THE ROLE OF GLUTAMATE ON THE GENERATION OF TINNITUS

谷氨酸对耳鸣产生的作用

• 批准号: 11671701
• 负责人: OGAWA Kaoru
• 金额: $ 0.83万
• 依托单位: KEIO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671701/
• 关键词: TINNITUS; MECHANISM; GLUTAMATE; ACECHYLCHOLONE; MICRODIALYSIS

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.

在本研究中，我们研究了一种新的技术，定量测量谷氨酸在外淋巴液中使用在体内微透析。通过GDH和NAD的反应产生的NADH被用作谷氨酸浓度的指示剂。然而，在这种方法中，我们不能检测到氨基糖苷类耳毒性引起的谷氨酸浓度的变化。此外，我们还测定了耳蜗外淋巴液中乙酰胆碱（Ach）的含量，以了解耳蜗传出神经系统在耳鸣发生中的作用。乙酰胆碱被认为是耳蜗传出神经末梢的主要神经递质。采用体外微透析法测定外淋巴液中Ach的释放。用固定化乙酰胆碱酯酶和胆碱氧化酶的酶柱，用HPLC-ECD测定样品中乙酰胆碱的含量。用50 μM AMPA（一种非NMDA谷氨酸受体激动剂）代替电刺激第四脑室底，测定灌流液中Ach含量的变化。当林格氏液中含有10 M毒扁豆碱时，基础Ach水平为22±4皮科mole/ml（平均值±SD）。注射50 μM AMPA（n=4）后，Ach水平立即升高2-3倍。基础Ach水平与先前生物测定报告中无电刺激时的Ach样活性一致。提示微透析技术可用于外淋巴液Ach释放的监测。此外，初步的数据表明，传入刺激，如AMPA的管理，可能会导致Ach从传出神经末梢的释放。AMPA引起Ach释放与耳鸣发生的关系有待进一步研究。