外源性谷氨酸引发的内耳毛细胞损伤与支持细胞GLAST表达的相关性研究

Relationship Between the Damage to IHC Initiated by Extraneous Glu and the Reaction of the Supporting Cell GLAST

目的观察外源性谷氨酸引发的内耳毛细胞损伤与支持细胞谷氨酸转运体(glutamate-aspartate trans-porter,GLAST)表达的相关性。方法将30只成年健康豚鼠随机分为3组,每组10只,经耳蜗钻孔,第一组动物左耳为对照组,给予人工外淋巴液(对照组);第二组动物左耳给予20mmol/L谷氨酸(Glu组);第三组动物为正常组,不作特殊处理。采用听性脑干反应(ABR)测试、耳蜗铺片、硝酸银染色及免疫组织化学技术,观察Glu处理前后的听力学、形态学改变,以及引发的支持细胞GLAST的表达变化。结果给药后,三组豚鼠1kHz ABR平均阈值分别为8.00±2.68、18.00±3.50、8.00±2.58dB nHL;8kHz分别为10.00±3.33、27.00±4.22、7.50±2.64dB nHL;Glu组与对照组有显著差异(P<0.05)。形态学改变:给予20mmol/L谷氨酸3d后,外毛细胞有较明显改变,出现错乱、缺失,内毛细胞出现形态改变,与听力学变化一致。免疫组织化学:正常组GLAST在支持细胞有表达,给予Glu后GLAST强阳性表达。结论谷氨酸对豚鼠内耳毛细胞具有毒性作用,GLAST参与外源性谷氨酸的转运,并受细胞外谷氨酸浓度的调控。

Objective To observe the relationship between the damage to IHC and the reaction of the supporting cell GLAST after giving extraneous Glu. Methods 30 adult guinea pigs were divided randomly into three groups and given different socution respectively through the holes drilled into the scala tympani of cochlea. In 10 animals, the left ears were taken as the control group in one group. In other 10 animals,the left ears were taken as treatment group given Glu （20 mmol/L）.The third group was the normal group without any solution. The changes in audiology and morphology at different periods before and after giving the drug were observed by auditory brainstem response（ABR）, cochlear preparation, silver nitrate dying and immuniohistochemistry. Results After giving the drug to the operated ears, the average threshold levels of ABR at 1 kHz were（ ± s） 8.00 ± 2.68 dB nHL, 18.00±3.50 dB nHL,8.00±2.58 dB nHL respectively, and at 8 kHz were 10.00±3.33 dB nHL, 27.00±4,22 dB nHL,7.50 ± 2.64 dB nHL respectively（ P 〈 0.05）. The sharp differences were observed when comparing with the control ears. When cochlear scala tympani was injected with Glu of 20 mmol/L concentration, there were obvious changes in morphology and absence in the OHCs, and the similar morphological changes occured in IHCs, which corresponded with the changes of audiology. Immunohistochemistry demonstrated that in supporting cell, the normal group showed the GLAST positive expression. Whereas in the group with the extraneous Glu givien, there was the stronger GLAST positive expression. Conclusion Glu probably has a toxic effect on inner ear hair cells, and participates in extraneous Glu transportion, prepared by extraneous Glu.