雷公藤内酯醇对癫痫大鼠电压门控钾通道kv1.1表达的影响

目的 探讨雷公藤内酯醇(TL)对癫痫大鼠神经的保护作用及其机制.方法 60只SD大鼠分成对照组、模型组、雷公藤组,每组各20只.雷公藤组大鼠腹腔注射TL(每日15μg/kg),模型组大鼠腹腔注射等量生理盐水,注射7d后,雷公藤组与模型组通过颈内皮下注射海人酸(KA)致痫,对照组则颈内皮下注射等量的生理盐水.用免疫组化和Western blot方法检测大鼠海马CA3区瞬时外向钾离子通道kv1.1蛋白表达.结果 模型组大鼠海马CA3区kv1.1蛋白表达水平低于对照组(P＜0.05);雷公藤组海马CA3区kv1.1蛋白表达高于模型组(P＜0.05);雷公藤组与对照组大鼠kv1.1蛋白表达水平无明显差异(P＞0.05).结论 TL对KA致痫大鼠神经元有保护作用,其作用的发挥可能与TL可增加海马CA3区神经元kv1.1的表达有关.     

电压门控性钠通道mRNA在海人酸致痫大鼠表达的研究

目的:探讨电压门控性钠通道在癫痫发病机制中的作用。方法:采用海人酸颞叶癫痫模型,运用原位杂交技术检测不同时点海马DG区、CA1区、CA2区和CA3区SCN2A、SCN3A mRNA的表达。结果:SCN2A和SCN3AmRNA均表达于海马的DG区、CA1、CA2、CA3区。海人酸致痫后3小时在海马各区表达开始增强,6小时明显增强(P<0.05),12小时达到高峰(P<0.01),24小时开始下降,48小时恢复至正常水平。结论:电压门控性钠通道SCN2AmRNA和SCN3A mRNA的表达增加可能参与了颞叶癫痫急性期的发病。     

电压门控钾通道Kv1.1在戊四唑致痫大鼠中的表达

目的通过检测戊四唑(PTZ)致痫大鼠钾通道Kv1.1蛋白表达,探讨钾通道Kv1.1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Westernblot法检测大鼠钾通道Kv1.1蛋白。结果实验组大鼠海马区钾通道Kv1.1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),但3个时间段之间钾通道Kv1.1蛋白表达水平差异无统计学意义(P0.05)。结论大鼠海马区钾通道Kv1.1表达的减少与全身强直阵挛发作大鼠癫痫发病密切相关。     

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。     

电压门控钠离子通道与药物抗性癫痫

癫痫患者中约三分之一无法以现有抗癫痫药物(anti-epilep-tic drugs,AEDs)有效控制发作,称为药物抗性癫痫(drug-resist-ance epilepsy,DRE).药物抗性癫痫的主要形成机制之一是AEDs的作用靶点发生结构或功能改变,AEDs无法结合预定靶点抑制神经元过度放电,无法控制癫痫发作.电压门控性钠离子通道(voltage-gated sodium channel,VGSC)主要在可兴奋性细胞中表达,其结构和功能异常可引起神经元的膜兴奋性改变,参与癫痫的发病机制.     

电压门控性钠通道β亚基与特发性癫痫

特发性癫痫与电压门控性钠通道的功能改变密切相关,β亚基作为电压门控性钠通道重要的功能调节亚基已经开始受到许多研究学者的重视。本文概述了电压门控性钠通道β亚基的基因克隆与定位、亚基分型、分子结构以及与特发性癫痫相关的功能研究现状,并介绍了由于β亚基改变而引发的特发性癫痫的相关症状。     

A型钾通道Kv1.4在致痫大鼠海马区的表达

目的 通过检测A型钾通道Kv1.4在戊四唑(PTZ)致痈大鼠海马CA1、CA3及齿状同区的表达变化,探讨A型钾通道与癫痫发病的关系.方法 SD大鼠40只,随机分为对照组、致痫后1h、24h、72h组,每组各10只.腹腔注射PTZ制备大鼠癫痫模型,应用免疫组化及Western Blot技术检测Kv1.4在各时间段海马CA1、CA3及齿状回区的蛋白表达.结果 致痫组大鼠海马区Kv1.4蛋白水平在致痫后1h、24h、72h 3个时间段均明显低于正常组(P<0.05);各致痫组之间Kv1.4蛋白水平均无明显差异(P>0.05).结论 (1)A型钾通道Kv1.4在SD大鼠海马中广泛分布.表达丰富,以轴突处最为明显.(2)大鼠癫痫模型海马区A型钾通道Kv1.4蛋白表达减少,提示Kv1.4的表达下调可能与癫痫的发病相关.     

伽玛刀照射对癫痫大鼠海马氨基酸递质及超微结构的影响

目的研究伽玛刀照射对癫痫大鼠的放射生物学作用,探讨伽玛刀治疗癫痫的作用机制。方法利用海人酸制备癫痫大鼠模型50只(癫痫组25只,伽玛刀照射组25只),对照组为正常大鼠25只(注射生理盐水),观察各组行为学改变;采集伽玛刀照射后1d、1w、2w、4w、8w后各组大鼠海马细胞外液,并测定Glu和GABA含量及观察海马超微结构变化。结果 1w后,照射组和癫痫组GABA含量均低于对照组;2w后,照射组和癫痫组Glu含量均高于对照组;4w后,照射组Glu含量低于癫痫组,GABA含量高于癫痫组;差异均具有统计学意义(P<0.05)。海马组织超微结构显示照射组早中期与癫痫组基本一致,其晚期部分结构恢复,线粒体修复较为明显。结论伽玛刀照射可调节海马中Glu与GABA以达到新的平衡,并在癫痫发作引起的病理性损伤修复中起到重要作用。     

MK-801对海人酸诱导大鼠癫痫发作脑组织HSP70表达的影响

目的：癫痫是一种神经系统常见病，其发病机制较为复杂。近年来研究发现，在许多化学药物及电刺激所造成的癫痫发作模型中均可诱导热休克蛋白７０（ＨＳＰ７０）的合成。表明ＨＳＰ７０与癫痫发作有着密切的关系。方法：选择针对ＨＳＰ７０的鼠单克隆特异抗体，采用免疫组织化学法对海人酸（ＫＡ）诱导ＳＤ大鼠癫痫发作后ＨＳＰ７０表达进行了观察，并就ＭＫ－８０１对ＨＳＰ７０表达的影响做了初步探讨。结果：ＫＡ注射后２４ｈ，ＨＳＰ７０免疫反应阳性细胞主要分布在边缘结构，多数皮质区也有表达；预先用ＭＫ－８０１后，全部大鼠均没有出现明显惊厥行为，多数皮质区及丘脑区ＨＳＰ７０表达消失，但在齿状回，ＣＡ３区仍有高表达。结论：ＭＫ－８０１具有抗惊厥及神经保护作用，其对某些脑区ＨＳＰ７０表达的抑制作用显示出它对ＫＡ引起的同步性痫样放电活动的破坏     

电压门控性钾通道抗体病及其治疗

越来越多的证据显示,某些抗体如电压门控性钙通道及钾通道等抗体病与神经系统疾病密切相关,并且这些疾病大多免疫抑制治疗有效。本文就电压门控性钾通道抗体病:神经性肌强直、边缘叶脑炎及莫旺综合征的研究与治疗进展作一介绍     

Age-related changes in the distribution of Kv1.1 and Kv1.2 channel subunits in the rat cerebellum

We have revealed age-related changes in the expression patterns of Kv1.1 and Kv1.2 in the rat cerebellum for the first time. In the aged rat, immunoreactivity for Kv1.1 was increased in the cell bodies of Purkinje cells, while the staining intensity was significantly decreased in the granule cells. The cell bodies of cerebellar output neurons showed strong Kv1.1 immunoreactivity in the nucleus medialis, interpositus and lateralis of the aged rat. Kv1.2 immunoreactivity was found in some interneurons with their processes in this region of the aged rat. Image analysis demonstrated that immunoreactivities for Kv1.1 and Kv1.2 were increased specifically in the cell bodies of cerebellar output neurons of the aged rat. This study may provide useful data for future investigations on the channels that cause brain diseases and age-related disorders.     

Developmental change in expression and subcellular localization of two shaker-related potassium channel proteins (Kv1.1 and Kv1.2) in the chick tangential vestibular nucleus

The chick tangential nucleus is a major avian vestibular nucleus whose principal cells participate in two vestibular reflexes. Intracellular recordings have shown that the principal cells acquire their mature firing pattern gradually during development. At embryonic day 16 (E16), most principal cells fire a single spike, whereas shortly after hatching (H) the vast majority fire repetitively on depolarization. The transition in firing pattern was likely due in part to a downregulation of a low-threshold, sustained, dendrotoxin-sensitive (DTX) potassium current, I(DS). Since the DTX-sensitive potassium channel subunits Kv1.1 and Kv1.2 generate sustained currents, in the present study we applied fluorescence immunocytochemistry and confocal microscopy to characterize their developmental expression at E16, H1, and H9. At E16, both Kv1.1 and Kv1.2 staining were confined to the principal cell bodies. Immunolabeling decreased significantly for both proteins at H1, and more so by H9. Double-labeling with a monoclonal antibody against microtubule-associated protein 2 (MAP2) in hatchlings showed that some Kv1.1 remained as clusters within the cell body, at the base of the dendrites, and in the axon initial segment. In hatchlings, Kv1.2 staining decreased in the cell bodies and simultaneously appeared in the neuropil, colocalized with biocytin-labeled primary vestibular fibers and vestibular "spoon" terminals. Also, double-labeling with synaptotagmin showed that Kv1.2 colocalized with many nonvestibular terminals surrounding the principal cell bodies. These results identified developmental decreases in the staining of these two potassium channel protein subunits and changes in their subcellular localization corresponding to the downregulation of I(DS) defined electrophysiologically around hatching. Accordingly, both of these protein subunits could be involved in regulating excitability of the principal cells.     

Developmental regulation and adult maintenance of potassium channel proteins (Kv 1.1 and Kv 1.2) in the cochlear nucleus of the rat

The development and maintenance of the adult expression and distribution of Kv 1.1 and Kv 1.2, two voltage-dependent potassium channel subunits, were investigated in the anteroventral cochlear nucleus (AVCN) of the rat. Both Kv 1.1 and Kv 1.2 were found in AVCN neuronal cell bodies at birth, as detected by in situ hybridization and immunocytochemistry. However, Kv 1.1 and Kv 1.2 were not seen in axons until the end of the third postnatal week. From postnatal day 21 through adulthood, labeling for both potassium channels was in axonal processes, whereas the number of cell bodies labeled for Kv 1.1 decreased and there were no cell bodies labeled for Kv 1.2. Therefore, these two potassium channel proteins are targeted to their final subcellular destinations in axons well after hearing onset. Once the adult distribution pattern of Kv 1.1 and Kv 1.2 is attained, its maintenance does not depend on signals from auditory nerve synapses. Eliminating auditory nerve input to the cochlear nucleus by means of bilateral cochleotomy did not change Kv 1.1 or Kv 1.2 expression or distribution, as seen by in situ hybridization, immunocytochemistry and Western blot. Thus, normal excitatory synaptic input in adult animals is not a requirement to regulate the expression and cellular and subcellular distribution of these potassium channel proteins.     

A型钾通道Kv4.1在戊四唑致痫大鼠海马区的表达

目的通过检测A型钾通道Kv4.1在戊四唑(PTZ)致痫大鼠海马CA1、CA3及齿状回区的表达变化,探讨A型钾通道在癫痫发病机制中的作用。方法 SD大鼠40只,随机分为正常组、致痫后1 h、24 h、72 h组。腹腔注射PTZ制备大鼠癫痫模型,应用免疫组化及Western Blot技术检测Kv4.1在各时间段海马CA1、CA3及齿状回区的蛋白表达情况。结果致痫组大鼠海马区Kv4.1蛋白表达水平在致痫后1 h、24 h、72 h三个时间段均明显高于正常组(P<0.05);各致痫组之间Kv4.1蛋白表达水平无明显差异(P>0.05)。结论大鼠癫痫模型海马区A型钾通道Kv4.1蛋白表达增多,Kv4.1的表达上调可能在癫痫的发生中起作用。     

Expression and localization of voltage dependent potassium channel Kv4.2 in epilepsy associated focal lesions

An increasing number of observations suggest an important role for voltage-gated potassium (Kv) channels in epilepsy. We studied the cell-specific distribution of Kv4.2, phosphorylated (p) Kv4.2 and the Kv4.2 interacting protein NCS-1 using immunocytochemistry in different epilepsy-associated focal lesions. In hippocampal sclerosis (HS), Kv4.2 and pKv4.2 immunoreactivity (IR) was reduced in the neuropil in regions with prominent neuronal cell loss. In both HS and malformations of cortical development (MCD), intense labeling was found in neuronal somata, but not in dendrites. Strong NCS-1 IR was observed in neurons in all lesion types. Western blot analysis demonstrated an increase of total Kv4.2 in all lesions and activation of the ERK pathway in HS and ganglioglioma. These findings indicate that Kv4.2 is expressed in both neuronal and glial cells and its regulation may involve potassium channel interacting proteins, alterations in the subcellular localization of the channel, as well as phosphorylation-mediated posttranslational modifications.     

Transient expression of A-type K channel alpha subunits Kv4.2 and Kv4.3 in rat spinal neurons during development

A-type K(+) currents (I(A)s) have been detected from the ventral horn neurons in rat spinal cord during embryonic day (E) 14 to postnatal day (P) 8 but not in adulthood. It is not known which types of neurons and which A-type K(+) channel alpha subunits express the I(A)s and what the possible function might be. Here, we examined the expression of two A-type K(+) channel alpha subunits, Kv4.2 and Kv4.3, in rat spinal cord at various developmental stages by immunohistochemistry. We found a transient expression of Kv4.2 in somatic motoneurons during E13.5-P8 with a peak around E17.5, which coincides temporally with the natural selection of motoneurons. Transient expression of Kv4.2 and Kv4.3 was also observed in the intermediate gray (IG) interneurons. During E19.5-P14, some IG interneurons express Kv4.2, some express Kv4.3 and a subset co-express Kv4.2 and Kv4.3. Peak expression of Kv4.2 and Kv4.3 in the IG interneurons was detected around P1, which coincides temporally with the developmental selection of IG interneurons. In contrast to the I(A)-expressing subunits Kv4.2 and Kv4.3, a delayed-rectifier K(+) channel alpha subunit Kv1.6 is persistently expressed in somatic motoneurons and IG interneurons. Together, these data support the hypothesis that expression of I(A)s may protect I(A)-expressing somatic motoneurons, and possibly also IG interneurons, from naturally occurring cell death during developmental selection.     

Kv1.2-immunoreactive primary sensory neurons in the rat trigeminal ganglion

Immunohistochemistry for Kv1.2, a subunit of voltage-gated K(+) channels, was performed on the trigeminal ganglion (TG). Immunoreactivity (ir) was detected in half (48%) the TG neurons. These neurons were mostly medium-sized to large (range 137.6-2664.8 microm(2), mean+/-S.D. 892.6+/-413.3 microm(2)). A double immunofluorescence method also revealed co-expression of Kv1.2 and parvalbumin. Half (54%) the Kv1.2-immunoreactive (ir) neurons exhibited parvalbumin-ir, and parvalbumin-ir neurons mostly showed Kv1.2-ir (95%). Kv1.2-ir neurons which co-expressed CGRP-ir were rare in this ganglion. Some 40% of TG neurons retrogradely labeled from the facial skin exhibited Kv1.2-ir, whereas ir was detected in 16% of those labeled from the tooth pulp. The present study indicates that Kv1.2-ir TG neurons include low-threshold mechanoreceptors and nociceptors which innervate the facial skin and tooth pulp, respectively.     

Abnormal axonal physiology is associated with altered expression and distribution of Kv1.1 and Kv1.2 K+ channels after chronic spinal cord injury

Dysfunction of surviving axons which traverse the site of spinal cord injury (SCI) has been linked to altered sensitivity to the K+ channel blocker 4-aminopyridine (4-AP) and appears to contribute to post-traumatic neurological deficits although the underlying mechanisms remain unclear. In this study, sucrose gap electrophysiology in isolated dorsal column strips, Western blotting and confocal immunofluorescence microscopy were used to identify the K+ channels associated with axonal dysfunction after chronic (6-8 weeks postinjury) clip compresssion SCI of the thoracic cord at T7 in rats. The K+ channel blockers 4-AP (200 microM, 1 mM and 10 mM) and alpha-dendrotoxin (alpha-DTX, 500 nM) resulted in a significant relative increase in the amplitude and area of compound action potentials (CAP) recorded from chronically injured dorsal column axons in comparison with control noninjured preparations. In contrast, TEA (10 mM) and CsCl (2 mM) had similar effects on injured and control spinal cord axons. Western blotting and quantitative immunofluorescence microscopy showed increased expression of Kv1.1 and Kv1.2 K+ channel proteins on spinal cord axons following injury. In addition, Kv1.1 and Kv1.2 showed a dispersed staining pattern along injured axons in contrast to a paired juxtaparanodal localization in uninjured spinal cord axons. Furthermore, labelled alpha-DTX colocalized with Kv1.1 and Kv1.2 along axons. These findings suggest a novel mechanism of axonal dysfunction after SCI whereby an increased 4-AP- and alpha-DTX-sensitive K+ conductance, mediated in part by increased Kv1.1 and Kv1.2 K+ channel expression, contributes to abnormal axonal physiology in surviving axons.     

缺氧抑制脑动脉Kv2.1通道作用机制的研究

目的探讨缺氧对脑动脉Kv通道抑制作用的机制是否由内源性15-羟二十碳四烯酸(15-HETE)介导。方法选取健康Wistar大鼠,通过酶法分离培养脑动脉和颈内动脉平滑肌细胞,分为正常对照组、缺氧组和去甲二氢愈创木酸(NDGA)缺氧组,正常对照组平滑肌细胞常规培养48h、缺氧组在缺氧箱内培养48h、NDGA缺氧组加入50μmol/L的NDGA后在缺氧箱内培养48h,使用RT-PCR和Western blotting技术检测大鼠脑动脉和颈内动脉平滑肌细胞上Kv2.1通道mRNA及蛋白质的表达情况。结果缺氧可以抑制大鼠脑动脉和颈内动脉平滑肌细胞上Kv2.1通道的表达,缺氧组与正常对照组相比,Kv2.1通道mRNA及蛋白质的表达下调(P<0.05);采用NDGA抑制15-脂氧化酶(15-LOX)后,导致脑动脉和颈内动脉平滑肌细胞上Kv2.1通道的表达上调,NDGA可以保护缺氧对于Kv通道的抑制,NDGA缺氧组与缺氧组相比,Kv2.1通道mRNA及蛋白质的表达明显上调(P<0.05)。结论缺氧可能通过内源性15-HETE发挥对Kv通道的抑制作用,15-HETE可能是影响脑动脉张力的重要中介因素。