新型抗癫痫药Q808对颞叶癫痫大鼠的改善作用及其机制

doi:10.13481/j.1671-587X.20240507

摘要/Abstract

摘要：

目的探讨新型抗癫痫药6-（4-氯苯氧基）-四唑^{（5，1-a）}酞嗪（Q808）对颞叶癫痫（TLE）大鼠神经元损伤的改善作用，并阐明其作用机制。方法采用腹腔注射Q808的方法制备TLE大鼠模型。将45只造模成功大鼠随机分为模型组、低剂量Q808组和高剂量Q808组，每组15只，低剂量Q808组和高剂量Q808组大鼠分别采用20和80 mg·kg^-1 Q808灌胃，模型组大鼠采用等量0.3%羧甲纤维素钠灌胃，另选15只健康SD大鼠作为对照组。持续灌胃治疗4周后，观察各组大鼠行为表现，采用PONEMAH 6.X实验动物遥测平台记录各组大鼠脑电图，采用高尔基染色观察各组大鼠海马CA1神经元树突形态表现和神经元树突棘密度，采用Western blotting法检测各组大鼠海马组织中突触可塑性特异性蛋白钙/钙调素依赖性蛋白激酶Ⅱ（CaMKⅡ）蛋白表达水平。结果对照组大鼠活动正常，无抽搐等异常表现；模型组、低剂量Q808和高剂量Q808组大鼠出现不同程度活动减少、震颤点头、失去平衡、肌肉强直和前肢的抽搐，逐渐转变为全身肌肉强直和站立，随后向后跌倒，发作间期无抽搐发作。与对照组比较，模型组、低剂量Q808和高剂量Q808组大鼠癫痫发作总持续时间明显延长（P<0.01）；与模型组比较，低剂量Q808和高剂量Q808组大鼠癫痫发作总持续时间明显缩短（P<0.01）。对照组大鼠海马组织CA1区神经元树突分布较为规律，树突网密集有序；模型组大鼠海马组织CA1区神经元树突排列紊乱，大量树突缠结，形成较粗大的神经纤维束；与模型组比较，低剂量Q808和高剂量Q808组大鼠海马组织CA1区神经元树突网络有所恢复，排列相对规律。与对照组比较，模型组大鼠海马组织CA1区神经元树突棘密度明显降低（P<0.01）；与模型组比较，低剂量Q808和高剂量Q808组大鼠海马组织CA1区神经元树突棘密度明显升高（P<0.01）。与对照组比较，模型组、低剂量Q808组和高剂量Q808组大鼠海马组织中CaMKⅡ蛋白表达水平明显降低（P<0.01）；与模型组比较，低剂量Q808组和高剂量Q808组大鼠海马组织中CaMKⅡ蛋白表达水平明显升高（P<0.01）。结论新型抗癫痫药Q808对TLE模型大鼠有改善作用，其机制可能与Q808能减轻海马组织CA1区神经元树突病变和增加突触可塑性相关蛋白CaMKⅡ蛋白表达水平有关。

关键词: 颞叶癫痫, 6-(4-氯苯氧基)-四唑^(5,1-a)酞嗪, 突触可塑性, 钙/钙调素依赖性蛋白激酶Ⅱ, 长时程增强

Abstract:

Objective To discuss the ameliorative effect of a novel antiepileptic drug Q808 on neuronal injury in temporal lobe epilepsy （TLE） rats， and to clarify its mechanism of action. Methods TLE rat model was prepared by intraperitoneal injection of the innovative antiepileptic drug candidate 6-（4-chlorophenoxy）- tetrazolo^{（5，1-a）} phthalazine （Q808）. Forty-five successfully modeled rats were randomly divided into model group， low dose of Q808 group， and high dose of Q808 group， and there were 15 rats in each group. The rats in low dose of Q808 group and high dose of Q808 group were gavaged with 20 and 80 mg·kg^-1 Q808，respectively， and the rats in model group were gavaged with an equal amount of 0.3% sodium carboxymethyl cellulose. Another 15 healthy SD rats were selected as control group. After 4 weeks of continuous gavage treatment， the morphology of the rats in varioius groups was observed； PONEMAH 6.X experimental animal telemetry platform was used to record the electroencephalogram of the rats in various groups； Golgi staining was used to observe the morphology of dendritic and dendritic spine density of hippocampal CA1 neurons of the rats in various groups； Western blotting method was used to detect the expression levels of synaptic plasticity-specific protein calcium/calmodulin-dependent protein kinaseⅡ （CaMKⅡ） in hippocampus tissue of the rats in various groups. Results The rats in control group showed normal activity without convulsions or other abnormal manifestations. The rats in model group， low dose of Q808 group， and high dose of Q808 group showed varying degrees of reduced activity， trembling and nodding， loss of balance， muscle rigidity and forelimb convulsions， gradually transforming into whole-body muscle rigidity and standing， followed by falling backwards， and there were no convulsions during the interictal period. Compared with control group， the total durations of epileptic seizures of the rats in model group， low dose of Q808 group， and high dose of Q808 group were significantly prolonged （P<0.01）. Compared with model group， the total durations of epileptic seizures in low dose of Q808 group and high dose of Q808 group were significantly shortened （P<0.01）. The hippocampal CA1 neurons of the rats in control group showed regular distribution of dendrites with dense and orderly dendritic networks. The hippocampal CA1 neurons of the rats in model group showed disordered arrangement of dendrites with massive dendritic entanglement， forming thicker nerve fiber bundles. Compared with model group， the dendritic networks of hippocampal CA1 neurons of the rats in low dose of Q808 group and high dose of Q808 group were partially recovered with relatively regular arrangement. Compared with control group， the dendritic spine density of hippocampal CA1 neurons of the rats in model group was significantly decreased （P<0.01）. Compared with model group， the dendritic spine densities of hippocampal CA1 neurons in low dose of Q808 group and high dose of Q808 group significantly increased （P<0.01）. Compared with control group， the expression levels of CaMKⅡ protein in hippocampus tissue of the rats in model group， low dose of Q808 group， and high dose of Q808 group were significantly decreased （P<0.01）. Compared with model group， the expression levels of CaMKⅡ protein in hippocampus tissue of the rats in low dose of Q808 group and high dose of Q808 group were significantly increased （P<0.01）. Conclusion The novel antiepileptic drug Q808 has an ameliorating effect on the TLE model rats；its mechanism may be related to Q808’s ability to reduce the dendritic lesions in hippocampal CA1 neurons and increase the expression level of synaptic plasticity-related protein CaMKⅡ protein.

Key words: Temporal lobe epilepsy, 6-（4-chlorophenoxy）-tetrazolio^{（5，1-a）} phthalazine, Synaptic plasticity, Calcium/calmodulin-dependent protein kinaseⅡ, Long-term potentiation

中图分类号:

R742.1

郑微微,高帆,杨振林,李佳芮,郭晶晶,李今子. 新型抗癫痫药Q808对颞叶癫痫大鼠的改善作用及其机制[J]. 吉林大学学报(医学版), 2024, 50(5): 1243-1249.

Weiwei ZHENG,Fan GAO,Zhenlin YANG,Jiarui LI,Jingjing GUO,Jinzi LI. Ameliorative effect of novel antiepileptic drug Q808 on rats with temporal lobe epilepsy and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2024, 50(5): 1243-1249.

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参考文献 16

1	MAJEED C N， RUDNICK S R， BONKOVSKY H L. Review of antiseizure medications for adults with epilepsy［J］. JAMA， 2022， 328（7）： 680.
2	FALCO-WALTER J. Epilepsy-definition， classification， pathophysiology， and epidemiology［J］. Semin Neurol， 2020， 40（6）： 617-623.
3	FIEST K M， SAURO K M， WIEBE S， et al. Prevalence and incidence of epilepsy： a systematic review and meta-analysis of international studies［J］. Neurology， 2017， 88（3）： 296-303.
4	WIESER H G. ILAE Commission on Neurosurgery of Epilepsy. ILAE Commission Report. Mesial temporal lobe epilepsy with hippocampal sclerosis［J］. Epilepsia，2004， 45（6）： 695-714.
5	SUN X Y， WEI C X， DENG X Q， et al. Evaluation of the anticonvulsant activity of 6-（4-chlorophenyoxy）- tetrazolo［5，1-a］phthalazine in various experimental seizure models in mice［J］. Pharmacol Rep， 2010， 62（2）： 273-277.
6	LI X， WANG Q， WU D， et al. The effect of a novel anticonvulsant chemical Q808 on gut microbiota and hippocampus neurotransmitters in pentylenetetrazole-induced seizures in rats［J］. BMC Neurosci， 2022， 23（1）： 7.
7	丁永强，陈超，李信晓，等. 脑卒中后癫痫患者发生抗癫痫药物耐药的预测因素分析［J］. 郑州大学学报（医学版）， 2024， 59（4）： 523-526.
8	DE SOUZA D L S， COSTA H M G E S， IDALINA NETA F， et al. Brain neuroplasticity after treatment with antiseizure： a review［J］. Clin Psychopharmacol Neurosci， 2023， 21（4）： 665-675.
9	PRIBIAG H， PENG H S， SHAH W A， et al. Dystroglycan mediates homeostatic synaptic plasticity at GABAergic synapses［J］. Proc Natl Acad Sci U S A， 2014， 111（18）： 6810-6815.
10	RACINE R J. Modification of seizure activity by electrical stimulation： Ⅱ. Motor seizure ［J］. Electroencephalogr Clin Neurophysiol， 1972， 32（3）： 281-294.
11	徐圣洁，席加秋，于晓雯，等. 内质网应激与胶质细胞激活介导的神经炎症在癫痫中的作用研究进展［J］. 解放军医学杂志， 2024， 49（4）： 475-481.
12	URBAN P， REZAEI T V， DENKIEWICZ M， et al. The mixture of autoregressive hidden markov models of morphology for dentritic spines during activation process ［J］. J Comput Biol， 2020， 27（9）： 1471-1485.
13	BLAZEJCZYK M， MACIAS M， KOROSTYNSKI M， et al. Kainic acid induces mtorc1-dependent expression of elmo1 in hippocampal neurons ［J］. Mol Neurobiol， 2017， 54（4）： 2562-2578.
14	NAKAHATA Y， YASUDA R. Plasticity of spine structure： local signaling， translation and cytoskeletal reorganization［J］. Front Synaptic Neurosci， 2018， 10： 29.
15	BIAN H， YU L C. Intra-nucleus accumbens administration of the calcium/calmodulin-dependent protein kinaseⅡ inhibitor AIP induced antinociception in rats with mononeuropathy ［J］. Neurosci Lett， 2015， 599： 129-132.
16	TAKEUCHI Y， YAMAMOTO H， FUKUNAGA K， et al. Identification of the isoforms of Ca²⁺/Calmodulin-dependent protein kinase Ⅱ in rat astrocytes and their subcellular localization ［J］. J Neurochem， 2000， 74（6）：2557-2567.

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[3]	梁建民\|张洪波\|李海波\|崔新明\|张淑琴. 小剂量地塞米松对颞叶癫痫大鼠空间记忆能力的保护作用[J]. J4, 2012, 38(1): 42-45.
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