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高胆红素致耳蜗核神经元超兴奋的机制研究

The Role of Synaptic Transmission and Intrinsic Firing in Bilirubin-induced Hyperexcitability in Cochlear Nucleus

尹新璐;陈博婕;刘汉玮;徐雅男;王家东;

1:上海交通大学医学院附属仁济医院头颈外科

2:上海交通大学附属第六人民医院耳鼻咽喉头颈外科

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摘要

目的探讨胆红素引起耳蜗核(cochlear nuclear, CN)神经元超兴奋性的机制。方法选用出生后1～9天(P1～9)C57Bl/6J小鼠60只，制备含有CN的脑片。利用膜片钳技术，用细胞贴附式或者全细胞式记录神经元自发性动作电位、超极化激活环磷腺苷依赖性阳离子通道(hyperpolarization-activated and cyclic nucleotide-gated channels, HCN)电流、自发性抑制性突触后电流(spontaneous inhibitory postsynaptic current, sIPSC)和自发性兴奋性突触后电流(spontaneous excitatory postsynaptic current, sEPSC)。神经元在人工脑脊液中自发性放电或者阻断突触的内源性放电记为对照组，随后在脑脊液中灌流加6μm胆红素作用9 min记为胆红素组，最后回归至人工脑肾液中记为洗脱组。加药过程由一个多阀门控制，依靠重力单向输出的灌流装置完成，平均流速在2 ml/min。结果胆红素可以提高CN神经元自发性放电频率(胆红素组较对照组升高了250%±31.2%;洗脱组降为对照组的162%±21.4%);胆红素可以促进sEPSC(胆红素组增加至对照组的193.2%±26.4%)和sIPSC(胆红素组增加至对照组的135.3%±16.4%)频率增大，说明胆红素可以促进谷氨酸和GABA/甘氨酸能突触传递。CN神经元存在不依赖于突触传递的自发性内源性放电，并且这种放电是由HCN离子通道起搏介导的。HCN通道的抑制剂CsCl和ID7288可以明显降低自发性内源性放电的频率(CsCl:降至对照组的48.75%±9.23%,ZD7288:降至对照组的68.45%±10.39%)。胆红素作用后，HCN通道电流的激活曲线右移(V_(0.5):对照组-104.9±1.5 mV,胆红素组：-95.4±2.2 mV)。抑制HCN通道电流后，高胆红素不再发挥超兴奋作用。结论胆红素通过促进神经突触和HCN通道介导的内源性放电发挥超兴奋作用。

关键词

胆红素;超兴奋性;突触传递;内源性放电;膜片钳

基金项目(Foundation):

作者

尹新璐;陈博婕;刘汉玮;徐雅男;王家东;

参考文献

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胆红素超兴奋性突触传递内源性放电膜片钳

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尹新璐陈博婕刘汉玮徐雅男王家东