皮质酮对原代培养的海马神经元及其[Ca2+]i和钙/钙调蛋白依赖性蛋白激酶Ⅱ表达的影响

目的:探讨皮质酮(CORT)对培养的海马神经元及其[Ca2 ]i和钙/钙调蛋白依赖性蛋白激酶Ⅱ(CaMKⅡ)表达的影响和可能的机制。方法:海马神经元被分为不同终浓度的CORT处理组、CORT MK-8 0 1或高浓度葡萄糖组。采用MTT法、流式细胞术、荧光标记和免疫细胞化学法,观察海马神经元活力、死亡方式[、Ca2 ]i和CaMKⅡ表达的变化规律。结果:1 0-6和1 0-5mol/L CORT组,其海马神经元的活力明显降低,分别以凋亡和坏死为主,并使海马神经元[Ca2 ]i显著升高;CaMKⅡ的表达明显减少。MK-8 0 1和高浓度葡萄糖均能拮抗1 0-6mol/L CORT对海马神经元的损伤作用。结论:在CORT的作用下,海马神经元发生凋亡和坏死;[Ca2 ]i升高可能既是海马神经元损伤的结果,又是引起其发生凋亡和CaMKⅡ表达下降的原因。     

急性给予ApoE4升高大鼠皮层神经元内静息[Ca~(2+)]i

目的研究载脂蛋白E(ApoE)对大鼠皮层神经元内游离钙离子水平[Ca2+]i的影响。方法用激光共聚焦显微镜(LSCM)和Fluo-3/AM荧光探针标记检测皮层神经元钙信号瞬间动态变化;用N-甲基-D-天冬氨酸(NMDA)受体阻断剂MK-801观察ApoE4对其影响。结果ApoE4可以呈时间及浓度依赖性升高神经元内静息[Ca2+]i(P<0.01或P<0.05),MK-801可以部分阻断ApoE4所致的静息[Ca2+]i升高(P<0.05或P<0.01);而ApoE3无影响。结论急性给予ApoE4能升高神经元内静息[Ca2+]i,NMDA受体的激活可能参与了ApoE4所致的胞内钙信号改变与其神经毒作用。     

皮质酮对原代培养的海马神经元及其Ca~(2+)/CaMKII的影响

在培养液中加入 10 7、10 6和 10 5浓度(mol/L)的皮质酮,采用MTT染色测定、Fura 2 /AM的荧光标记以及Western印迹的方法,观察了不同浓度的皮质酮作用下海马神经元形态学和细胞存活率的变化以及胞浆内游离钙离子浓度 [Ca2+ ]i和CaMKII表达的变化规律, 探讨其对原代培养的大鼠海马神经元及其Ca2+ /CaMKII的影响和可能的机制。结果发现: 10 6、10 5浓度的皮质酮对海马神经元的形态学影响较大,与对照组比较,细胞存活率明显降低; [Ca2+ ]i分别为 113. 1022±16. 9716、155.3794±20. 7727;CaMKII的表达也明显减少;三者的变化均显著 (P<0. 01 ),而 10 7浓度的皮质酮对上述指标影响不大 (P>0.05)。此外,相关性分析表明: [Ca2+ ]i和CaMKII的表达呈现负相关(P<0. 05)。以上结果提示,皮质酮对大鼠海马神经元的作用存在浓度依赖效应,浓度越高,对大鼠海马神经元的损伤越大,同时也验证了皮质酮是啮齿类动物的主要的应激激素。     

红景天苷对谷氨酸损伤海马神经元的保护作用

目的观察红景天苷(salidroside)对谷氨酸(Glu)损伤海马神经元的保护作用。方法胚鼠原代培养海马神经元,与不同浓度的红景天苷(10、20和40mg/L)共同孵育24h,加入125μmol/L谷氨酸损伤海马神经元15min。采用MTT法检测细胞活性;生化法测定培养液中乳酸脱氢酶(LDH)活力的变化;膜联蛋白-V-FITC(annexin V-FITC)和碘化吡啶(PI)双标染色以及Hoechst染色观察细胞凋亡情况;激光共焦显微镜观察细胞内Ca2 的变化。结果红景天苷可明显改善谷氨酸损伤后细胞的活性,抑制谷氨酸损伤后培养液中LDH的活力,降低谷氨酸引起的细胞凋亡率,降低谷氨酸损伤后细胞内的Ca2 浓度。结论红景天苷可明显拮抗谷氨酸损伤海马神经元的作用,其机制可能与抑制谷氨酸引起的Ca2 内流有关。     

马桑内酯对大鼠海马神经元细胞内游离钙离子浓度的影响

以荧光探针Fluo-3/AM为胞浆内游离钙特异性指示剂,通过激光扫描共聚焦显微镜动态监测急性分离SD大鼠(出生后7～14 d)海马神经元细胞内游离钙离子浓度([Ca2 ]i)的变化情况,以了解钙离子及钙通道在癫痫发病中的作用,探讨马桑内酯(coriaria lactone, CL)调节神经细胞内钙稳态的机理.实验发现,CL能使急性分离大鼠海马神经元[Ca2 ]i增加;nifedipine和低浓度NiCl2虽可延迟,但不能完全阻断这种作用.除促使电压依赖的T-、L-型钙离子通道开放外,CL尚可通过其他途径增加海马神经元内[Ca2 ]i,改变细胞兴奋性而致痫.     

4-氨基吡啶对皮层神经元细胞内游离钙的影响

目的研究4-氨基吡啶(4-AP)对体外培养的皮层神经元细胞内游离钙浓度([Ca2+]i)的影响,了解4-AP的药理学作用机制.方法荧光探针Fluo-3-AM标记体外培养的皮层神经细胞内游离钙后,用共聚焦显微镜观察记录4-AP及L型谷氨酸对小鼠原代培养的皮层神经元[Ca2+]i的影响.结果4-AP与谷氨酸均能提高[Ca2+]i,两者峰值与持续时间存在差异,共同作用于细胞时的上升曲线与单用谷氨酸时相仿.结论4-AP的药理作用机制可能与提高神经细胞[Ca2+]i有关,其中机制与兴奋性氨基酸的[Ca2+]i升高作用可能不同.     

离子型谷氨酸受体拮抗剂MK-801浓度与全脑缺血再灌注大鼠海马内源性神经干细胞的增殖**

背景：脑缺血再灌注后,过度释放的兴奋性氨基酸可通过N-甲基-D-天冬氨酸(NMDA)受体激活内源性神经干细胞,促使其增殖、分化,修复神经细胞,但同时也导致细胞内钙离子超载,引起神经细胞的损伤。 目的：观察NMDA受体拮抗剂MK-801浓度对脑缺血再灌注大鼠海马内源性神经干细胞增殖的影响。 方法：SD大鼠随机分为正常对照组、手术对照组及MK-801 0.2,0.4,0.6,0.8,1.0,1.2 mg/kg组。除正常对照组外,大鼠首先进行侧脑室插管,3 d后进行4条血管阻断方法制备大鼠全脑缺血再灌注模型。在模型制作前30 min按照不同浓度侧脑室注射MK-801。正常对照组和手术对照组侧脑室注射同剂量的生理盐水。免疫组织化学、RT-PCR技术检测各组脑海马nestin阳性细胞及其mRNA表达。 结果与结论：MK-801浓度在0.8 mg/kg以下时,用药组大鼠脑海马nestin mRNA及蛋白的表达与手术对照组差异无显著性意义(P > 0.05),呈现高表达;当MK-801浓度达到0.8 mg/kg时,与手术对照组相比,用药组大鼠脑海马nestin基因及蛋白的表达明显下降(P < 0.05),并随浓度的增高呈递减趋势。提示MK-801在浓度为0.6 mg/kg时,即可抑制钙超载保护神经元,又有良好的刺激神经干细胞增殖作用。 关键词：离子型谷氨酸受体拮抗剂;脑缺血;再灌注;神经干细胞;MK-801 doi:10.3969/j.issn.1673-8225.2012.06.012     

外源性锌对缺氧神经元的保护作用

目的:观察不同浓度外源性锌对神经元细胞内游离钙([Ca2 ]i)的影响,探讨低浓度外源性锌对于缺氧神经元保护作用的可能机制.方法:原代培养大鼠皮层神经元,加入不同浓度的外源性锌,激光扫描共聚焦显微镜检测神经元[Ca2 ]i的变化;建立细胞缺氧模型,检测10 μmol/L外源性锌对于缺氧神经元[Ca2 ]i和[Zn2 ]i的影响.结果:神经元的[Ca2 ]i,μmol/L锌组与对照相比无明显变化;100μmol/L锌组一过性升高,后降至对照水平;500 μmol/L锌组,明显高于对照水平.除正常加锌组外,其余各组神经元[Zn2 ]i和[Ca2 ]i均较正常对照组增高;但正常加锌组、缺氧加锌组神经元[Zn2 ]i和[Ca2 ]i均较缺氧组降低.结论:外源性锌对于神经元的作用随浓度而不同;10 μmol/L外源性锌维持了神经元的钙稳态,它对于缺氧神经元的保护作用,可能通过抑制钙超载实现.     

电刺激大鼠小脑顶核对下丘脑神经元放电活动的影响

目的:研究电刺激小脑顶核(fastigial nucleus,FN)及微电泳Glu(glutamic acid)、MK-801对大鼠下丘脑(hypothalamus,HT)神经元自发放电活动的影响,探讨电刺激FN治疗脑卒中大鼠的机制。方法:应用细胞外记录的方法,记录电刺激FN及微电泳药物对HT神经元放电的影响。结果:电刺激频率为20Hz(低频)时,50%的HT神经元的放电频率增高(P0.01),电刺激频率为100Hz(高频时),80%的HT神经元的放电频率增高(P0.01);Glu对HT有紧张性兴奋作用,其拮抗剂MK-801能够明显抑制Glu诱致的兴奋作用;70%HT神经元在微电泳MK-801的基础上进行100Hz电刺激FN时,高频刺激诱发的兴奋作用被明显削弱。结论:采用电刺激FN治疗脑卒中,其可能机制是通过Glu的兴奋作用来调节HT的异常活动。     

4-氨基吡啶对皮层神经元细胞内游离钙的影响

目的研究4-氨基吡啶(4-AP)对体外培养的皮层神经元细胞内游离钙浓度([Ca2+]i)的影响,了解4-AP的药理学作用机制.方法荧光探针Fluo-3-AM标记体外培养的皮层神经细胞内游离钙后,用共聚焦显微镜观察记录4-AP及L型谷氨酸对小鼠原代培养的皮层神经元[Ca2+]i的影响.结果 4-AP与谷氨酸均能提高[Ca2+]i,两者峰值与持续时间存在差异,共同作用于细胞时的上升曲线与单用谷氨酸时相仿.结论 4-AP的药理作用机制可能与提高神经细胞[Ca2+]i有关,其中机制与兴奋性氨基酸的[Ca2+]i升高作用可能不同.     

Involvement of the N-methyl-D-aspartate receptor in neuronal cell death induced by cytotoxic T cell-derived secretory granules.

The mechanisms underlying neurotoxicity mediated by cytotoxic T lymphocytes (CTL) and their secretory granule proteins perforin and granzymes remain unclear. We evaluated the possible role of the neurotransmitter glutamate in cell death observed in differentiated neurons exposed to CTL-derived granules. Excitotoxicity induced by excessive concentrations of extracellular glutamate is associated with a rise in intracellular calcium and can lead to generation of NO through the activation of glutamatergic N-methyl-D-aspartate (NMDA) receptors. Consistent with an involvement of glutamate, we found that cell death in mature cerebral granule cells was inhibited by 65-80% by two NMDA receptor blockers (MK-801 and D-2-amino-5-phosphonovaleric acid) or a NO synthase blocker (N(G)-nitro-L-arginine methylester). Furthermore, neurons treated with secretory granules responded with a biphasic rise in the intracellular calcium concentration ([Ca2+]i). Whereas MK-801 did not interfere with the immediate rise of [Ca2+]i, the second wave of calcium accumulation starting at 40 min was delayed by 20 min and reduced in amplitude in the presence of MK-801. In immature, NMDA receptor-negative neurons, MK-801 prevented neither the cytotoxicity nor the calcium influx observed 5 min after addition of cytotoxic granules. The demonstration that NMDA receptors and NO are involved in granule-mediated killing of mature neurons opens new avenues in the treatment of neuronal cell death in CTL-mediated diseases such as viral encephalitis.     

低氧与缺血诱导培养海马和皮质神经元钙应答反应的比较

目的：钙作为重要信使参与多种生理和病理代谢过程,而且在缺血性神经元损伤机制中起着重要的作用。本实验模拟脑缺血的病理生化改变,用激光扫描共聚焦显微镜（LSCM）和Fluo-3荧光探针标记技术,观察低氧缺血状态下体外培养的海马,皮质神经元内钙浓度的变化。方法：用100μmol/L氰化钠造成细胞低氧;100μmol/L氰化钠和3.5mmol/L碘醋酸盐模拟在体完全性脑缺血;1mmol/LL-谷氨酸模拟在体脑缺血时兴奋性氨基酸大量释放;无葡萄糖介质剥夺细胞能量代谢的底物。结果：低氧使海马神经元[Ca^2 ]i显著升高,但有两种不同的钙振荡现象,谷氨酸引起海马神经元[Ca^2 ]i持续升高,但峰值低于低氧组,缺血组未见[Ca^2 ]i大幅升高,葡萄糖缺如不引起[Ca^2 ]i升高,结论：低氧和谷氨酸引起的神经元损害是能量依赖性的,轻度酸中毒可阻止胞内Ca^ 升高,糖对神经元具有保护作用,但单纯无糖引起的神经元损害与Ca^2 超载机制无关。     

Impaired dopamine release and uptake in R6/1 Huntington's disease model mice

The aim of this study was to evaluate the intracellular cytosolic calcium concentration ([Ca(2+)](i)) changes induced by activation of ionotropic glutamate receptors in cultured hippocampal neurons after repeated brief episodes of hypoxia. To investigate what kinds of ionotropic glutamate receptors are involved we used specific agonists for AMPA- and NMDA-type glutamate receptors. Measurements of [Ca(2+)](i) in cultured hippocampal neurons were made by imaging Fura-2AM loaded hippocampal cells. In the rat hippocampal slice method, field potential measurements in CA1 pyramidal neurons were used. The main result of our study is that brief hypoxic episodes progressively depress the [Ca(2+)](i) increases induced by agonists of AMPA and NMDA glutamate receptors in cultured hippocampal neurons. An effectiveness of this depression is increased from the first hypoxic episode to the third one. Hypoxic preconditioning effect is observed during 10-20 min after termination of hypoxic episode and depends on [Ca(2+)](i) response amplitudes to agonists before hypoxia. In contrast to AMPA receptor activation, NMDA receptor activation before hypoxia induce the spontaneous [Ca(2+)](i) increase about 3 min after each hypoxic episode. These spontaneous [Ca(2+)](i) increases may be an indicator of the development of posthypoxic hyperexcitability in hippocampal neurons. Our results suggest that brief hypoxia-induced depression of the glutamate receptor-mediated [Ca(2+)](i) responses contributes to the development of rapid hypoxic preconditioning in hippocampal CA1 neurons.     

地塞米松对谷氨酸升高海马神经元胞内[Ca~(2+)]i作用的影响

为了研究谷氨酸致痫和人工合成的糖皮质激素地塞米松抑痫作用的细胞内机制,本文在ＥＰＣ ９ 光电联合检测系统上用Ｆｕｒａ ２ 阳离子检测法观察了地塞米松对谷氨酸引起的培养乳鼠海马神经细胞内［Ｃａ２＋ ］ｉ 的影响。结果：（１）谷氨酸引起海马神经元内［Ｃａ２＋ ］ｉ 显著升高,ＥＧＴＡ（５ ｍ ｍ ｏｌ／Ｌ）耗竭细胞外钙后,谷氨酸升钙作用消失,给予氯化钙（１ ｍ ｍ ｏｌ／Ｌ）后其升钙作用恢复：Ｖｅｒａｐａｍ ｉｌ（１０ μｍ ｏｌ／Ｌ）对谷氨酸升钙作用无明显的影响,ＭＫ ８０１（１０ μｍ ｏｌ／Ｌ,ＮＭ ＤＡ 受体特异性非竞争性阻断剂）可明显阻断谷氨酸的升钙作用。（２）地塞米松（１００ μｍ ｏｌ／Ｌ）作用２ ｈ 明显抑制了谷氨酸（２００ μｍ ｏｌ／Ｌ）的升钙作用,地塞米松（１００ μｍ ｏｌ／Ｌ）＋ 放线菌酮（１０ μｍ ｏｌ／Ｌ,蛋白合成抑制剂）共同作用２ ｈ,再加入谷氨酸,则地塞米松的抑制作用消失,地塞米松（１００ μｍ ｏｌ／Ｌ）作用２ ｍ ｉｎ 对谷氨酸（２００ μｍ ｏｌ／Ｌ）的升钙作用无明显影响。本实验结果提示,谷氨酸通过ＮＭ ＤＡ 受体介导的外钙内流升高了海马神经元胞内［Ｃａ２＋ ］ｉ,地塞米松可能通过基因组机制抑制了谷氨酸的这种升     

Selective vulnerability of hippocampal cornu ammonis 1 pyramidal cells to excitotoxic insult is associated with the expression of polyamine-sensitive N-methyl-d-asparate-type glutamate receptors

Excess glutamate release and stimulation of post-synaptic glutamatergic receptors have been implicated in the pathophysiology of many neurological diseases. The hippocampus, and the pyramidal cell layer of the cornu ammonus 1 (CA1) region in particular, has been noted for its selective sensitivity to excitotoxic insults. The current studies examined the role of N-methyl-d-aspartate (NMDA) receptor subunit composition and sensitivity to stimulatory effects of the polyamine spermidine, an allosteric modulator of NMDA NR2 subunit activity, in hippocampal CA1 region sensitivity to excitotoxic insult. Organotypic hippocampal slice cultures of 8 day-old neonatal rat were obtained and maintained in vitro for 5 days. At this time, immunohistochemical analysis of mature neuron density (NeuN); microtubule associated protein-2(a,b) density (MAP-2); and NMDA receptor NR1 and NR2B subunit density in the primary cell layers of the dentate gyrus (DG), CA3, and CA1 regions, was conducted. Further, autoradiographic analysis of NMDA receptor distribution and density (i.e. [¹²⁵I]MK-801 binding) and spermidine (100 μM)-potentiated [¹²⁵I]MK-801 binding in the primary cell layers of these regions was examined. A final series of studies examined effects of prolonged exposure to NMDA (0.1–10 μM) on neurodegeneration in the primary cell layers of the DG, CA3, and CA1 regions, in the absence and presence of spermidine (100 μM) or ifenprodil (100 μM), an allosteric inhibitor of NR2B polypeptide subunit activity. The pyramidal cell layer of the CA1 region demonstrated significantly greater density of mature neurons, MAP-2, NR1 and NR2B subunits, and [¹²⁵I]MK-801 binding than the CA3 region or DG. Twenty-four hour NMDA (10 μM) exposure produced marked neurodegeneration (∼350% of control cultures) in the CA1 pyramidal cell region that was significantly reduced by co-exposure to ifenprodil or dl-2-Amino-5-phosphonopentanoic acid (APV). The addition of spermidine significantly potentiated [¹²⁵I]MK-801 binding and neurodegeneration induced by exposure to a non-toxic concentration of NMDA, exclusively in the CA1 region. This neurodegeneration was markedly reduced with co-exposure to ifenprodil. These data suggest that selective sensitivity of the CA1 region to excitotoxic stimuli may be attributable to the density of mature neurons expressing polyamine-sensitive NR2B polypeptide subunits.     

MK-801 is neuroprotective in gerbils when administered during the post-ischaemic period

The neuroprotective effects of the non-competitive N-methyl-D-aspartate receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) have been evaluated in the gerbil hippocampus when the drug was administered i.p. at various times during and after a 5 min period of transient forebrain ischaemia, induced by bilateral common carotid artery occlusion. A single dose of 1, 3 or 10 mg/kg of MK-801 gave significant protection of hippocampal CA1 and CA2 pyramidal neurons when administered during the occlusion and up to 24 h following the period of ischaemia. A dose of 0.3 mg/kg was effective when administered during the occlusion period but gave no protection at 30 min or 2 h post-ischaemia. Experiments in which MK-801 was administered in repeated doses indicated that significant protection was achieved with 1 mg/kg of MK-801 repeated post-ischaemically and with 1 mg/kg MK-801 supplemented with repeated doses of 0.3 mg/kg of MK-801. However 0.3 mg/kg of MK-801 followed by repeated doses of 0.03 mg/kg administered post-ischaemically was not neuroprotective. These results indicate that MK-801 can protect hippocampal neurons from ischaemia-induced neuronal degeneration when it is administered up to 24 h after the insult. These data provide further evidence that therapeutic intervention in the post-ischaemic period can successfully prevent neurodegenerative events, and that the delayed degeneration of hippocampal neurons following an ischaemic insult occurs by an N-methyl-D-aspartate receptor-mediated process.     

Geometry of dendritic spines affects calcium dynamics in hippocampal neurons: theory and experiments.

The role of dendritic spine morphology in the regulation of the spatiotemporal distribution of free intracellular calcium concentration ([Ca2+]i) was examined in a unique axial-symmetrical model that focuses on spine-dendrite interactions, and the simulations of the model were compared with the behavior of real dendritic spines in cultured hippocampal neurons. A set of nonlinear differential equations describes the behavior of a spherical dendritic spine head, linked to a dendrite via a cylindrical spine neck. Mechanisms for handling of calcium (including internal stores, buffers, and efflux pathways) are placed in both the dendrites and spines. In response to a calcium surge, the magnitude and time course of the response in both the spine and the parent dendrite vary as a function of the length of the spine neck such that a short neck increases the magnitude of the response in the dendrite and speeds up the recovery in the spine head. The generality of the model, originally constructed for a case of release of calcium from stores, was tested in simulations of fast calcium influx through membrane channels and verified the impact of spine neck on calcium dynamics. Spatiotemporal distributions of [Ca2+]i, measured in individual dendritic spines of cultured hippocampal neurons injected with Calcium Green-1, were monitored with a confocal laser scanning microscope. Line scans of spines and dendrites at a <1-ms time resolution reveal simultaneous transient rises in [Ca2+]i in spines and their parent dendrites after application of caffeine or during spontaneous calcium transients associated with synaptic or action potential discharges. The magnitude of responses in the individual compartments, spine-dendrite disparity, and the temporal distribution of [Ca2+]i were different for spines with short and long necks, with the latter being more independent of the dendrite, in agreement with prediction of the model.     

Electrophysiology of glutamate neurotoxicity in vitro: induction of a calcium-dependent extended neuronal depolarization.

1. Physiological responses of hippocampal pyramidal neurons in primary culture to prolonged glutamate (GLU) exposure (500 microM in all experiments) were studied with the use of patch electrodes and whole-cell current-clamp recording techniques. In some experiments, perforated patch recordings were employed with electrodes containing the pore-forming antibiotic nystatin. 2. After washout of GLU after a 10-min exposure, pyramidal neurons remained depolarized by greater than or equal to 20 mV from rest for the duration of the recording (30 min to less than 4 h). This depolarization was accompanied by a 57.8% increase in membrane conductance and was termed an extended neuronal depolarization (END). The percentage of neurons in which END was induced varied with the duration of GLU exposure, with a 4-, 6-, 8-, 10-, and 20-min GLU exposure eliciting END in 12.5, 41.7, 81.8, 100, and 100% of neurons. END induction appeared to be an all-or-none phenomenon, because END levels did not differ when compared across GLU exposure times. 3. During the END, cells retained both the ability to fire action potentials and the ability to respond to GLU, appeared viable when examined anatomically, and still excluded vital dyes. This supports the conclusion that END is not a nonspecific consequence of cell death. Rather, END is a discrete physiological process triggered by prolonged GLU exposure. The results raise questions concerning the reversibility of END induction, i.e., can neurons be "rescued" once END is induced, or will these cells inevitably go on to die? 4. END induction was dependent on a rise in intracellular free calcium ([Ca]i). END was prevented by strong buffering of [Ca]i or by substitution of external Ba2+ for Ca2+. However, substitution of Mn2+ for Ca2+ still permitted END induction. In cells recorded with the perforated-patch technique, maintaining normal [Ca]i levels, END could be induced, but less readily than under unbuffered [Ca]i conditions. 5. END could not be induced by a 10- to 20-min current-clamp depolarization to 0 mV, nor by 10-min GLU application while the membrane potential was voltage clamped at rest in a solution containing 1 mM Mg2+. In addition, END induction by GLU could be blocked by application of MK-801 (10-30 microM) but not 6-cyano-7-nitroquinoxaline-2,3-dione [CNQX (100-200 microM)]. 6. The dependence of both delayed neuronal cell death and END induction on GLU exposure duration were similar.(ABSTRACT TRUNCATED AT 400 WORDS)     

Neuroprotective effects of carvedilol, a new antihypertensive, at the N-methyl-D-aspartate receptor.

Carvedilol's potent antioxidant activity could explain its protective action in brain ischemia, but may not apply to glutamate-induced excitotoxicity in cultured cerebellar granule cells, since glutamate neurotoxicity was not associated with the formation of lipid peroxidative products. Rather, carvedilol diminished the N-methyl-D-aspartate (NMDA)/glycine-induced increase in intracellular calcium ([Ca2+]i), lowering [Ca2+]i by a maximum of 66 +/- 5% (n = 8) with a 50% inhibitory concentration of 0.8 microM. Prior addition of 5 microM dihydropyridines did not shift the dose-response of carvedilol, but did significantly lower the NMDA/glycine-stimulated response to 64% of untreated (n = 8, P = 0.014). Inclusion of 5 microM carvedilol before the additions of NMDA/glycine prevented 85% of the increase in [Ca2+]i. Furthermore, carvedilol displaced 3[H]MK-801 binding to rat brain cortical membranes with a Kd of 29.4 +/- 2.2 microM (n = 6) and no selectively for the glutamate or glycine binding sites. These data therefore suggest that, in addition to its antihypertensive and anti-lipid peroxidative functions, carvedilol has neuroprotective activity as a calcium channel blocker and as a non-competitive inhibitor at the NMDA receptor.