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目的研究低镁介质致痫的培养海马神经元癫痫模型中神经元内游离钙离子([Ca2+]i)的时空分布及其动力学改变,以探讨钙离子在癫痫发病过程中的作用。方法联合应用共聚焦激光扫描显微镜和膜片钳,运用较高时间分辨率动态观察培养海马神经元癫痫模型[Ca2+]i和电生理变化,以及化学门控钙离子通道阻滞剂的影响。结果致痫后海马神经元胞浆和核内游离钙离子迅速上升到(612±65)nmol/L和(620±69)nmol/L水平,NMDA受体阻断剂MK-801(10μmol/L)和非NMDA受体阻断剂NBQX(10μmol/L)可使[Ca2+]i的升高明显减少;升高的[Ca2+]i恢复有明显的延迟现象,90min和150min癫痫样放电后[Ca2+]i恢复的时间分别为(114.8±5.2)和(135.0±22.7)(P<0.05)。结论持续的癫痫样放电可导致海马神经元细胞内钙超载,这个效应可被MK-801阻断,化学门控钙离子通道也参与了细胞外Ca2+内流的过程。
Objective To investigate the spatial and temporal distribution of intracellular free calcium ions ([Ca2 +] i) in hippocampal neurons with epilepsy induced by low magnesium medium and their kinetic changes to explore the role of calcium in the pathogenesis of epilepsy. Methods Combined confocal laser scanning microscopy and patch-clamp were used to observe the changes of [Ca2 +] i and electrophysiological changes of hippocampal neurons in cultured hippocampal neurons with high temporal resolution and the effects of chemosensitive calcium channel blockers. The results showed that the levels of cytoplasm and intracellular free calcium in hippocampal neurons rapidly increased to (612 ± 65) nmol / L and (620 ± 69) nmol / L, and the NMDA receptor antagonist MK- ) And non-NMDA receptor antagonist NBQX (10μmol / L) significantly decreased the increase of [Ca2 +] i; the delayed recovery of [Ca2 +] i increased obviously after 90min and 150min epileptiform discharge [Ca2 + ] i recovery times were (114.8 ± 5.2) and (135.0 ± 22.7), respectively (P <0.05). CONCLUSIONS: Sustained epileptiform discharges can cause intracellular Ca2 + overload in hippocampal neurons. This effect can be blocked by MK-801. Chemically-gated Ca2 + channels are also involved in extracellular Ca2 + influx.