Objective To investigate the effect of D-AP5 (D-2-amino-5-phosphonopentanoate, a specific NMDA-antagonist) on the increase of intracellular free Ca 2+ concentration ([Ca 2+ ] i) induced by glutamate in isolated cochlear inner hair cells (IHCs), and to detect the autoreceptors of the IHC membrane. Methods When a laser scanning confocal microscope (LSCM) was used, the exogenous glutamate (Glu)-induced changes in [Ca 2+ ] i of isolated IHCs and OHCs of guinea pig cochlea were observed with fluo-3, a fluorescent probe for [Ca 2+ ] i. After D-AP5 or CNQX (6--cyano--7--nitroguinoxaline--2, 3--dione, a specific AMPA- antagonist) was administrated, the exogenous glutamate (Glu)-induced changes in [Ca 2+ ] i of isolated IHCs were recorded. Results In the presence of a low concentration Glu (3.85?μmol/L), there was an increase of [Ca 2+ ] i in IHCs, whereas there was no change in OHCs. When 50?μmol/L of D-AP5 was administrated in advance, Glu did not induce a corresponding increase in [Ca 2+ ] i in IHCs, and 50?μmol/L of CNQX did not completely block the increase of [Ca 2+ ] i in IHCs. Conclusions These results suggest that the autoreceptors existing in the IHC membrane are mainly of NMDA type, while there are relatively few AMPA receptors. Exogenous Glu is capable of increasing [Ca 2+ ] i in IHCs by acting on the NMDA autoreceptor of IHCs in a positive feedback manner.
目的通过脉冲噪声分别对豚鼠进行30、15次的暴露,分析比较豚鼠的听力学变化,探索建立隐性听力损失模型的适合条件。同时给予氢气,探究其对隐性听力损失的预防作用。方法选取ABR听阈正常的豚鼠16只,随机分为4组:空白对照组、脉冲噪声30次组、脉冲噪声15次组、氢气吸入+脉冲噪声15次组。脉冲噪声压力峰值为163 dB SPL,脉宽为0.25ms,间隔时间为6.5s。分别于脉冲噪声暴露前及暴露后24h进行听性脑干反应测定。结果通过统计学分析发现,豚鼠在30次脉冲噪声暴露24h后,其ABR短声阈值及短纯音(16 kHz 70 dB SPL)Ⅰ波幅值产生明显改变,具有统计学意义;15次脉冲噪声暴露组豚鼠其各项听力学指标都发生显著改变;氢气预处理组同单纯脉冲噪声暴露15次组的暴露24h后听力学指标相比,其ABR短声阈值及短纯音(16 kHz 70 dB SPL)Ⅰ波幅值存在统计学差异。结论本研究中脉冲噪声暴露30次及15次均对豚鼠听力产生显著影响。其中脉冲噪声暴露15次的豚鼠,其各项听力学指标都符合隐性听力损失的听力学特点,脉冲噪声暴露15次是建立隐性听力损失动物模型的可行条件。此外,氢气防护组较单纯脉冲噪声暴露组,听力学指标存在显著差别,表明氢气对隐性听力损失具有预防作用,为进一步的分子机制研究提供直接实验依据。
目的观察豚鼠耳蜗脂肪酸转运蛋白1和4(fatty acid transport protein,FATP1,FATP 4)在噪声损伤和促脂肪酸代谢药物苯扎贝特干预前后的变化,探讨噪声性听力损失与FATP1、FATP4的关系。方法正常听力健康成年雄性Dunkin Hartley豚鼠21只随机分为对照组(15只)和干预组(6只),对照组在噪声暴露前随机选取3只(6耳)动物行耳蜗取材作为正常对照组,其余12只(24耳)与干预组(6只,12耳)一起予以白噪声(110 dB SPL,每天4小时,连续12天)暴露,干预组在噪声暴露的同时给予苯扎贝特(5 mg·kg-1·d-1)灌胃干预;在开始噪声暴露第6、12、24天行听性脑干反应(ABR)检测,采用脂肪酸特异性探针荧光标记技术检测脂类物质在豚鼠耳蜗的分布,采用免疫荧光染色技术、Western blot技术检测噪声暴露前后FATP1、PATP4在两组豚鼠耳蜗的表达变化。结果噪声暴露后对照组和干预组click声及4、8、16 kHz短纯音(tone burst)ABR(tb-ABR)反应阈较噪声暴露前显著升高,在噪声暴露的第12天,干预组4 kHz tb-ABR反应阈(16.25±5.82 dB SPL)明显低于对照组(32.27±5.92 dB SPL)(P<0.05);特异性脂肪酸探针荧光染色提示正常对照组豚鼠耳蜗脂类物质主要分布于基底膜外侧的Hensen细胞、螺旋神经节、血管纹,噪声暴露前免疫荧光染色显示FATP1主要表达于Corti器、螺旋唇,在螺旋神经节细胞、螺旋韧带、血管纹等少量表达;FATP4主要表达于耳蜗Corti器及螺旋韧带,在Corti器外侧的Hensen细胞、螺旋神经节、血管纹极少表达或不表达。噪声暴露后对照组(第12天)FATP1在Corti器、Hensen细胞、螺旋突、螺旋韧带的表达较噪声暴露前明显增加(P<0.01),对照组(第12天)和干预组(第24天)FATP4在Hensen细胞、螺旋唇、螺旋神经节的表达均较噪声暴露前明显增加(P<0.01),干预组(第24天)FATP4在血管纹的表达均较噪声暴露前增加(P<0.05),但对照组(第12天)FATP4在血管纹的表达均较噪声暴露前及�
