Carbogen气体吸入对癫痫影响的机制研究

Hyperventilation (HV) is most effective in activation of generalized absence seizures and also used as an activation technique in the electrocephalography record. The anticonvulsant effect of carbogen (containing 5%CO2 and 95%O2) was demonstrated in our previous study by decreasing frequency of seizure onset and suppression of the characteristic electrocephalography spike-wave pattern during administration of carbogen containing 5%CO2. However, the mechanisms remain unclear. We establish epilepsy model with kainic acid (KA) in immature rats. Carbogen will be applied immediately after the onset of seizures to observe whether it can help seizure termination. In control group with saline administration, the rats will receive the same amount of carbogen as in the KA group. Cortical pH was measured when the rats inhaled carbogen. After 8 weeks, motor function will be assessed by open field test. Morris water maze will be used to evaluate their visual-spacial learning and memory ability, and improved Y maze be used to evaluate the working memory. We aims to observe the effect of carbogen on seizure onset and figure out its reliability for long time use. The release of neurotransmitter GABA and glutamate will be detected by HPLC. We use in vivo electrophysiology to assess the current in the cortex of hippocampus and temporal lobe, and in vitro whole cell recording in hippocampus slice to record the Ih current. The mossy fiber sprouting in hippocampus will be assessed by Timm's staining. We try to reveal its underlying mechanisms from several aspects, thus provide theoretical fundation for clinical use of carbogen as an anticonvulsant agent.

过度换气是常用的诱发癫痫发作及脑电图异常的方法之一，我们的前期研究发现吸入carbogen(5%CO2+95%O2)气体可以减少失神癫痫患者发作次数及棘慢波持续时间，但是其机制尚不明了。本立题以此为出发点，选择海藻酸致痫发育期大鼠为研究对象，发作时给予大鼠吸入carbogen 气体，观察其对癫痫发作的影响；同时给予正常发育期大鼠反复吸入carbogen。8周后评价大鼠的活动水平，视觉－空间学习记忆能力和工作记忆能力。试图从多个角度观察carbogen对癫痫发作的影响及长期应用的安全性。同时监测吸入前后皮层pH值的变化，测定神经递质GABA及谷氨酸释放的变化，在体电生理测定海马及颞叶皮层电流的变化，体外电生理记录海马CA1区Ih电流，采用Timm's 染色法检测海马区苔藓纤维发芽的情况。从多个层面深入探讨可能的机制，为carbogen气体应用于癫痫及其他原因引起的惊厥的预防提供理论依据。

项目背景：过度换气是常用的诱发癫痫发作及脑电图异常的方法之一，我们的前期研究发现吸入 carbogen(5%CO2+95%O2)气体可以减少失神癫痫患者发作次数及棘慢波持续时间，但是其机制尚不明了。.主要研究内容：本课题选择海藻酸致痫发育期大鼠为研究对象，发作时给予大鼠吸入 carbogen 气体，观察其对癫痫发作的影响；同时给予正常发育期大鼠反复吸入 carbogen作为对照。8 周后评价大鼠的活动水平，视觉－空间学习记忆能力和工作记忆能力。同时监测吸入前后皮层pH 值的变化，采用 Timm's 染色法检测海马区苔藓纤维发芽的情况，测定神经递质 GABA 及谷氨酸释放的变化，在体电生理测定海马及颞叶皮层电流的变化，体外电生理记录海马 CA1 区 Ih 电流。.重要结果、关键数据：Carbogen气体可以延长急性发作后首次出现SRS的潜伏期(由KA组的26.12min±2.11min延长至43.65±2.78min)，减少痫性发作次数。吸入Carbogen气体可以缓解大鼠焦虑情绪。吸入Carbogen气体可显著下调Escape latency （27.16±2.02 vs 50.39±3.05, p<0.001），显著上调目标象限游泳时间 (33.74±2.64 s vs 27.63±2.95 s, p<0.001)，上调站台区域搜寻时间百分比(Search time)，改良Y迷宫实验提示KA组大鼠学习能力下降，而吸入Carbogen气体可恢复大鼠学习能力。进一步对大鼠皮层pH检测发现：单独吸入carbogen 气体即可降低皮层pH值,单独KA也降低皮层pH，二者联合产生协同作用。吸入carbogen 气体后, 兴奋性氨基酸谷氨酸含量显著下降, 兴奋性氨基酸拮抗剂GABA的含量显著上升。此外，吸入carbogen 气体后EHG显著下降; ECoG显著下降。吸入Carbogen气体可显著降低海马CA1区的超极化活化的阳离子电流。.科学意义：本课题揭示了 carbogen 气体影响癫痫发生的机制，为 carbogen 气体应用于癫痫的预防提供理论依据，为开发新的抗癫痫药物奠定基础。