腺苷对原代培养的大鼠海马神经元大电导钙激活钾通道的作用

目的在离子通道水平研究腺苷对原代培养的大鼠海马神经元大电导钙激活钾通道(BKCa)的作用。方法采用新生24h的乳鼠,取出海马,清除表面血管,以胰酶消化和巴氏管吹打的方法制备海马神经元,细胞培养6～8d后,MAPⅡ染色鉴别神经元表面标志;采用全细胞膜片钳技术,对细胞进行封接、破膜并记录BKCa通道电流,以细胞旁方式给药观察不同浓度腺苷对BKCa通道的作用。结果原代培养的海马神经元上可以成功记录到BKCa电流,其为一组幅值较大,可以快速激活且几乎不失活的外向电流;腺苷(1～100μmol/L)可以增大BKCa电流,浓度越高,增长幅度越大;100μmol/L腺苷对BKCa电流的影响具有电压依赖性,同时它还可以使BKCa电流的激活曲线负向漂移。结论腺苷可以增大海马神经元BKCa电流幅值,这可能是腺苷作为内源性抗癫痫物质发挥抗癫痫作用的机制之一。

Effect of Adenosine on Large Conductance Calcium-activated Potassium Channels in Cultured Hippocampal Neurons

Objective To investigate the effects of adenosine on large conductance calcium-activated potassium channels(BKCa)in cultured hippocampal neurons.Methods The hippocampi were isolated from 24 h neonate rats and blood vessels outside were removed.The neurons were prepared by trypsinization and blowing using pastometer.Immunofluorescence of MAPⅡ was used to identify neurons 6-8 days after culture.Current of BKCa was recorded by a whole-cell voltage-clamp technique,and effect of different doses of adenosine on BKCa was observed.Results BKCa channels were recorded on cultured hippocampal neurons,which was outward current and characteristic of high amplitude,quick activation and almost never inactivation.The current of BKCa was increased by adenosine in a concentration-dependent manner at the doses of 1-100 μmol/L.Adenosine(100 μmol/L)could increase BKCa in a voltage-dependent manner;moreover adenosine(100 μmol/L)made the activation curve of this current shift towards more negative values.Conclusion Adenosine could increase the amplitude of BKCa,which may be the anti-epileptic mechanism of adenosine as an endogenous anticonvulsant.