| Budget Amount *help |
¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2000: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Research Abstract |
In the present research, we investigated a new technique for the quantitative measurement of glutamate in the perilymph using in vivo microdyalysis. NADH produced by a reaction of GDH and NAD was used as a indicator of glutamate concentration. In this method, however, we could not detect a change of the glutamate concentration induced by aminoglycoside ototoxicity. Further study will be planned for improving the sensitivity of this technique.
In addition, we measured the concentration of acetylcholine (Ach) in the perilymph in order to figure out the effects of cochlear efferent nervous system on the generation of tinnitus. Ach has been considered as a major neurotransmitter in the cochlear efferent nerve endings. In order to measure Ach release in perilymph, we employed in vitro microdialysis. Ach level of the samples was determined by the HPLC-ECD with the enzyme column on which acetylcholine esterase and choline oxidase were immobilized. The change of Ach level were measured before and after 50 μM AMPA, a non-NMDA glutamate receptor agonist, was added to the perfusate instead of the electrical stimulation on the floor of 4th ventricle. The basal Ach level was 22±4 pico mole/ml (mean±SD), when the Ringer solution contained 10 M physostigmin. The level of Ach increased 2-3 folds immediately after 50 μM AMPA was administered (n=4). Basal Ach level was consistent with Ach-like activity without electrical stimulation on the previous report with bioassay. These results indicate that microdialysis is useful for monitoring Ach release in perilymph. In addition, preliminary data suggest that the afferent stimulation, such as the administration of AMPA, may induce the release of Ach from the efferent nerve endings. Further investigation will be needed to figure out the relationship between Ach release induced by AMPA and the generation of tinnitus.
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