大鼠癫痫持续状态后海马组织各区脑红蛋白表达的实验研究

目的观察大鼠癫痫持续状态(Status epilepticus,SE)后海马组织脑红蛋白(Neuroglobin,NGB)表达动态变化,探讨NGB在癫痫发作中的作用。方法健康成年雄性SpragueDawley大鼠40只,随机分为对照组(n=5)、癫痫模型实验组(n=35);实验组再依据观察时间分为:0、1、3、12、24 h和10、30 d。应用锂-匹罗卡品(20~127 mg/kg)建立大鼠SE模型,观察大鼠致痫期间行为学变化;采用尼氏(Nissl)染色检测海马组织神经元损伤情况;SABC免疫组化法检测海马组织NGB表达水平。结果 SE后,海马组织各区均出现不同程度神经元细胞损伤坏死,随着发作时程进展,CA1、CA3区存活神经元呈近直线下降趋势。其中CA1区(12、24 h,10、30d)、CA3区(0、12、24 h,10、30 d)和(DG区12、24 h,10、30 d)神经元存活数较对照组明显减少(P0.05)。大鼠SE后,海马各区NGB表达水平均上调,CA1、DG区NGB表达均于SE后24 h达顶峰后轻度下降,但仍持续高于对照组,CA3区NGB表达呈持续升高趋势。其中CA1区(24 h,10、30 d)、CA3区(24 h,10、30 d)和DG区(12、24 h,10、30 d)NGB表达水平均较对照组显著升高(P0.05)。另外,海马CA1和CA3区神经元存活数与NGB表达水平呈正相关(R=0.206,P=0.015;R=0.306,P=0.011)。结论大鼠SE后海马各区NGB表达上调,且与CA1、CA3区神经元存活数呈正相关,提示NGB表达上调可能是癫痫发作所致缺血缺氧损害的一种代偿保护机制,参与癫痫相关神经元损害的保护。     

颞叶癫痫大鼠海马XIAP相关因子-1蛋白表达变化研究

目的动态观察颞叶癫痫大鼠海马神经元XIAP相关因子-1(XAF1)的表达变化,探讨其在癫痫发生发展中的作用。方法清洁级SD雄性成年大鼠36只,随机分为正常对照组、致痫(SE)后3h、6h、12h、24h、72h组。用氯化锂(LiCl)和匹罗卡品(PILO)制备癫痫动物模型,应用免疫组化染色技术检测致痫后各时间点XAF1蛋白表达情况。结果对照组海马CA1、CA3区神经元XAF1蛋白表达极少,SE组CA1、CA3区XAF1蛋白表达在3～24h表达逐渐增高,72h表达有所下降,但仍高于对照组,各时间点两组间差异均有统计学意义(P0.01)。结论SE大鼠海马神经元XAF1表达增高,它可能参与了SE后脑神经元凋亡的调控。     

大鼠癫痫持续状态后海马TNF-α、IL-1β的动态变化

目的观察大鼠癫痫持续状态(status epilepticus,SE)后海马组织各区细胞因子的动态变化,探讨炎症反应与癫痫发作的关系。方法雄性SD大鼠60只,随机分为致痫组(n=54)和对照组(n=6),致痫组再根据观察时间分为:SE-0 h组、1 h组、3 h组、12 h组、24 h组和10 d组。应用锂-匹罗卡品建立大鼠SE模型,观察大鼠行为学变化,采用尼氏染色检测海马组织神经元损伤情况,酶联免疫吸附法(ELISA)及免疫组化法检测海马组织TNF-α、IL-1β的表达水平。结果 SE后,大鼠海马组织IL-1β、TNF-α蛋白含量显著升高,其中(3 h组、12 h组、24 h组、10 d组)IL-1β和1h TNF-α蛋白含量均较对照组有统计学差异(P0.05)。免疫组化证实,海马各区IL-1β和TNF-α表达水平也均上调,其中(12 h组、24 h组)CA1、(12 h组、24 h组)CA3和12 h组DG区IL-1β表达水平均较对照组有统计学差异(P0.05);3 h组CA1、(1 h组、3 h组、24 h组、10 d组)CA3和(0h组、1 h组、3 h组、12 h组)DG区TNF-α表达水平均较对照组有统计学差异(P0.05)。结论癫痫发作后海马组织TNF-α、IL-1β表达上调,且持续处于高表达水平,提示癫痫发作可能诱发炎症反应。     

钾通道Kv1.2与致痫大鼠发病相关性的研究

目的通过检测戊四唑(PTZ)致痫大鼠钾通道Kv1.2蛋白表达,探讨钾通道Kv1.2与癫痫发病的相关性。方法 40只SD大鼠分成实验组30只和正常对照组10只。实验组30只大鼠通过腹腔注射PTZ建立全身强直阵挛发作大鼠癫痫模型,取成功致痫鼠24只均分成3组,分别于致痫后3个时间段(1h、24h、48h)取脑组织。用免疫组化法和Western blot法检测大鼠钾通道Kv1.2蛋白。结果实验组大鼠海马区钾通道Kv1.2蛋白表达水平在致痫后3个时间段(1h、24h、48h)均明显低于对照组(P0.05)。实验组大鼠海马区钾通道Kv1.2蛋白表达水平在致痫后3个时间段(1h、24h、48h)之间无显著性差异(P0.05)。结论大鼠海马区钾通道Kv1.2表达的减少与全身强直阵挛发作大鼠癫痫发病密切相关。     

大鼠癫痫持续状态后认知功能变化模式及海马脑红蛋白表达水平的实验研究

目的观察大鼠癫痫持续状态(SE)后学习记忆功能改变情况及海马组织脑红蛋白(NGB)表达水平,探讨癫痫发作对认知功能影响的可能机制。方法健康成年雄性SD大鼠40只,随机分为对照组(n=5)、癫痫模型实验组(n=35),模型组再依据观察时间分为:0 h、1 h、3 h、12 h、24 h、10 d、30 d。应用氯化锂-匹罗卡品(Li-Pilo)建立SE模型,观察致痫期间大鼠行为学变化;采用Nissl染色检测神经元损伤情况;SABC免疫组化法检测NGB表达水平。同时随机选取同期相同品系SD大鼠40只,在造模前及造模后第5d、10 d、15 d、25 d、35 d进行RMT-100迷宫实验,以评价大鼠SE前后学习记忆功能变化情况。结果大鼠SE后,海马CA1、CA3区和DG区均出现不同程度神经元细胞损伤坏死,且NGB表达上调,而海马CA1和CA3区神经元存活数与NGB表达水平呈正相关(r=0.206,P=0.015;r=0.306,P=0.011)。迷宫实验显示工作记忆错误(WME)和参照记忆错误(RME)次数随SE后时间延长均呈递增趋势。相关性分析证实RME次数与CA1和CA3区神经元存活数呈负相关(r=-0.579,P=0.000;r=-0.454,P=0.002),WME次数与CA1和CA3区神经元存活数也呈负相关(r=-0.470,P=0.001;r=-0.507,P=0.000)。结论 SE后NGB表达上调,且与海马组织神经元存活数呈正相关,提示其可能是SE所致缺血缺氧损害的一种代偿神经保护机制。SE后可导致明显认知功能损害,其可能与SE所致海马组织神经元的病理改变相关。     

红藻氨酸致痫大鼠海马神经元凋亡及其与caspase-3关系的研究

目的　研究大鼠癫痫发作后海马神经元凋亡及其与天冬氨酸特异性半胱氨酸蛋白酶 -3 (cysteinylasparate-specific proteinase,caspase-3 )表达的关系。方法　采用红藻氨酸 (kainic acid,KA)诱导大鼠癫痫模型 ,以原位末端标记 (TUNEL)及透射电镜检测癫痫发作后 6h及 1、3、7d海马神经元凋亡 ;半定量 RT-PCR及免疫组化法检测 caspase-3 m RNA及 caspase-3阳性表达。结果　KA致痫后 1 d,海马 CA1、CA3及 CA4区开始出现凋亡细胞 ,3 d时明显增多 ,7d时最多。 3个时间组相应区域间凋亡神经元数比较差异均有显著性 (P<0 .0 0 1 )。透射电镜观察可见典型的凋亡细胞形态学改变。 RT-PCR结果显示 ,KA致痫后 6h,海马组织 caspase-3 m RNA表达较对照组显著增高 (P <0 .0 5 ) ,1、3、7d caspase-3 m RNA仍持续高水平表达 (P <0 .0 5 )。免疫组化结果显示 ,KA致痫后 1 d,海马 CA1、CA3、CA4区开始出现 caspase-3阳性表达 ,3 d时阳性表达进一步增强 ,7d时表达最强。结论　凋亡参与 KA致痫大鼠癫痫发作后海马神经元迟发性死亡过程 ,caspase-3可能在癫痫后神经元凋亡过程中具重要的作用。     

实验性癫痫大鼠颞叶及海马损害的病理学观察

目的　观察癫痫持续不同时间脑组织学损害特点。方法　Wistar大鼠 80只 ,以 1 0 %Pilocarpine 350mg/kg腹腔注射 ,诱发强直 痉挛全面大发作持续状态 ,脑标本制成 30 μm层厚的横轴位切片 ,HE染色 ,光镜下观察癫痫持续 5min至 60h颞叶、海马组织学改变 ,并对海马CA1 区和CA3 区神经元丢失做了定量计数。结果　癫痫持续 (SE) 5～ 30min,颞叶、海马区以神经元变性为主 ;SE 30min～ 3h,轻度神经元坏死、丢失及胶质增生 ;癫痫持续 3h～ 6h后 ,可见颞叶、海马区严重神经元脱失、胶质增生 (即近中颞叶硬化 ,MTS)和脑水肿 ;CA1 区和CA3 区神经元丢失与发作持续时间相关。结论　①随着癫痫持续时间延长 ,神经元损伤加重。②大鼠癫痫持续 3h以上 ,可形成MTS。③大鼠对pilocarpine诱导的持续性癫痫的神经元损伤的易感区依次为CA1 >CA3>齿状回 >CA2 >颞叶     

全蝎醇提物对Li-Pilo癫痫持续状态大鼠海马神经细胞凋亡的影响

目的 探讨全蝎醇提物(Ethanol Extracts of Scorpion,EES)对大鼠癫痫持续状态(SE)后海马神经元凋亡的影响.方法 建立氯化锂.匹罗卡品癫痫持续状态(Lithium-Pilocarpine induced status epilepticus,Li-Pilo SE)模型.使用TUNEL技术观察正常对照组、Li-Piio SE模型组、丙戊酸钠组(VPA)和EES低(L)、中(M)、高(H)剂量组大鼠SE后6h、24h、48h、72h和7d海马CA_1区和CA_3区TUNEL阳性细胞的动态变化,并进行组间比较.结果 正常对照组未见TUNEL阳性细胞.造模各组大鼠SE后6h海马可见部分TUNEL阳性细胞,主要分布在CA_1、CA_3区,其中模型组和EES(L)组72h达高峰,而VPA、EES(M)和EES(H)组高峰提前到SE后48h,以后各组逐渐下降.VPA、EES(M)和EES(H)组SE后各观察时间点TUNEL阳性细胞数较模型组极显著减少(P<0.01),而EES(L)组除个别时间点外,与模型组比较无统计学差异(P>0.05),其中VPA和EES(H)组各时间点TUNEL阳性细胞数减少较EES(M)组(P<0.01,和P<0.05)及EES(L)组(P<0.01,和P<0.05)更显著,EES(M)组各时间点减少又较EES(L)组更显著(P<0.01),而VPA与EES(H)组TUNEL阳性细胞数在各时间点无统计学差异(P>0.05).结论 EES能防止Li-Pilo SE大鼠海马神经元凋亡,并呈明显的量-效关系,高剂量EES抗凋亡作用与VPA相近.     

半胱氨酸蛋白酶在实验性癫痫神经元损伤中的作用

目的:通过观察实验性癫痫发作时海马半胱氨酸蛋白酶(Caspase-3)表达变化,探讨神经元损伤及癫痫发病机制。方法:利用锂-匹罗卡品(Li-PC)制作大鼠急性癫痫持续状态(SE)30min、1h、2h的癫痫模型,造模后1d、3d、7d、14d,经流式细胞技术(FCM)半定量检测Caspase-3的表达含量,并应用光镜及电镜进行神经元形态学观察。结果:模型组海马内Caspase-3表达与SE早期持续时间呈正相关(SE30min3.51±0.35,1h4.49±0.14,2h7.39±0.26p<0.001);各组中均可见Caspase-3表达1d活化,7d表达达峰值(SE30min组1d时3.51±0.35,7d时10.34±0.36;1h组1d时4.49±0.14,7d时14.33±0.33;2h组1d时7.39±0.26,7d时31.15±0.79;p<0.01)。电镜下细胞超微结构凋亡、缺失明显,以海马门区、CA1区为著。结论:Caspase-3是神经元凋亡关键的执行因子,它可以反映SE对神经元造成损伤程度。急性期SE所致的海马区缺血缺氧、水肿以及兴奋毒性导致神经元损伤,可能是慢性颞叶癫痫的病理基础。     

钾通道Kv4.2、Kv1.4在硫化氢后处理对大鼠短暂全脑缺血神经保护中的作用研究

目的研究硫氢化钠(sodium hydrosulfide,Na HS)后处理对短暂全脑缺血大鼠海马中钾通道Kv4.2和Kv1.4 mRNA表达变化的影响及其脑保护作用,从而探讨Na HS对大鼠短暂全脑缺血神经保护作用的机制。方法用4VO方法建立大鼠短暂性全脑缺血(transient global cerebral ischemia,t GCI)模型,大鼠被随机分配到3组,分别为:假手术组(sham)、t GCI组、Na HS后处理组。Na HS后处理组为t GCI之后1 d,给予大鼠腹腔注射Na HS 24μmmol/kg或者180μmmol/kg。通过尼氏染色与Neu N免疫染色确定海马神经元的死亡,通过RT-PCR方法检测海马组织Kv4.2和Kv1.4mRNA水平的表达变化。结果 (1)与t GCI组比较,在t GCI之后1 d给予24μmol/kg Na HS后处理使海马CA1区存活细胞数目显著增加,而高剂量的Na HS(180μmol/kg)后处理对t GCI大鼠海马CA1区则无明显的保护作用。(2)在Re 26 h和Re 48 h,海马组织中Kv4.2、Kv1.4的mRNA表达水平均明显低于假手术组(P<0.05)。在Re 26 h+Na HS组,kv4.2(1.24±0.08)和kv1.4(1.11±0.07)的mRNA表达水平均分别高于Re 26 h组的kv4.2(0.75±0.04)和kv1.4(0.79±0.06),差异均有显著性(P<0.05)。结论外源性Na HS可能通过上调大鼠t GCI后海马区Kv4.2和Kv1.4 mRNA的表达,从而导致膜电位超极化,降低神经元兴奋性和氧耗,继而保护神经元免受脑缺血损伤。     

Epilepsy and anomalies of neuronal migration: MRI and clinical aspects

Neuronal migration disorders are the result of disturbed brain development. In such disorders, neurons are abnormally located. In diagnosing these conditions, magnetic resonance imaging is superior to any other imaging technique. This enables us to improve our knowledge of the clinical correlates of neuronal migration. With reference to migrational disorder, a retrospective study of all 303 patients with epileptic seizures referred for magnetic resonance imaging during a 3-year period was performed, 13 patients (aged 12-41, mean age 27) were identified. They represent 4.3% of the entire study group. Of the patients with known epilepsy, 6.7% and of the mentally retarded, 13.7% had migrational disorders. Four patients had schizencephaly as the dominant finding, one was classified as hemimegalencephaly, 2 had isolated heterotopias, and 6 had localized pachy- and/or poly-microgyria. The clinical pictures are complex. Ectopias of grey matter are recognised foci of epilepsy, but from an epileptological and a clinical viewpoint little attention has been given to these disorders. The present study shows that malmigration is not rare in epilepsy patients, especially not in the mentally retarded.     

Classification of methods in transcranial Electrical Stimulation (tES) and evolving strategy from historical approaches to contemporary innovations

Transcranial Electrical Stimulation (tES) encompasses all methods of non-invasive current application to the brain used in research and clinical practice. We present the first comprehensive and technical review, explaining the evolution of tES in both terminology and dosage over the past 100 years of research to present day. Current transcranial Pulsed Current Stimulation (tPCS) approaches such as Cranial Electrotherapy Stimulation (CES) descended from Electrosleep (ES) through Cranial Electro-stimulation Therapy (CET), Transcerebral Electrotherapy (TCET), and NeuroElectric Therapy (NET) while others like Transcutaneous Cranial Electrical Stimulation (TCES) descended from Electroanesthesia (EA) through Limoge, and Interferential Stimulation. Prior to a contemporary resurgence in interest, variations of transcranial Direct Current Stimulation were explored intermittently, including Polarizing current, Galvanic Vestibular Stimulation (GVS), and Transcranial Micropolarization. The development of these approaches alongside Electroconvulsive Therapy (ECT) and pharmacological developments are considered. Both the roots and unique features of contemporary approaches such as transcranial Alternating Current Stimulation (tACS) and transcranial Random Noise Stimulation (tRNS) are discussed. Trends and incremental developments in electrode montage and waveform spanning decades are presented leading to the present day. Commercial devices, seminal conferences, and regulatory decisions are noted. We conclude with six rules on how increasing medical and technological sophistication may now be leveraged for broader success and adoption of tES.     

Hepatic Considerations in the Use of Antiepileptic Drugs

Summary: Virtually all of the major antiepileptic drugs (AEDs) can cause hepatotoxicity, although fatal hepatic reactions are rare. The mechanisms, incidences, and risk profiles for such reactions differ from drug to drug. With carbamazepine and phenytoin, hepatotoxicity may be due to drug hypersensitivity. Although the profiles of patients at risk have not been well-defined for these two antiepileptic drugs, it would appear from reports in the literature that older adolescents and adults are at higher risk than children of developing serious or fatal hepatotoxicity. Once hepatotoxicity develops, mortality rates are 10–38% with phenytoin and 25% for carbamazepine. The risk profile for valproate fatal hepatotoxicity has been more clearly defined. Those at primary risk of fatal hepatic dysfunction are children under the age of 2 years who are receiving multiple anticonvulsants and also have significant medical problems in addition to severe epilepsy. The risk is considerably lower for patients over the age of 2 years on valproate monotherapy. In contrast to the risk profile with other AEDs, adults receiving valproate as monotherapy have the lowest risk of hepatotoxicity. Fatal hepatic dysfunction coincident with valproate may be the result of aberrant drug metabolism. Concomitant use of AEDs that induce microsomal P450 enzymes (e.g., phenytoin and phenobarbital) may enhance the production of a toxic metabolite, and hence the greater risk of hepatotoxicity with polypharmacy.     

Vascular Malformations and Epilepsy: Clinical Considerations and Basic Mechanisms

Summary: Vascular malformations (VMs) are associated with epilepsy. The natural history of the various VMs, clinical presentation, and tendency to provoke epilepsy determine treatment strategies. Investigations have probed the mechanisms of epileptogenesis associated with these lesions. Electrophysiologic changes are associated with epileptogenic cortex adjacent to VMs. Putative pathophysiologic mechanisms of epileptogenesis include neuronal cell loss, glial proliferation and abnormal glial physiology, altered neurotransmitter levels, free radical formation, and aberrant second messenger physiology.     

Neonatal Seizures: Problems in Diagnosis and Classification

Summary: The clinical identification of neonatal seizures is critical for the recognition of brain dysfunction; however, diagnosis is often difficult because of the poorly organized and varied nature of these behaviors. Current classification systems are limited in their ability to communicate motor, autonomic, and electroencephalo-graphic features of seizures precisely and to provide a basis for uniform effective diagnosis, therapy, and determination of prognosis. Recent investigations of neonates, utilizing bedside electroencephalographic/polygraphic/ video monitoring techniques, have provided the basis for improved diagnosis and classification of seizures in the newborn. These studies have demonstrated that not all clinical phenomena currently considered to be seizures require electrocortical epileptiform activity for their initiation or elaboration. In addition, the specific clinical character of the phenomena considered to be seizures, the clinical state of the infant, and the character of the EEG indicate the probable pathophysiological mechanisms involved and suggest probable etiologies, prognosis, and therapy. Similarities between animal models that demonstrate reflex physiology and neonates with motor automatisms and tonic posturing suggest that these clinical behaviors may not be epileptic in origin but, rather, primitive movements of progression and posture mediated by brainstem mechanisms. Although not all clinical behaviors currently considered to be neonatal seizures may have similar pathophysiological mechanisms, they are clinically significant because they all indicate brain dysfunction.     

Valproate Monotherapy in the Management of Generalized and Partial Seizures

Summary: For decades, therapeutic tradition has promoted the concept of polypharmacy in the management of epilepsy. In recent years, however, studies have shown that, for most patients, monotherapy can provide comparable or better seizure control than administration of multiple anticonvulsants, while diminishing the potential for adverse reactions, drug interactions, and poor compliance. Valproate is an important monotherapeutic agent that is highly effective in the control of idiopathic primary and secondarily generalized epilepsies, and partial seizures that do not generalize. Comparative studies have found that valproate is at least as effective as phenytoin and carbamazepine in the treatment of generalized and partial seizures. Given the similar efficacy, other factors such as pharmacokinetics and side effects may therefore determine anticonvulsant selection for monotherapy.     

Carbamazepine Efficacy and Utilization in Children

Summary: Carbamazepine is effective for preventing partial and generalized tonic-clonic seizures in children. Although absence epilepsies are more common in children than adults, an estimated 80% of children with epilepsy have seizure types or epilepsies that are potentially responsive to carbamazepine. The differential diagnosis of ictal staring is an especially important issue in children because absence and atypical absence seizures are more prevalent in children than adults. Age-related pharmacokinetic differences and drug interactions are major considerations in children. On average, children have higher clearance rates of carbamazepine, shorter half-lives, and higher ratios of carbamazepine-10, 11-epoxide to carbamazepine than adults. In addition, children with severe epilepsy are more likely to require multiple-drug therapy, which can lead to complex drug interactions. When carbamazepine is administered along with valproate, drug protein binding interactions can cause intermittent side effects.     

Un nouveau cadre conceptuel de travail pour la psychiatrie

In an attempt to place psychiatric thinking and the training of future psychiatrists more centrally into the context of modern biology, the author outlines the beginnings of a new intellectual framework for psychiatry that derives from current biological thinking about the relationship of mind to brain. The purpose of this framework is twofold. First, it is designed to emphasize that the professional requirements for future psychiatrists will demand a greater knowledge of the structure and functioning of the brain than is currently available in most training programs. Second, it is designed to illustrate that the unique domain which psychiatry occupies within academic medicine, the analysis of the interaction between social and biological determinants of behavior, can best be studied by also having a full understanding of the biological components of behavior.     

Special Pharmacokinetic Considerations in Children

Summary: Pediatric patients have greater degrees of pharmacokinetic variability and unpredictability than adults. This variability results from the effects of pharmacogenetics, age and growth, prior and current comedication, and disease. Newborns with seizures have the least predictable dosage requirements, and their needs change as drug-eliminating mechanisms mature in the neonatal period. Infants have the highest relative capacities to eliminate antiepileptics of any age group and require the largest relative doses. In addition to age-related trends, children demonstrate the same drug-specific, pharmacokinetic phenomena that adults do, including nonlinear phenytoin elimination, nonlinear valproate binding, and autoinduction of carbamazepine. Intercurrent illness and drug interactions further modify the age-related pharmacokinetic patterns in children and make dosage requirements even more unpredictable. Recent studies have shown that febrile illness can affect drug elimination, sometimes decreasing drug levels by 50% or more. Intermittent treatment with benzodiazepines administered either orally or rectally can be an important adjunct and help minimize this type of problem for children with marginally controlled epilepsy. Intermittent benzodiazepines are also helpful for children who have febrile seizures and who need only occasional antiepileptic protection.