氯化锂-匹罗卡品致癎大鼠的模型研究

目的:探讨氯化锂(LiCl)-匹罗卡品(PILO)致大鼠的行为学特征及海马病理改变。方法:建立改良的LiCl-PILO致大鼠模型,观察大鼠行为,尼氏染色观察不同时间点大鼠海马病理改变,FJB染色观察大鼠海马神经元及其轴突、树突变性。结果:大鼠腹腔注射LiCl-PILO后,癫持续状态(SE)诱发成功率为92.5%,死亡率21.25%;尼氏染色结果显示,实验组大鼠海马神经元于SE后7d和60d缺失最明显,与对照组相比,齿状回颗粒细胞减少(P<0.05),CA1、CA3区锥体细胞和门区神经元均显著减少(P<0.01);FJB染色结果显示,实验组大鼠海马神经元于SE后7d和60d变性最明显,主要集中在CA1、CA3区锥体细胞层和门区,SE后7d海马腔隙-分子层亦可见变性的神经元轴突和树突。结论:改良后的LiCl-PILO致模型SE诱发成功率高,死亡率低,可基本复制人类颞叶癫的发作特点及病理改变;颞叶癫海马神经元的缺失可能是由于神经元的变性死亡。     

氯化锂-匹罗卡品致大鼠的模型研究

目的:探讨氯化锂(LiCl)-匹罗卡品(PILO)致大鼠的行为学特征及海马病理改变。方法:建立改良的LiCl-PILO致大鼠模型,观察大鼠行为,尼氏染色观察不同时间点大鼠海马病理改变,FJB染色观察大鼠海马神经元及其轴突、树突变性。结果:大鼠腹腔注射LiCl-PILO后,癫持续状态(SE)诱发成功率为92.5%,死亡率21.25%;尼氏染色结果显示,实验组大鼠海马神经元于SE后7d和60d缺失最明显,与对照组相比,齿状回颗粒细胞减少(P0.05),CA1、CA3区锥体细胞和门区神经元均显著减少(P0.01);FJB染色结果显示,实验组大鼠海马神经元于SE后7d和60d变性最明显,主要集中在CA1、CA3区锥体细胞层和门区,SE后7d海马腔隙-分子层亦可见变性的神经元轴突和树突。结论:改良后的LiCl-PILO致模型SE诱发成功率高,死亡率低,可基本复制人类颞叶癫的发作特点及病理改变;颞叶癫海马神经元的缺失可能是由于神经元的变性死亡。     

大鼠癫持续状态后miR-21、miR-29的表达变化

目的:探讨miR-21、miR-29在癫持续状态(SE)大鼠脑组织和外周血中表达的变化及意义。方法:SD大鼠30只,随机分为SE组20只和对照组10只,SE组建立氯化锂-匹罗卡品致大鼠SE模型,对照组以生理盐水代替匹罗卡品,应用RT-PCR检测2组大鼠脑组织和外周血miR-21、miR-29的表达。结果:与对照组比较,SE组大鼠海马区脑组织和外周血中miR-21表达均下降(P0.05),miR-29表达均升高(P0.05);miR-21在外周血中的表达显著高于脑组织(P0.01),miR-29在外周血中的表达稍低于脑组织(P0.05);结论:miR-21、miR-29可能参与SE后神经元凋亡的调控过程,外周血miR-21和miR-29变化可反映脑组织中的变化趋势。     

GIRK通道在匹罗卡品癫痫模型中的表达改变

目的：研究匹罗卡品诱导癫痫发作后不同时期G蛋白耦联内向整流钾通道（GIRK通道）在海马神经元细胞膜表面表达变化情况，并探索可能的调控机制。 方法：将成年雄性SD大鼠随机分为对照组和匹罗卡品造模组，利用致痫剂匹罗卡品诱导癫痫持续状态（Status epilepticus，SE）后，分别在急性期（24h）和慢性期（30d）取海马组织提取膜蛋白，利用Western Blot方法检测海马膜蛋白GIRK通道亚基蛋白GIRK1、GIRK2的表达改变，并同时检测海马组织PI3K/Akt通路激活情况。建立细胞癫痫模型，观察PI3K抑制剂渥曼青霉素和LY294002对海马神经元内向整流钾通道（Kir）电流的影响。 结果：匹罗卡品诱导大鼠SE发作后急性期细胞膜上GIRK1和GIRK2蛋白的表达较对照组相比显著下调（P<0.05）。海马膜蛋白中GIRK1的表达在大鼠SE发作后慢性期较对照组下降（P<0.05），而GIRK2蛋白表达量没有显著改变（P>0.05）。p-Akt（308）、Akt蛋白表达量的比值在匹罗卡品诱导大鼠SE发作后急性期与对照组相比显著上升（P<0.05），而在SE发作后慢性期无显著差异（P>0.05）。在细胞癫痫模型中，PI3K抑制剂渥曼青霉素和LY294002对痫样放电神经元的Kir电流未发现显著影响。 结论：在大鼠匹罗卡品诱导SE发作后急性期，海马中GIRK通道蛋白表达量下调， PI3K/Akt通路存在明显激活。癫痫急性期GIRK通道表达的调控机制尚需进一步研究。     

丹参多酚酸盐及神经肽Y干预后癫痫大鼠海马神经肽Y的表达变化

目的探讨丹参多酚酸盐及神经肽Y（neuropeptide-Y,NPY）干预注射对海马NPY的表达影响研究。方法采用匹鲁卡品（PILO）诱发癫痫模型,随机分为NPY干预组、丹参多酚酸盐干预组、盐水干预组和对照组,观察各组大鼠的行为学改变,用免疫组织化学法标记显示各组大鼠脑内海马NPY的表达变化。结果在海马门区、CA3区、CA1区有NPY阳性细胞表达,生理盐水及丹参多酚酸盐干预组可见颗粒细胞中NPY的异位表达。干预方法不同,海马NPY阳性细胞数表达不同,生理盐水（NS）、丹参多酚酸盐干预组NPY阳性细胞数高于对照组,差异有统计学意义（P〈0.05）;丹参多酚酸盐组高于生理盐水干预组,差异有统计学意义（P〈0.05）。结论 NPY有抗癫痫作用。丹参多酚酸盐可以明显降低急性癫痫痉挛性大鼠的发作等级和发作时间,可能通过NPY的表达增高来发挥治疗作用。     

氯化锂-匹罗卡品致大鼠急性癫痫模型

目的：建立氯化锂-匹罗卡品化学诱导的大鼠急性癫痫模型。方法：雄性Wistar大鼠55只，随机分为生理盐水对照组、地西泮组和致痫组。氯化锂腹腔注射后10～10h后给予匹罗卡品。结果：生理盐水对照组大鼠均为0级发作，地西泮组中8只大鼠为0级发作，2只出现Ⅰ级发作；致痫组均达到Ⅲ级以上的痫性发作，其中Ⅲ级2只(2／35)，Ⅳ级3只(3／35)，Ⅴ级30只(30／35)。致痫组大鼠在匹罗卡品腹注射后10～90min内全部出现急性痫性发作。结论：氯化锂-匹罗卡品诱导的大鼠急性癫痫模型具有制作方便、致痫成功率高和动物死亡率低等特点，具有同人类癫痫持续状态和颞叶癫痫相似的行为和脑电图改变。     

癫痫持续状态大鼠模型海马区miR-181b表达的变化及意义

目的：探讨癫痫持续状态（status epilepticus,SE）大鼠模型海马区miR-181b表达的变化及意义。方法随机将30只SD（Sprague-Dawley）大鼠分为实验组和对照组,各15只,实验组建立氯化锂-匹罗卡品致SE大鼠模型,对照组以生理盐水代替氯化锂-匹罗卡品,提取两组大鼠右侧海马区脑组织总miRNA和左侧海马组织总蛋白,采用Real-Time RT-PCR检测miR-181b表达及采用Western Blot方法检测caspase-3蛋白的表达变化。结果 SE组大鼠海马区脑组织miR-181b表达较对照组有明显下降（P〈0.05）, caspase-3蛋白表达明显上升（P〈0.05）。结论 miR-181b可能通过调控caspase-3蛋白的表达抑制SE后海马区神经元凋亡过程。     

应用2种方法建立大鼠癫痫模型对比研究

目的探讨应用杏仁核电点燃法与氯化锂-匹罗卡品法制备大鼠癫痫模型的造模情况及癫痫大鼠不同时期脑电图表现。方法雄性SD大鼠60只,随机分为电点燃组20只,化学点燃组20只,电点燃对照组10只和化学点燃对照组10只,电点燃组将电极植入大鼠右侧杏仁基底外侧核,7d后给予电刺激建立电点燃癫痫模型;电点燃对照组只植入电极,不给予电刺激;化学点燃组腹腔注射氯化锂-匹罗卡品建立化学点燃模型;化学点燃对照组腹腔注射生理盐水。电点燃组和化学点燃组大鼠癫痫模型制备成功后,采用苯巴比妥钠、苯妥英钠分别筛选出杏仁核电点燃敏感、耐药型癫痫大鼠和氯化锂-匹罗卡品敏感、耐药型癫痫大鼠。观察电点燃组和化学点燃组造模情况及癫痫大鼠不同时期脑电图表现。结果电点燃组大鼠癫痫模型成功点燃12只(60%),化学点燃组成功点燃18只(90%),化学点燃组成功率高于电点燃组(P0.05);化学点燃组慢性期发作时脑电图频率[(19.00±2.12)Hz]明显高于急性期发作时[(14.89±1.96)Hz]和电点燃组急、慢性期发作时[(13.38±1.60)、(14.50±2.38)Hz](P0.05),化学点燃组慢性期发作后脑电图频率[(15.89±3.69)Hz]和波幅[(139.73±43.70)μV]高于电点燃组急性期发作后[(12.75±1.03)Hz、(116.87±54.43)μV](P0.05);给药后,化学点燃组耐药型大鼠发作前[(15.33±1.63)Hz]、发作时[(19.32±6.02)Hz]和发作后[(14.38±1.51)Hz]频率均高于电点燃组敏感型大鼠[(12.17±1.17)、(12.67±1.21)、(11.63±0.92)Hz](P0.05);化学点燃组耐药型大鼠发作时波幅[(388.61±42.51)μV]高于敏感型大鼠[(242.82±23.81)μV]及电点燃组敏感型大鼠[(228.23±12.74)μV]和耐药型大鼠[(298.48±18.64)μV](P0.05)。结论与杏仁核电点燃法相比,氯化锂-匹罗卡品点燃成功率高,更适合实验及临床研究需要,2种不同类型癫痫大鼠不同时期脑电图均发生相应改变。     

奥卡西平单药及添加治疗部分性癫(癎)的疗效和安全性前瞻研究

目的:评估奥卡西平单药与添加治疗部分性癫患者的疗效、耐受性和安全性。方法:前瞻性对在同济医院癫诊疗中心就诊的67名部分性癫患者应用奥卡西平后进行临床随访观察,分为单药治疗组和添加治疗组,用药的前3个月及后3个月进行对比观察。结果:2组患者治疗前后发作频率减少的平均百分率有显著性差异(P=0.002)。单药治疗组与添加治疗组相比,前者用药前3个月发作完全控制的百分率、用药后3个月发作频率减少50%的百分率和发作完全控制的百分率(P=0.02,0.017,0.019)均明显低于后者。单药治疗组或添加治疗组自身用药的3个月及后3个月发作减少50%的百分率、发作减少75%的百分率、发作完全控制百分率均无明显差异(P>0.05)。治疗引起的不良反应发生率为19.40%(13/67),主要出现于用药的前3月。结论:奥卡西平是治疗部分性癫的一线药物,可用于新诊断或其他药物无法耐受和疗效不佳患者的单药或添加治疗。     

参蒲汤抗大鼠癫痫的实验研究

目的探讨参蒲汤抗癫痫的作用及机理。方法采用匹罗卡品制备边缘系统癫痫大鼠模型，50只大鼠随机分为中药对照组8只、空白对照组8只、预防中药组（中药＋匹罗卡品）8只、匹罗卡品模型组26只（再进一步分为中药治疗组和模型对照组），观察模型大鼠的惊厥发生率及各组大鼠的脑组织游离氨基酸含量。结果预防中药组的惊厥发生率为0，匹罗卡品模型组为88.5%，两组间差异有显著性意义（P〈0.05）；匹罗卡品组和预防中药组大鼠脑组织的谷氨酸含量高于空白对照组（P〈0.05～0.01）；中药治疗组和预防中药组的γ-氨基丁酸含量高于模型对照组（P〈0.05）。结论参蒲汤能有效预防和终止匹多罗品诱导的边缘性癫痫模型大鼠的惊厥发作，具有良好抗惊厥、抗癫痫作用。     

Use of verapamil as a potential P-glycoprotein inhibitor in a patient with refractory epilepsy

OBJECTIVE: To describe a patient in whom we used adjunctive verapamil therapy was used for its P-glycoprotein inhibitory effects. CASE SUMMARY: Verapamil was added to the antiepileptic drug regimen of a 24-year-old woman with intractable epilepsy. The average time interval between hospitalizations for complex partial status doubled. The addition of verapamil greatly improved overall seizure control and subjective quality of life in this pharmacoresistant patient. DISCUSSION: The overexpression of P-glycoprotein in the central nervous system may be one mechanism of pharmacoresistance in patients with epilepsy. The calcium-channel blocker verapamil is a known inhibitor of P-glycoprotein and may function to block P-glycoprotein-modulated efflux of antiepileptic drugs in the brain, thereby raising the intracellular concentration of antiepileptic drugs and ultimately decreasing seizure burden in patients with refractory epilepsy. CONCLUSIONS: Verapamil may offer pharmacoresistant patients hope of improved seizure control due to its potential P-glycoprotein inhibitory effects.     

Attenuation of Verapamil-induced Myocardial Toxicity in an Ex-vivo Rat Model Using a Verapamil-specific Ovine Immunoglobin

OBJECTIVE: To determine whether an ovine verapamil-specific immunoglobin G (V-IgG) attenuates verapamil toxicity in an ex-vivo rat left ventricular papillary muscle model. METHODS: The authors dissected left ventricular papillary muscle strips from male Sprague-Dawley rats (350-410 g) and suspended them in an oxygen-perfused Tyrode buffer bath at 37.5 degrees C. Muscle strips equilibrated for 90 minutes under electrical stimulation of 1 Hz. Resting and developed tension (mg) were monitored continuously. A concentration-response trial was performed with verapamil concentrations ranging from 31 to 1,020 nM; 510 nM produced consistent reduction in developed tension. A trial of V-IgG was then conducted by administering the following treatments to papillary muscle strips in a randomized manner: V-IgG + 510 nM verapamil, nonspecific ovine IgG (N-IgG) + 510 nM verapamil (protein control), and 510 nM verapamil alone. Immunoglobin G was administered in equimolar concentrations to verapamil. Attenuation was expressed as inhibition of the verapamil-induced reduction of developed tension. RESULTS: The V-IgG comparative trial indicated the V-IgG + verapamil treatment had a mean reduction in developed tension of 14.1% (SD +/- 12.2) compared with 36.2% (SD +/- 14.9) for N-IgG + verapamil and 34.9% (SD +/- 8.1) for verapamil alone (p < 0.05). There was no difference between the two control groups. CONCLUSION: Verapamil-specific IgG attenuated verapamil-induced reduction of developed tension in an ex-vivo rat model.     

Verapamil suppresses the inhomogeneity of electrical remodeling in a canine long-term rapid atrial stimulation model

Verapamil is known to suppress shortening of the atrial effective refractory period (AERP) during relatively short-term atrial pacing, although the effect of a long-term stimulation model is unclear. The effect of verapamil on electrical remodeling was evaluated in a canine rapid atrial stimulation model. The right atrial appendage (RAA) was continuously paced (400 beats/min) for 2 weeks. Four pairs of electrodes were sutured at four atrial sites; the RAA, right atrium close to the inferior vena cava, Bachmann's bundle, and LA. AERP, AERP dispersion (AERPd), conduction time, and inducibility of AF were evaluated during the pacing phase and the recovery phase. The same protocol was performed under the administration of verapamil. In five control dogs, the AERP shortening was inhomogeneous and the shortening of the AERP was most prominent in the LA. AERPd increased during the rapid pacing phase by 5 +/- 2 ms, but recovered quickly in the recovery phase. The max AERPd was 46 +/- 4 ms in the control group and was larger than that in the verapamil group (31 +/- 3 ms, P = 0.001). At the LA site, the shortening of the AERP was decreased by verapamil administration (-19 +/- 3 vs -5 +/- 2 ms, P = 0.04). However, the AF inducibility was not significantly different between the two groups. The effect of verapamil on electrical remodeling was inhomogeneous, depending on the anatomic portion. As a result, AERPd widening during the rapid pacing phase was suppressed by verapamil, while the AF inducibility was unchanged.     

VAMP加逆转剂治疗难治性急性淋巴细胞白血病的体内、外研究

目的：观察VAMP方案加维拉帕米或环孢菌素A逆转治疗难治性ALL的疗效。方法：选择原发和继发耐药的ALL患者13例，采用VAMP方案加维拉帕米或环孢菌素A逆转体内、外研究。结果：13例ALL中7例达CR,3例达PR,3例无效。体外VAMP方案加VRP或CsA可有效逆转耐药。结论：采用VAMP方案加维拉帕米或环孢菌素A逆转治疗难治性ALL,可取得较好的疗效,为争取移植治疗创造了条件。     

全蝎醇提物与丙戊酸干预氯化锂-匹罗卡品慢性点燃模型大鼠癫痫发作的疗效比较

目的 探讨全蝎醇提物(Ethanol Extracts of Scorpion,EES)与丙戊酸(Valproic Acid,VPA)干预氯化锂一匹罗卡品(Lithium Chloride-Pilocarpine,Licl-Pilo)慢性点燃模型大鼠癫痫发作的疗效.方法 186只成年雄性大鼠除6只设为正常对照组外.其余均建立Licl-Pilo慢性点燃大鼠模型,遣模成功后分为模型组、VPA干预组、低 EES 剂量干预组(EESL干预组)、中EES剂量干预组(EESM干预组)和高EES剂量干预组(EESH干预组)各36只.正常对照组、模型组分别灌胃给予等容积的生理盐水,VPA干预组灌胃给予VPA溶液[浓度为120 ms/(kg·d)],EESL干预组、EESM干预组、EESH干预组灌胃给予EES溶液[浓度分别为290 ms/(kg·d)、580 nag/(kg·d)及1160 ms/(kg·d)],观察30天内致痫大鼠慢性癫痛自发性发作(spontaneous seizures,SRS)、发作级别、发作次数,并进行组问比较.结果 Licl-Pilo慢性点燃模型鼠的造模成功率为85.65%,各实验组SRS发生频率5～15次/周.治疗后,EESM干预组、EESH干预组和VPA干预组癫痫大鼠痫性发作级别及SRS发作次数与模型组比较差异有统计学意义(P<0.05);EESL干预组治疗效果不明显,与模型组比较差异无统计学意义(P>0.05).结论 一定剂量的EES能显著降低癫痫大鼠的自发性发作,且与疗效呈明显的量效关系,即剂量越大疗效越好,该结果 为EES的抗癫痫作用提供了行为学依据.     

异搏定和环孢素A对急性缺血性肾损伤后内皮素的影响

目的：观察内皮素（ＥＴ）在急性缺血性肾损伤后的变化及异搏定对肾的保护作用和环孢素Ａ（ＣｓＡ）肾毒性与ＥＴ之间的关系。方法：通过阻断大鼠肾动脉６０分钟造成急性缺血性肾功能衰竭（ＩＡＲＦ）模型，用放射免疫方法测定肾脏缺血再灌注１小时、６小时和２４小时血浆及肾组织中ＥＴ的变化；同时观察异搏定和ＣｓＡ对ＥＴ肾功能的影响。结果：ＩＡＲＦ时，血浆及肾组织中ＥＴ在３个时间点均有不同程度的升高；异搏定可降低ＩＡＲＦ大鼠血浆和肾组织中的ＥＴ含量；ＣｓＡ引起ＥＴ活性进一步升高，并加重肾损伤。结论：ＥＴ可能是导致ＩＡＲＦ的一个重要因子；异搏定能抑制ＩＡＲＦ时ＥＴ的分泌和释放而保护肾脏；ＣｓＡ的肾毒性可能与其诱导ＥＴ的分泌和释放有关。     

Intralipid Prolongs Survival in a Rat Model of Verapamil Toxicity

Objectives: Verapamil is a lipid-soluble calcium channel blocker with significant mortality in overdose. Previous investigators have demonstrated the benefit of lipid emulsion therapy in ameliorating toxicity from lipid-soluble agents. The authors investigated the effect of Intralipid treatment in a rat model of verapamil toxicity.
Methods: Thirty sedated Wistar rats were infused with verapamil at 37.5 mg/kg/h. Five minutes after the start of infusion, animals were treated with a bolus of either 12.4 mL/kg 20% Intralipid or 12.4 mL/kg 0.9% saline. Verapamil infusion was continued until the animals were killed. Respiratory rate, heart rate, and electrocardiography were sampled every 2.5 minutes throughout.
Results: Survival was prolonged in the Intralipid-treated group (44 ± 21 vs. 24 ± 9 minutes; p = 0.003). The median lethal dose was increased in the Intralipid group (25.7 mg/kg [95% confidence interval {CI} = 24.7 to 26.7] vs. 13.6 mg/kg [95% CI = 12.2 to 15.0]). A less marked decrease in heart rate was observed during verapamil infusion in the Intralipid-treated group (6.8 beats/min [95% CI = 8.3 to 5.2] for Intralipid vs. 10.7 beats/min [95% CI = 12.6 to 8.9] for saline; p = 0.001).
Conclusions: Intralipid treatment prolongs survival and doubles median lethal dose in a rat model of verapamil toxicity. The mechanism of action remains to be elucidated.     

异博定对视网膜缺血再灌注损伤ERG的影响

目的：建立视网膜缺血再灌注损伤模型，用视网膜电(流)图(ERG)观察异博定对视网膜ERG的影响。方法：通过结扎大鼠左颈总动脉1h，然后再灌注，同时向腹腔注射异博定，检测1h，6h，12h，24h，48h，72h视网膜ERG的变化。结果：再灌注6h后，异博定组视网膜的ERG与对照组相比，有明显的提高。结论：异博定可以促进缺血再灌注损伤视网膜ERGb波的恢复。     

生酮饮食添加治疗儿童难治性癫痫的临床效果

目的 探讨生酮饮食(KD)添加治疗儿童难治性癫痫的临床效果.方法 回顾性选取2013年6月至2019年4月于西安市儿童医院神经内科接受KD添加治疗的47例难治性癫痫患儿为研究对象.采用Engel分级评估KD添加治疗的临床疗效,同时评估KD添加治疗的终止原因及不良反应发生情况.结果 接受KD添加治疗的47例患儿中,KD添...     

Does Pilocarpine-Induced Epilepsy in Adult Rats Require Status epilepticus?

Pilocarpine-induced seizures in rats provide a widely animal model of temporal lobe epilepsy. Some evidences reported in the literature suggest that at least 1 h of status epilepticus (SE) is required to produce subsequent chronic phase, due to the SE-related acute neuronal damage. However, recent data seems to indicate that neuro-inflammation plays a crucial role in epileptogenesis, modulating secondarily a neuronal insult. For this reason, we decided to test the following hypotheses: a) whether pilocarpine-injected rats that did not develop SE can exhibit long-term chronic spontaneous recurrent seizures (SRS) and b) whether acute neurodegeneration is mandatory to obtain chronic epilepsy. Therefore, we compared animals injected with the same dose of pilocarpine that developed or did not SE, and saline treated rats. We used telemetric acquisition of EEG as long-term monitoring system to evaluate the occurrence of seizures in non-SE pilocarpineinjected animals. Furthermore, histology and MRI analysis were applied in order to detect neuronal injury and neuropathological signs. Our observations indicate that non-SE rats exhibit SRS almost 8 (+/22) months after pilocarpine-injection, independently to the absence of initial acute neuronal injury. This is the first time reported that pilocarpine injected rats without developing SE, can experience SRS after a long latency period resembling human pathology. Thus, we strongly emphasize the important meaning of including these animals to model human epileptogenesis in pilocarpine induced epilepsy.