg2+ causes a 50-60-fold increase in TnC's affinity for the other components of the regulatory complex, allowing it to attach firmly to thin filaments. Calcium can replace Mg2+ at a concentration ratio of 1:5000, and at this ratio the Ca2.TnC complex is more tightly bound to the filaments than the Mg2.TnC form. In the C-terminal mutants, higher concentrations of Ca2+ (above tension threshold) are required to effect this transformation than in the recombinant wild-type protein, suggesting that the mutants reveal an attachment mediated by Ca2+ in the N-domain sites.

[Neo-adjuvant chemotherapy with cisplatin, 5-fluorouracil, vindesine in head and neck cancer].

Neo-adjuvant chemotherapy with CDDP combination was introduced into the treatment of advanced head and neck cancer. Twenty-three patients with s.c.c of head and neck were given combination chemotherapy consisting of CDDP, VDS and 5-FU before surgical treatment in our department. CR and PR of this trial in all patients were 4 and 35%, respectively. The WBC nadir occurred around 2 weeks later, but all the patients recovered prior to the next cycle or surgical treatment. Renal dysfunction, nausea, vomiting and depilation were generally mild. VDS is useful as one of the neo-adjuvant drugs for the treatment of head and neck cancer. Long-term observation in connection with this treatment is required.

[Use of a butterfly ventilation tube in the treatment of otitis media with effusion].

Sixty-three ears of 49 patients with serous otitis media were treated making use of butterfly ventilation tube in our 4 hospitals and 1 private office. The average time from intubation to extubation was 9 months and the longest case was 33 months. Main complications were infection and spontaneous extubation. The rate of hearing improvement after tympanostomy was more than 80% in all cases. The frequency of the most improvement was observed in 1kHz on the average. It was cleared that the butterfly ventilation tube was easy to use for the wide age patients and at any clinics. It was concluded that the butterfly ventilation tube was useful as a long-term ventilation tube.

Interaction of polyanion against kanamycin ototoxicity.

We studied whether interference of the electron bindings between kanamycin (KM) and the outer plasma membranes of the hair cells with polyanion such as heparin can reduce ototoxicity. In a short course experiment, KM 200 mg/kg/day was injected intramuscularly 23 times with or without 1 U or 0.5 U of heparin/g/day differently to 22 guinea pigs. The reducing effect of heparin against KM ototoxicity was significant from a comparison of the N1 threshold obtained by the cochleogram. The number of surviving outer hair cells in the heparin groups was significantly greater in the third turn than in the group given KM alone. A long course experiment in which 50 injections of KM with or without 0.5 U of heparin were given intramuscularly to 16 guinea pigs revealed from the pinna reflex, cochlear microphonics and cell counting that heparin could reduce ototoxicity slightly at an early stage or before reaching crucial accumulation.

Duane's syndrome associated with crocodile tear and ear malformation. A case of stapes anomaly and its reconstruction.

Duane's syndrome is a congenital eye movement disorder characterized by marked limitation or absence of abduction, variable limitation of adduction, and narrowing of the palpebral fissure with retraction of the globe on attempted adduction. We treated a patient with bilateral Duane's syndrome associated with crocodile tear and external, middle, and inner ear anomalies. Exploratory tympanotomy showed an absence of the oval window and a rudimentary superior structure of the stapes that was connected to the abbreviated incudal lenticular process with connective tissue. A small hole was drilled into the vestibulum and the ossicular chain was reconstructed by inserting a Teflon wire piston to minimize the air-bone gap. Pertinent literature is reviewed, and the pathogenesis of Duane's syndrome and the aspect of concomitant anomalies are discussed.

Localization of triphosphoinositide in the cochlea. An electronmicroscopic immunocytochemical study.

Triphosphoinositide (TPI) has been demonstrated to be a receptor for aminoglycosides in the cochlea and may regulate ionic permeability by its binding with Ca++. This phospholipid was localized by a protein A-gold technique in the cochlea at the electronmicroscopic level. TPI was prepared by a neomycin column and antibodies to it were raised in rabbits. The antibody used in this study reacted virtually only to TPI among the tested lipids. TPI was localized mainly at stereocilia, cuticular plates, head plates of Deiters' cells, plasma membrane, and mitochondria of various cells in the organ of Corti. In the vascular stria, TPI was found mainly at the plasma membrane of basal infoldings of the marginal cells. Possible physiological and pathophysiological roles of TPI in the cochlea are briefly discussed.

Responses of gerbil and guinea pig auditory nerve fibers to low-frequency sinusoids.

The characteristics of time-locked auditory nerve fiber responses to 50 Hz acoustic sinusoids were studied in gerbils and guinea pigs. Whereas the time-locked responses of all guinea pig fibers produced single-peaked period histograms, those of the gerbil produced distorted, multiple-peaked response histograms, especially fibers with characteristic frequencies (CFs) between 2 and 10 kHz. Although the shapes of the period histograms vary with stimulus intensity, the phases of the fundamental components are essentially invariant over the range of stimulus intensities used. In contrast to the phase of the cochlear microphonic produced by the 50 Hz stimulus, which was constant along the length of the cochlea in both species, the phase of the neural responses depends on the fiber CF in each of the two species. In guinea pigs, the phase of the neural responses relative to the acoustic stimulus decreases with the fiber CF from a phase lead of 90 degrees for fibers with CFs below 300 Hz to a phase lag of nearly 60 degrees for fibers with CFs greater than 3 kHz. In gerbils, the response phase also decreases with increasing CF below 2 kHz and above 10 kHz but undergoes an abrupt 160 degrees phase increase between those frequencies.

Quantitative analysis of kanamycin ototoxicosis.

The morphological changes after kanamycin intoxication of the inner ear, including both the cochlea and the vestibule, were quantitatively analysed by the surface preparation technique after succinic dehydrogenase staining. 75 guinea pigs were used. The outer hair cells in the basal coil and the inner hair cells in the upper coils of the cochlea were the most severely damaged, but many unusual modes of damage were also revealed. For example, the initial hair cell damage in the cochlea appeared in the upper hair cells. The clearly observed vestibular damage contradicts the general belief that kanamycin is not so toxic to the vestibular hair cells. The utricular macula and the lateral crista were most severely damaged. The delayed ototoxicity of kanamycin was observed for the first time in the vestibular hair cells.