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; 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，我们研究了一种新技术，用于使用体内微观分析在周期性中对谷氨酸进行定量测量。由GDH和NAD的反应产生的NADH用作谷氨酸浓度的指标。但是，在这种方法中，我们无法检测到氨基糖苷耳毒性诱导的谷氨酸浓度的变化。将计划进一步的研究以提高该技术的敏感性。此外，我们测量了甲基胆碱（ACH）在PerilyMPH中的浓度，以找出耳蜗有效神经系统对耳鸣的产生的影响。在人工耳蜗有效的神经结尾中，ACH被认为是主要的神经递质。为了衡量围绕PerilyMPH的ACH释放，我们使用了体外微透析。样品的ACH水平由HPLC-ECD用酶柱确定，将乙酰胆碱酯酶和氧化胆碱固定在其上。在50μmAMPA（非NMDA谷氨酸受体激动剂）之前和之后测量了ACH水平的变化，将其添加到灌注液中，而不是第四通气底的电刺激。当林格溶液含有10 m Physostigmin时，基底ACH水平为22±4 Pico mole/ml（平均值±SD）。给予50μmAMPA后，ACH的水平立即增加了2-3倍（n = 4）。基础ACH水平与ACH样活性一致，而在先前的生物测定报告中没有电刺激。这些结果表明，微透析可用于监测PerilyMPH中的ACH释放。此外，初步数据表明，诸如AMPA给药之类的传入刺激可能会导致ACH从有效的神经端释放。将需要进一步研究以找出AMPA诱导的ACH释放与耳鸣的产生之间的关系。