腺苷对氯化锂-匹罗卡品致痫大鼠海马神经元癫痫样放电的影响

Effect of adenosine on epileptiform discharge of hippocampal neurons from lithium chloride-pilocarpine induced epileptic rats

目的 使用膜片钳全细胞记录方式,观察腺苷对氯化锂-匹罗卡品致痫大鼠海马锥体神经元癫痫样放电的影响.方法 选20只200 9左右的成年SD大鼠,雌雄不拘,使用氯化锂-匹罗卡品制备大鼠癫痫模型,造模成功24 h后,使用10％水合氯醛腹腔注射麻醉,断头取海马,再修块切片,厚度约350 μm,移入37℃预热的人工脑脊液中孵育1h后,转入23℃水浴中孵育30 min备用.全细胞膜片钳记录海马脑片CA1区锥体神经元的动作电位及诱发的兴奋性突触后电流,并观察25μmol/L腺苷对上述指标的干预情况.结果 通过电流钳模式记录发现,腺苷对海马CA1区神经元的动作电位频率有明显的抑制作用;通过电压钳模式记录发现,癫痫大鼠[（1.44±0.06） nA]海马神经元诱发的兴奋性突触后电流幅值较正常大鼠[（0.50±0.06）nA]的明显升高（独立样本t检验,t=30.99,P＜0.01）;25 μmol/L腺苷对海马CA1区神经元诱发的兴奋性突触后电流的幅值抑制明显[（0.66 ±0.06） nA],加入腺苷A1受体抑制剂DPCPX（10 μmol/k）又可以使幅值回升[（0.92±0.06）nA]（单因素方差分析及q检验,F =266.3,q=9.81,P＜0.01）.结论 腺苷抑制氯化锂-匹罗卡品致痫大鼠海马锥体神经元癫痫样放电,是通过对突触后兴奋性的干预完成的.

Objective To use the whole-cell patch clamp recording to observe the effect of adenosine on hippocampus pyramidal neurons epileptiform discharge from lithium chloride-pilocarpine induced epileptic rats.Methods Twenty adult male and female SD rats （weighing about 200 g） were selected and were narcotized by 10% chloral hydrate intraperitoneal injection.The whole brain was removed, then chopped into 350 μm thick slices.The brain slices were transferred to 37 ℃ preheated artificial cerebrospinal fluid for 1 h and then transferred to 23 ℃ for 30 min.Action potentials （AP） and evoked excitatory postsynaptic currents （eEPSCs） of brain slices neurons were recorded by whole-cell patch clamp, in addition, 25 μmol/L adenosine was used to observe its effect on AP and eEPSCs.Results With the application of current clamp technique, the application of adenosine （25 μmol/L） significandy decreased the numbers of AP in epileptic neurons （P 〈 0.01） compared with the normal neurons （（0.50 ± 0.06） nA）, and eEPSCs amplitude of epileptic neurons increased significantly （（1.44 ± 0.06） nA;independent sample t test, t =30.99, P 〈 0.01） by voltage clamp.Bath application of adenosine （25 μmol/L） significantly reduced eEPSCs amplitude （（0.66 ±0.06） nA）, while application of A1 receptor inhibitor DPCPX partially reversed this effect （（0.92 ± 0.06） nA;one-way analysis of variance and q test, F =266.3, q =9.81, P 〈 0.01）.Conclusion Adenosine possesses a strong inhibitory effect on epileptic hippocampal brain slices discharge, which is mediated by its effect on eEPSCs.