OTOTOXIC MECHANISMS OF GENTAMICIN ON SENSORY HAIR CELLS

这项建议旨在解决一个主要问题， 对鱼耳感觉毛细胞的选择性耳毒性损伤 一种氨基糖苷类抗生素，硫酸庆大霉素。 最近我们 结果表明，只有椭圆囊和拉格纳的纹状毛细胞 鱼耳的耳石内器官对硫酸庆大霉素敏感 人身攻击 耳蜗外毛的选择性损伤 前庭终器的I型毛细胞和I型毛细胞也被认为是 在哺乳动物物种中发现。 这些相同的损伤模式 鱼类和哺乳动物证明鱼耳作为一个有价值的模式系统， 建议的研究。 提出三个假说，并将用于阐明机制 导致庆大霉素对感觉毛细胞造成选择性损伤。 1)“受体存在“假说预测了 两种毛细胞之间的庆大霉素受体。 将 通过定位庆大霉素受体，即， 磷酸肌醇（PIP2）。 免疫金透射电子 用抗受体的抗血清进行的免疫细胞化学方法 将被用来检测受体之间的分布纹状体和 纹外毛细胞2)“路径路径”假说预测空间 两者之间庆大霉素摄取途径的时间差异 细胞类型。 将使用放射自显影术和 免疫细胞化学方法。 氚化庆大霉素将用于 在光下观察庆大霉素的细胞内存在 显微镜 抗庆大霉素的抗血清将用于 结合免疫金检测细胞内靶点 透射电镜下观察庆大霉素。3)“锂堵塞” 一种假说预测，如果庆大霉素受体可以被阻断， 应观察耳毒性损害。 该假设将通过以下方式进行检验： 使用锂，一种已知的PIP2抑制剂，通过全身注射， 直接应用于耳石终末器官， 注射庆大霉素 “锂堵塞”假说可能得到证实 如果在纹状体毛细胞处没有检测到耳毒性损伤。

This proposal is intended at addressing a major question with regard to selective ototoxic damage to sensory hair cells of the fish ear caused by one aminoglycoside antibiotic, gentamicin sulfate. Recently we demonstrated that only striolar hair cells in the utricular and lagenar otolithic endorgans of fish ear are susceptible to gentamicin sulfate assault. The same type of selective damage to the cochlear outer hair cells and type I hair cells of vestibular endorgans have also been reported in mammalian species. These identical damage patterns between fish and mammals justify the fish ear as a valuable model system for the proposed study. Three hypotheses are proposed and will be used to elucidate mechanisms leading to selective damage to sensory hair cells caused by gentamicin. 1) "Receptor Presence " hypothesis predicts differential distribution of gentamicin receptors between the two types of hair cells. It will be tested by localizing the presence of gentamicin receptors, i.e., phosphoinositide (PIP2). Immuno-gold transmission electron immunocytochemical procedures with antisera raised against the receptor will be used to detect the distribution of receptors between striolar and extrastriolar hair cells. 2) "Route Pathways" hypothesis predicts spatial and temporal difference in uptake routes of gentamicin between the two cell types. The hypothesis will be tested by using both autoradiography and immunocytochemistry methods. Tritiated gentamicin will be used to visualize the intracellular presence of gentamicin under light microscopy. Antisera raised against gentamicin will be used in conjunction with immuno-gold to examine intracellular targets of gentamicin under transmission electron microscope. 3) "Lithium blockage" hypothesis predicts that if gentamicin receptors could be blocked then no ototoxic damage should be observed. The hypothesis will be tested by using lithium, a known inhibitor of PIP2, through systemic injection and direct application into otolithic endorgans in conjunction with injections of gentamicin. "Lithium blockage" hypothesis could be proved if no ototoxic damage is detected at striolar hair cells.