顺铂对小鼠耳蜗螺旋神经节细胞延迟整流钾电流的影响

目的探讨耳毒性药物顺铂对耳蜗螺旋神经节细胞延迟整流钾电流的影响,了解顺铂耳毒性的离子机制。方法选择状态良好培养2—14d的耳蜗螺旋神经节细胞在电压钳制条件下记录的钾电流作为对照组,浴液中分别加入100,500,1000,5000μM不同浓度的顺铂溶液后记录的电流为实验组,观察其对钾通道电流电生理学特性的影响,并观察冲洗后电流的恢复情况。结果实验显示耳蜗螺旋神经节细胞钾通道电流的电生理学特性受不同浓度顺铂溶液的影响,并具有浓度依赖性,表现在其活化动力学和电流幅度的改变。本实验中顺铂的50％的电流被抑制时的浓度（IC50）为294μM,冲洗后钾通道电流逐渐恢复。结论顺铂可以改变耳蜗螺旋神经节细胞钾通道电流的电生理学特性,这种变化在短期内可逆,揭示其耳毒性的形成具有一定的离子机制。     

Preferential amino acid uptake identifies Type II spiral ganglion neurons in the gerbil

The localization of 3H-labeled amino acids was compared by light and electron microscopic autoradiography in the spiral ganglion of the gerbil, following in vivo intracochlear incubations. Following incubations with taurine, a population of heavily labeled neurons could be distinguished from lightly labeled neurons. The heavily labeled population comprised 5-6% of spiral ganglion neurons, and included the least myelinated cells. Ultrastructurally, the heavily labeled neurons were characterized by a loosely coiled Schwann cell sheath covering the cell body, and the presence of abundant cytoplasmic microfilaments. The unlabeled cells showed a typical perikaryal myelin sheath and cytoplasm rich in rough endoplasmic reticulum. It is concluded that the spiral ganglion of the gerbil contains both Type I and Type II neurons, and that Type II neurons preferentially incorporate the amino acid taurine. Type II neurons are therefore biochemically as well as morphologically distinct from Type I neurons.     

小鼠耳蜗螺旋神经节细胞电生理学特性的研究

目的：应用膜片钳技术记录小鼠耳蜗螺旋神经节细胞的全细胞电流,了解电压依赖性离子通道的基本电生理学特性,并比较耳蜗顶、底转螺旋神经节细胞电生理学特性的差异。方法：应用全细胞构型电压钳制技术,采用不同的电极内液及阻断剂,在不同的刺激参数下记录耳蜗顶、底转螺旋神经节细胞的电压依赖性离子通道电流,并进行分析比较。结果：实验记录到了内向的钠电流、延迟整流钾电流、超极化激活内向阳离子通道电流及瞬时外向钾电流,并发现耳蜗顶、底转螺旋神经节细胞的延迟整流钾电流及瞬时外向钾电流的电生理学特性具有显著性差异（P〈0．05）。结论：实验记录到的各种离子电流数据表明耳蜗螺旋神经节细胞具有完成动作电位的形成、传导并对其功能进行调节的离子通道基础;耳蜗顶、底转螺旋神经节细胞电生理学特性的差异有助于听觉的形成过程。     

Unbiased quantification of the microdissected human Scarpa's ganglion neurons

OBJECTIVE: The objective of the present study was to obtain unbiased estimates of the total number of Scarpa's vestibular ganglion neurons in individuals with normal vestibular function. STUDY DESIGN: Application of unbiased stereology using microdissected human temporal bone specimens. METHODS: Postmortem temporal bones were obtained from five young subjects with no history of audiovestibular disease (age range 42-49 years). The vestibular nerve containing the Scarpa's ganglion was microdissected, embedded in paraffin, and cut into 40 microm serial sections. Unbiased estimates of the total number of neurons were obtained using the optical fractionator technique of stereology. RESULTS: An average of 23,599 (coefficient of variation [CV] = 0.11) vestibular ganglion neurons was obtained. There was no significant difference between the results obtained from the microdissected specimens and results that had been obtained from an age-matched group derived from a previously published report from our laboratory using archival human temporal bone specimens. CONCLUSIONS: This study represents the first report to demonstrate the combination of the microdissection technique and the unbiased stereologic technique in the human temporal bone. This study demonstrates the reliability of the microdissection technique as an alternative method of human temporal bone processing for unbiased stereology. The utility of the microdissection technique is that specimens can be used for quantification, immunohistochemistry, and other powerful applications.     

顺铂对大鼠耳蜗螺旋神经节细胞外向延迟整流钾电流的影响

目的:观察记录顺铂作用下急性分离新生大鼠耳蜗螺旋神经节细胞(SGNs)延迟整流钾通道的电流曲线,分析顺铂对SGNs钾电流激活动力学的影响,并初步探讨其耳毒性机制。方法:采用全细胞膜片钳技术记录SGNs外向延迟整流钾电流及顺铂对此电流的影响。结果:钳制电压为-60mV,刺激电压从-60mV到 80mV逐渐去极化,阶跃电压为10mV,持续时间为500ms,可在SGNs上记录到外向钾电流,该电流对TEA-Cl(4-乙基胺)敏感,具有延迟整流特性;在细胞外液中加入10μmol/L顺铂,能明显抑制SGNs延迟整流钾电流;顺铂对此电流的抑制作用与细胞外液中顺铂的浓度呈剂量依赖性;外液洗脱后SGNs电流可基本恢复正常。结论:钾通道与SGNs动作电位的产生密切相关,顺铂可抑制SGNs钾通道电流,导致听觉功能障碍。     

Differential expression of voltage-gated potassium channel genes in auditory nuclei of the mouse brainstem

Voltage-gated potassium (Kv) channels may play an important role in the encoding of auditory information. Towards understanding the roles of Shaker and Shaw-like channels in this process, we examine here the expression of Kv1.1, Kv1.2, Kv3.1, and Kv3.3 in the central auditory nuclei of the mouse using quantitative in situ hybridization techniques. We establish rank order for each channel’s expression in each region, finding that the medial nucleus of the trapezoid body shows the highest signal for each of the four channel genes. In other auditory nuclei differential expression is found among and between members of both Shaker and Shaw subfamilies. Of particular interest is the stark contrast between high level expression of Kv1.1 and very low level expression of Kv3.1 in the octopus cell area of the cochlear nucleus and in the lateral superior olivary nucleus. These unique expression patterns suggest that Kv channel gene expression is regulated to allow brainstem auditory neurons to transmit temporally patterned signals with high fidelity. In instances where specific cell types can be tentatively identified, we discuss the possible contribution made by these channel genes to the physiological properties of those neurons.     

Quisqualate excites spiral ganglion neurons of the guinea pig

The effects of quisqualate on primary afferent auditory neurons were examined by comparing the unit activity of guinea pig spiral ganglion neurons before, during, and after the infusion of artificial perilymph containing 1 mM quisqualate into the basal turn scala tympani. In 10 of 13 preparations quisqualate infusion increased unit activity above baseline rates established prior to infusion while control infusions of quisqualate-free artificial perilymph had no appreciable influence on the unit activity of four preparations. Postexcitatory depression typically followed peak evoked excitation, but no purely inhibitory responses were observed. Postexcitatory depression developed into a temporary cessation of spike production in three cases, suggesting depolarization blockade had developed; however, all of these preparations gradually regained some degree of spontaneous unit activity following the termination of quisqualate infusion. Quisqualate-induced excitation may be attributable to the activation of receptors for the afferent neurotransmitter released by hair cells. This interpretation is consistent with our working hypothesis that the primary afferent neurotransmitter is L-glutamate or a structural analog of this excitatory amino acid.     

The effects of ethacrynic acid upon the potassium concentration in guinea pig cochlear fluids

After i.v. injection of 50 mg/kg ethacrynic acid (EA), potassium concentration in the endolymph (K_e⁺) measured with K⁺-specific microelectrodes decreases by 10 mM at the most and endocochlear potential falls to negative values. Potassium concentration in the perilymph (K_p⁺) generally does not change, but sometimes a transient decrease in K_p⁺ level of about 0.5 mM was observed, presumably due to the electrogenic effect of the time-related decrease of the endocochlear potential.When anoxia is induced approximately 120 min after EA administration, K_e⁺ slowly decreases. The decrease in K_e⁺ 50 min after the arrest of ventilation is smaller when compared with the K_e⁺ anoxic decrease without preceding EA administration. The endocochlear potential, which falls to negative values during anoxia after EA administration, does not return to the zero level as in the case when only anoxia is applied. Similarly, during anoxia, which follows EA administration, the perilymphatic K_e⁺ concentration increases more slowly than in the case when only anoxia is introduced. It is assumed from the results that EA abolishes activity of the positive electrogenic K⁺ pump and reduces the passive permeability of the walls of the cochlear duct to the potassium ions.