OTOTOXIC MECHANISMS OF GENTAMICIN ON SENSORY HAIR CELLS

这项提议旨在解决与以下方面有关的一个主要问题 沙门氏菌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 证明只有椭圆形和背侧的纹状毛细胞 鱼耳耳石内脏对硫酸庆大霉素敏感 人身攻击罪。同样类型的选择性耳蜗外毛发损伤 前庭内脏的细胞和I型毛细胞也被 在哺乳动物物种中有报道。这些相同的损坏模式 鱼和哺乳动物证明鱼耳朵是一种有价值的模型系统 建议的研究。 提出了三个假说，并将用来阐明机制 导致庆大霉素对感觉毛细胞的选择性损伤。 1)“受体存在”假说预测不同的分布 庆大霉素受体在两种类型的毛细胞之间。会是 通过定位庆大霉素受体的存在进行测试，即， 肌醇磷脂(PIP2)。免疫金穿透电子 用抗受体抗血清进行免疫细胞化学反应 将用于检测纹状体和纹状体之间受体的分布 纹状体外毛细胞。2)“路径路径”假说预测空间 以及庆大霉素在两者摄取途径上的时间差异 单元类型。这一假设将通过使用两种放射自显影进行验证 和免疫细胞化学方法。氚化庆大霉素将用于 光下细胞内庆大霉素存在的可视化 显微镜。针对庆大霉素的抗血清将用于 与免疫胶体金联用检测细胞内靶点 在透射电子显微镜下观察庆大霉素。3)“锂堵塞” 假说预测，如果庆大霉素受体可以被阻断，那么 应观察到耳毒性损害。这一假设将通过以下方式进行检验 使用已知的PIP2抑制剂锂，通过全身注射和 直接应用于耳石内脏联合 注射庆大霉素。“锂阻塞”假说可以得到证实 如果纹状毛细胞没有检测到耳毒性损害。

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.