Cultured hippocampal neurons were used in the study. First, the viability of neurons was determined by the CCK-8 assay after culturing in magnesium-free medium, DMSO, dynasore (dynamin agonist), and PIP2 (dynamin antagonist). Then, the effect of dynasore on seizure activity was evaluated. Next, we tested the levels of phospho-dynamin 1/total dynamin 1 and dynamin 1 mRNA in the control group and four epilepsy groups. Moreover, the uptake of tetramethylrhodamine-dextran in the different groups was measured.
Dephospho-dynamin 1 expression was significantly increased in hyperexcitable neurons, while there was no change in total dynamin 1 level. The level of dephospho-dynamin 1 in hyperexcitable neurons was reduced when cultured with dynasore but increased with PIP2 treatment. Activity-dependent bulk endocytosis (ADBE) was upregulated in hyperexcitable neurons. Along with a decrease in dephospho-dynamin 1 level, ADBE was also downregulated with dynasore treatment, while PIP2 did not affect ABDE. The close link between the dephosphorylation status of dynamin 1 and ADBE suggests that ADBE activation depends on dynamin 1 dephosphorylation.
Dephospho-dynamin 1 triggers ADBE to meet the requirements of high-frequency discharges during epileptic seizures.
在研究中使用培养的海马神经元。首先,在无镁培养基中培养后,通过CCK-8测定确定神经元的活力,DMSO,王朝(动力蛋白激动剂),和PIP2(动力蛋白拮抗剂)。然后,评估了朝代对癫痫发作活动的影响。接下来,我们检测了对照组和四个癫痫组的磷酸-动力蛋白1/总动力蛋白1和动力蛋白1mRNA的水平。此外,测量了不同组的四甲基罗丹明-葡聚糖的摄取。
去磷酸-动力蛋白1在高兴奋神经元中表达显著增加,而总动态蛋白1水平没有变化。与dynasore一起培养时,可兴奋神经元中的去磷酸-动力蛋白1水平降低,但随着PIP2处理而升高。活动依赖性大量内吞作用(ADBE)在可过度兴奋的神经元中上调。随着去磷动力学1水平的降低,ADBE也随着朝代治疗而下调,而PIP2不影响ABDE。动态蛋白1的去磷酸化状态与ADBE之间的紧密联系表明ADBE的激活取决于动态蛋白1的去磷酸化。
去磷动力蛋白1触发ADBE,以满足癫痫发作期间高频放电的要求。