【作者】 诸葛正兵；

【导师】 陈忠；

【作者基本信息】 浙江大学 ， 药理学， 2007， 硕士

【摘要】 癫痫是一种严重的慢性中枢神经系统疾病，其特征为反复发作的大脑神经元异常放电，导致暂时性大脑功能失调，临床表现为运动、感觉、意识、植物神经和精神等不同障碍。因其发作频繁，不仅影响患者的日常生活、学习和工作，还可能因在发作时遭遇危险而危及生命，给患者、家庭和社会带来极大负担。据WHO提供的数字，目前世界上大约有5000万癫痫患者，我国约有800万。癫痫的确切发病机制目前尚未完全阐明。神经元高度的兴奋性、过度的同步化活动、神经元兴奋和抑制之间平衡失调可能是癫痫发作最根本的原因。尽管传统的抗癫痫药物治疗对许多患者具有明显效果，但是仍有近25％的患者对药物治疗不敏感，并且抗癫痫药物的长期服用往往伴随着认知功能的损害(如记忆和注意缺陷)以及其他中枢神经系统副作用，如精神运动速度异常，嗜睡，衰弱和头昏。虽然外科手术也能有效地控制癫痫发作，但因其创伤大，常伴有严重的并发症，使其临床应用受到很大限制。因此，寻找和开发新的、安全有效的治疗手段，具有重要的临床价值。随着深部脑核团电刺激(deep brain stimulation，DBS)在治疗运动障碍性疾病中的广泛应用，其在治疗药物难治性癫痫中的作用也日益受到人们的重视。目前全世界约有5000多例药物难治性癫痫患者接受了DBS治疗，这些患者的年平均癫痫发作次数及发作严重程度均能得到有效的抑制或者控制。但是，由于电刺激参数及靶核团位置的选择不同，其产生作用的效果及方式都会有所不同，甚至有出现相反结果的报道。因此，我们的实验就从刺激参数(频率)和刺激靶点(中央区梨状皮质和乳头结节核E2区)这两方面入手，观察低频率电刺激不同核团对电点燃癫痫的作用，期望我们的研究能对临床DBS的应用提供一定的理论和实验依据。1．低频率电刺激中央区梨状皮质对大鼠杏仁核电点燃癫痫大发作及灶点转移的作用目的：研究低频率电刺激中央区梨状皮质对大鼠杏仁核电点燃癫痫大发作以及灶点转移的作用，并初步探讨其可能机制。方法：1．大鼠右侧基底旁侧杏仁核每天给予电点燃刺激使之达到完全点燃，随即在点燃刺激1秒后给予右侧或左侧中央区梨状皮质连续10 d的低频率电刺激(1 Hz，0.1 ms，50-150μA)，观察动物癫痫大发作几率、发作级别、后放电持续时间以及大发作持续时间的变化。2．大鼠右侧基底旁侧杏仁核(第一灶点)每天给予电点燃刺激后给予右侧中央区梨状皮质15 min的低频率电刺激，若干天后全部动物达到完全点燃，观察第一灶点的点燃对左侧杏仁核(第二灶点)后放电阈值以及点燃速度的变化以及低频率电刺激的效应。结果：1．低频率电刺激同侧中央区梨状皮质能显著降低大鼠的癫痫大发作几率和发作级别，缩短总的后放电持续时间和大发作持续时间；低频率电刺激对侧中央区梨状皮质则只能显著降低发作级别，而对大发作几率、后放电持续时间、大发作持续时间没有明显的作用。另一方面，低频率电刺激同侧中央区梨状皮质能显著增加大鼠的后放电阈值和大发作阈值，并进一步增加后放电阈值和大发作阈值之间的差值。2．在第一灶点达到充分点燃的情况下(连续3次5级)，低频率电刺激组的第二灶点后放电阈值下降幅度显著小于对照组，点燃形成所需的天数也显著比对照组延长；而在第一灶点未充分点燃的情况下(1-4级)，低频率电刺激组的第二灶点后放电阈值下降幅度和对照组相比没有明显差异，点燃形成速度也没有影响。结论：提示了低频率电刺激可以作为一种既能抑制癫痫棘波产生又能抑制癫痫棘波传播的有效手段，并且能够对癫痫灶点的转移产生抑制性作用。这也提示我们癫痫灶点外的核团，如中央区梨状皮质，可以作为深部脑核团电刺激治疗临床难治性癫痫的合理靶点来应用。2．单侧低频率电刺激乳头结节核E2区对大鼠杏仁核电点燃癫痫形成过程的作用目的：研究单侧低频率电刺激乳头结节核E2区对大鼠杏仁核电点燃癫痫形成过程的作用。方法：在大鼠右侧基底旁侧杏仁核每天给予点燃刺激，1 s后给予右侧或左侧乳头结节核E2区15 min的低频率电刺激(1 Hz，0.1 ms，300-400μA)，持续20 d直到全部动物达到完全点燃，观察动物每天的癫痫发作级别以及后放电持续时间。结果：低频率电刺激右侧或左侧乳头结节核E2区均能够促进杏仁核电点燃癫痫的发作级别和后放电持续时间，并且在刺激第7天到第13天具有显著性差异。结论：低频率电刺激乳头结节核E2区能促进大鼠杏仁核电点燃癫痫的形成过程，主要原因是显著减少了大鼠停留在3-5级所需的刺激天数。结合我们以前的结果，低频率电刺激脑内不同的核团对电点燃癫痫的作用不一致提示我们低频率电刺激具有靶点特异性，这也将为临床DBS治疗提供一定的理论和实验依据。更多还原

【Abstract】 Epilepsy is a serious chronic neurological disease, which is characterized by recurrent abnormal discharge of cerebral neurons and transient disfunction of brain. The clinical symptoms include a variety of abnormities in motor, sense, conscious state, vegetative nervous and psychology. According to WHO, 50 million persons worldwide, and 8 million in China have suffered from the diseases. The exact pathogenesis of epilepsy remains largely unclear. Excessive excitability, over-synchronized discharges and the imbalance between excitation and inhibition of cerebral neurons may contribute to seizures. With the currently available antiepileptic drugs, the disease in about 80% patients who have no serious injuries in the brain, especially those with fewer frequencies of seizures and normal interictal EEGs, are well controlled. By contrast, about 25% epileptic patients can’t get effective therapies using those drugs. This epilepsy is called drug-resistance or intractable epilepsy. Moreover, long application of those antiepileptic drugs often results in some adverse responses in psychological, neurological, splanchnic and skin system. Thus, developing new safe and effective antiepileptic methods is urgently expected.With the widely application of deep brain stimulation (DBS) in the treatment of motor disorder diseases, the roles of DBS in drug-resistance epilepsy become more and more important. So far, about 5000 epilepsy patients have received DBStreatment and the frequency and severity of epileptic attack among them were well-controlled. However, the fundamental questions of where to stimulate and what types of stimuli are most effective remain unclear. So our experiment was designed to observe the possible role of stimulation parameter (frequency) and stimulation targets (central piriform cortex and tuberomammillary nucleus E2 region) in kindling model of epilepsy, and expected to provide further evidence of DBS in the clinic.1. Low-frequency stimulation of central piriform cortex inhibits amygdaloid-kindled seizures and epileptic focus transfer in Sprague-Dawley rats Aim: The central piriform cortex (cPC) is considered to be critically involved in the generation and propagation of kindled seizures. Our previous study found that low-frequency stimulation (LFS) of the cPC inhibits the development process of amygdala kindling. In this study, we determined whether unilateral LFS of the cPC had an inhibitory effect on amygdaloid-kindled seizures and epileptic focus transfer in Sprague-Dawley rats. Methods: 1. When fully-kindled seizures were achieved by daily amygdala electrical stimulation (2 s train of 1 ms pulses at 60 Hz and 150-300 μA), LFS (15 min train of 0.1 ms pulses at 1 Hz and 50-150μA) was applied to the ipsilateral or contralateral cPC 1 s after cessation of kindling stimulation for 10 days.2. The ipsilateral cPC received LFS immediately 1 s after termination of once daily kindling stimulation in the right amygdala (primary site) until all animals reached kindled state. Then, left amygdala (secondary site) afterdischarge threshold and kindling rate were measured at different stages of primary site kindling. Results: 1. LFS of the ipsilateral cPC significantly decreased the incidence of generalized seizures and seizure stage, and shortened cumulative afterdischarge duration and cumulative generalized seizure duration. LFS of the contralateral cPC also significantly decreased the expression of seizure stage, but had no appreciable effect on the generalized seizure incidence, cumulative afterdischarge duration and cumulative generalized seizure duration. On the other hand, LFS of the ipsilateral cPC significantly increased the afterdischarge threshold and further increased the differences of current intensity between afterdischarge threshold and generalizedseizure threshold. 2. In the case of primary site reached kindled state (stage 5), the decrement of afterdischarge threshold at secondary site after LFS was fewer than control animals, and the number of days for this site reached kindled state also significantly increased; In the case that primary site did not reach kindled state (stage 1-4), there were no apparent differences of secondary site afterdischarge threshold and kindling rate between LFS and control groups. Conclusion: Our data suggest that LFS of the cPC may be an effective method of inhibiting kindled seizures by preventing both afterdischarge generation and propagation, and LFS also have an inhibitory effect on epileptic focus transfer in Sprague-Dawley rats. It provide further evidence that brain regions like the cPC, other than the seizure focus, can serve as targets for deep brain stimulation treatment of epilepsy.2. Unilateral low-frequency stimulation of histaminergic tuberomammillary nucleus ￡2 region facilitates basolateral amygdaloid-kindling in ratsAim: The tuberomammillary nucleus (TM), located in the posterior hypothalamus, is the sole seat of histaminergic neurons in the brain. In this study, we investigate whether low-frequency stimulation (LFS) of TM E2 region is involved in the acquisition of amygdaloid-kindling seizures. Methods: The ipsilateral or contralateral TM E2 region received LFS (15 min train of 0.1 ms pulses at 1 Hz and 300-400 μA) immediately 1 s after termination of once daily kindling stimulation (2 s train of 1 ms pulses at 60 Hz and 150-300 μA) in the right amygdala for twenty days. Results: LFS of either ipsilateral or contralateral TM E2 region significantly facilitated the progression of seizure stages and increased afterdischarge duration throughout the course of amygdaloid kindling. The marked facilitation induced by LFS on either side of the TM was predominantly due to the significant promotion of progression from stage 3 to stage 5 seizures. Conclusion: The significant facilitative effect of LFS on TM strongly indicated that the role of LFS in epilepsy is brain region specific and possibly another unknown mechanism that underlie classic explanation of LFS may be involved in this action.更多还原