Activations of nPKCepsilon and ERK1/2 were involved in oxygen-glucose deprivation-induced neuroprotection via NMDA receptors in hippocampal slices of mice

Accumulated reports have suggested that activation of protein kinase C (PKC) isoforms may involve the activation of extracellular signal-regulated kinases (ERKs) in the neuronal response to ischemic/hypoxic stimuli. We have previously demonstrated that the membrane translocation of novel PKC (nPKC) epsilon increased in the early phase of cerebral ischemic/hypoxic preconditioning of mice. In this study, we used Western blot analysis and propidium iodide stain to determine whether the activations of nPKCepsilon and ERKs were involved in oxygen-glucose deprivation (OGD)-induced neuroprotection via N-methyl-D-aspartate (NMDA) receptors. The hippocampal slices of mice were exposed to OGD for 10 (OGD10) or 45 minutes (OGD45) to mimic mild (causing ischemic/hypoxic preconditioning) and severe (causing severe OGD) ischemia/hypoxia, respectively. We found that OGD10-induced nPKCepslilon membrane translocation was mediated by NMDA receptors, and both OGD10 and NMDA (1 microM, 30 min) pretreatment could protect Cornu Ammonis region 1 neurons against the subsequent severe OGD45. In addition, nPKCepsilon translocation inhibitor, epsilonV1-2 (1 microM, 30 min), and ERKs upstream mitogen-activated protein/extracellular signal regulated kinase kinase inhibitor, PD-98059 (20 microM, 30 min), could significantly inhibit OGD10 and NMDA-induced neuroprotection. These results suggest that OGD10-induced neuroprotection against severe OGD45 in the Cornu Ammonis region 1 region of the hippocampal slices was mediated by the activations of NMDA receptors, nPKCepsilon, and the downstream ERKs.     

吗啡预处理对小鼠海马脑片氧-糖缺失/恢复时CaMKⅡ和NMDA受体的影响

目的 评价吗啡预处理对小鼠海马脑片氧-糖缺失，恢复时钙，钙调蛋白依赖性蛋白激酶Ⅱ（CaMKⅡ）膜转位及N-甲基-D-天冬氨酸（NMDA）受体磷酸化的影响。方法成年BALB／C小鼠，体重18-22g，雌雄不拘。每次将5只小鼠同批断头取脑，制备海马脑片，随机分为5组：正常对照组（Ⅰ组）、氧．糖缺失，恢复组（Ⅱ组）、吗啡预处理组（Ⅲ组）、纳络酮＋吗啡预处理组（Ⅳ组）及纳络酮预处理组（Ⅴ组）。Ⅰ组脑片正常体外培养，假操作换液。Ⅲ、Ⅳ、Ⅴ组小鼠海马脑片分别作相应预处理30min，间隔30min。Ⅱ、Ⅲ、Ⅳ、Ⅴ组建立小鼠海马脑片体外缺血再灌注损伤模型，分别氧-糖缺失20min，氧-糖恢复2h。各组于氧．糖缺失前即刻（T0）、氧-糖缺失20min（T1）、氧-糖恢复1h（T2）和2h（T3）取若干脑片匀浆、超声粉碎、离心，分离胞浆蛋白和膜相关蛋白；部分脑片分离总蛋白，Western blot检测CaMKⅡα蛋白表达量以及NMDA受体NR1亚单位的磷酸化。结果 海马脑片在T1、T2．T3时，CaMKⅡα膜相关蛋白含量增多。同时胞浆蛋白含量减少（P〈0．05）；T2、T3时，NMDA受体NR1亚单位磷酸化水平增高（P〈0．05）；而吗啡预处理明显地抑制上述CaMKⅡα膜转位及NMDA受体NR1亚单位磷酸化（P〈0．05）。纳络酮完全阻断吗啡预处理对CaMKⅡα膜转位及NR1磷酸化的抑制作用（P〈0．05）。CaMKⅡα总蛋白表达水平未见明显变化。结论 CaMKⅡ膜转位的抑制及NMDA受体磷酸化的抑制在吗啡预处理对小鼠海马脑片缺血再灌注的脑保护作用机制中起重要作用。     

The role of N-methyl-D-aspartate (NMDA) receptors in pain and morphine tolerance

To determine the importance of the N-methyl-D-aspartate (NMDA) receptor in pain hypersensitivity following injury, the NMDAR1 subunit was selectively deleted in the lumbar spinal cord of adult mice by the localized injection of an adeno-associated virus expressing the Cre recombinase into floxed NR1 mice. This procedure resulted in more than an 80% reduction in the expression of both NR1 mRNA and protein and a corresponding loss of NMDA, but not AMPA currents, in the lamina II neurons in the injected area. This spatially and temporally restricted knockout dramatically reduced the response to pain hypersensitivity resulting from the intraplantar injection of formalin but did not alter heat or cold paw withdrawal latencies, mechanical threshold, or motor function. Thus, the NMDA receptor in the spinal cord dorsal horn is essential for central sensitization, the central facilitation of pain transmission produced by peripheral injury. Agents that act on these spinal receptors may provide a therapeutic approach to ameliorate injury-induced pain.     

NMDA受体拮抗剂LY274614对吗啡耐受性和依赖性的抑制作用

目的 观察竞争性N－甲基－D－天门冬氨酸（NMDA）受体拮抗剂LY274614对大鼠吗啡耐受性和依赖性的抑制作用。方法 对大鼠皮下注射吗啡（15mg/kg,3次／d)产生吗啡耐受性和依赖性，并在注射吗啡的同时皮下注射LY274614（2，4，6mg/kg),测定1h后大鼠热辐射甩尾反应潜伏期（TF)以评价吗啡的镇痛效果。实验的第10d，观察皮下注射纳络酮诱发的戒断症状以评价大鼠对吗啡的躯体依赖行为。结果 LY27461本身不产生镇痛作用，但与吗啡合用时可抑制吗啡耐受性产生而引起的TF下降，并可减少纳络酮诱发的戒断症状发生次数。结论 竞争性NMDA受体拮抗剂LY274614可抑制大鼠经持续皮下注射吗啡而产生的吗啡耐受性和依赖性。     

Partial blocking of NMDA receptors reduces plastic changes induced by short-lasting classical conditioning in the SI barrel cortex of adult mice.

The effect of blockade of N-methyl-D-aspartate (NMDA) receptors in the barrel cortex upon the learning-induced changes of the cortical body map was examined in adult mice. We have previously found that three sensory conditioning sessions, in which stimulation of a row of vibrissae was paired with a tail shock, produced an enlargement of the functional representation of a row of vibrissae stimulated during training. Implantation of the slow release polymer Elvax, containing 2-amino-5-phosphonovalerate (APV, 50 mM), in the vicinity of the barrel cortex was performed 1 day before conditioning to block NMDA receptors. The cortical representation of a trained row of vibrissae was visualized with 2-deoxyglucose (2DG) functional brain mapping 1 day after the completion of the conditioning procedure. The partial blockade of NMDA receptors within the barrel cortex reduced (by half) the expansion of the cortical representation of a trained row of vibrissae as compared to the enlargement of the cortical representation of a trained row found in untreated (60%) and Elvax-PBS implanted (47%) mice. The results provide evidence that the learning-induced processes of cortical map reorganization involve mechanisms that depend on NMDA receptor activation.     

gamma-Aminobutyric acid-A receptors contribute to isoflurane neuroprotection in organotypic hippocampal cultures

The mechanisms by which anesthetics such as isoflurane reduce cell death in rodent models of cerebral ischemia remain incompletely defined. Reduction in glutamate excitotoxicity explains some but not all of isoflurane's neuroprotection. Because isoflurane potentiates gamma-aminobutyric acid (GABA) receptor-mediated ion fluxes and GABA(A) receptor agonists have neuroprotective effects, we hypothesized that GABA(A) receptors contribute to isoflurane neuroprotection. As a model of cerebral ischemia and recovery, we used rat hippocampal slice cultures. Survival of CA1, CA3, and dentate neurons was examined 2 and 3 days after 1-h combined oxygen-glucose deprivation (OGD) at 37 degrees C. To define the role of GABA(A) receptors in mediating protection, the effect of 1% isoflurane on cell survival was examined in the presence of the GABA(A) antagonist bicuculline during OGD. Cell death was measured with propidium iodide fluorescence. Isoflurane and the selective GABA(A) agonist muscimol (25 micro M) reduced cell death after OGD to values similar to slices not exposed to OGD, with the exception that muscimol did not reduce cell death in CA3 neurons 2 days after OGD. The GABA(A) antagonist bicuculline reduced the neuroprotective effects of isoflurane on hippocampal neurons 2 and 3 days after OGD. We conclude that GABA(A) receptors contribute to neuroprotection against OGD produced by isoflurane in the hippocampal slice model. Based on this and other studies, it is likely that neuroprotection produced by isoflurane is multifactorial and includes actions at both GABA(A) and glutamate receptors and possibly other mechanisms. IMPLICATIONS: Isoflurane is neuroprotective in rodent brain ischemia models, but the mechanisms for this effect remain incompletely defined. In organotypic cultures of rat hippocampus, we show that protection of CA1, CA3, and dentate neurons by 1% isoflurane from death caused by oxygen and glucose deprivation involves GABA(A) receptors.     

雷米芬太尼对依托咪酯引发肌震颤的影响

目的研究预注雷米芬太尼对依托咪酯引发肌震颤的影响。方法选择ASAⅠ或Ⅱ级,体重指数在20～24 kg/m2,无神经肌肉传导功能障碍性疾病的择期手术患者90例,随机均分为三组:雷米芬太尼0.5μg/kg组(R0.5组)、雷米芬太尼1μg/kg组(R1.0组)和对照组(C组)。预注雷米芬太尼或生理盐水,2 min后静脉注射依托咪酯0.3 mg/kg;记录预注前即刻(T0)、预注雷米芬太尼或生理盐水结束后1 min(T1)、2 min(T2)的SBP、DBP、HR、SpO2、RR、潮气量(VT),观察肌震颤程度并进行评分。结果与C组相比,R0.5和R1.0组依托咪酯引发肌震颤程度降低(P<0.01),R0.5和R1.0组相比肌震颤程度差异无统计学意义;体重60 kg以上的患者与其他体重段的患者相比肌震颤相对易发生且程度严重(P<0.05);各组同一时点的SBP、DBP、HR、SpO2、RR、VT差异无统计学意义。结论预注雷米芬太尼0.5或1μg/kg均能够显著降低依托咪酯引发的肌震颤,对呼吸系统和循环系统没有明显影响。     

大鼠肝缺血及再灌注所致小肠过氧化损伤及丹参的保护作用

目的 观察大鼠肝缺血再灌注小肠过氧化损伤及丹参预处理的保护作用.方法 首先将SD大鼠随机分为正常对照组(CO组)、假手术组(SO组)、缺血再灌注组(IR组)、丹参预处理组(SM组),分别在肝缺血30、45、60 min时取上段空肠进行大体病理学检测;然后在肝缺血45 min条件下,动物亦随机分为4组(CO组、SO组、IR组、SM组),按再灌注后不同时间(0、3、12、24、72 h)分为5个亚组,每组5只.SM组在阻断第一肝门30 min前经尾静脉推注丹参注射液6 g/kg加生理盐水40 ml/kg,其余各组按40 ml/kg给予生理盐水尾静脉注入,SO组开腹后仅解剖肝门,不钳夹肝蒂.分别在再灌注0、3、12、24、72 h取上段空肠行病理学检查、丙二醛(MDA)含量测定、髓过氧化物酶(MPO)活性测定.结果 空肠黏膜损伤评分随肝缺血时限延长而加重;在肝缺血45 min再灌注不同时限点SM组空肠黏膜损伤较IR组明显减轻,且肠组织MDA含量、MPO活性均低于IR组(P<0.05).结论 肝缺血再灌注所致小肠明显淤血性损伤,MDA含量、MPO活性升高,丹参预处理对肝缺血再灌注所致小肠损伤具有保护作用.     

Lidocaine pretreatment reduces the frequency and severity of myoclonus induced by etomidate

The objective of this study was to assess the effects of lidocaine on the incidence and severity of myoclonic movements induced by etomidate. Sixty patients were randomly assigned to receive either 20 mg lidocaine or saline (n = 30, each), 30 s before administration of etomidate (0.3 mg/kg). One minute after etomidate administration we assessed severity of myoclonus. Pretreatment with lidocaine significantly reduced both the incidence and severity of myoclonic movements. As a conclusion, lidocaine is an effective and safe drug to reduce the etomidate-induced myoclonus without significant side effects.     

促红细胞生成素预先给药对大鼠内毒素性急性肺损伤的影响

目的 探讨促红细胞生成素(EPO)预先给药对大鼠内毒素性急性肺损伤的影响.方法 成年雄性SD大鼠32只,体重180～220 g,随机分为4组(n=8),C组腹腔注射生理盐水4 ml/kg(EPO溶剂对照),30 min后静脉注射生理盐水2 ml/kg[脂多糖(LP3)溶剂对照];EPO组腹腔注射EPO3 000 U/kg,30 min后静脉注射生理盐水2 ml/kg;LPS组腹腔注射生理盐水4 ml/kg,30 min后静脉注射LPS 6 mg/kg;EPO+LPS组腹腔注射EPO 3 000 U/kg,30 min后静脉注射LPS 6 mg/kg.于静脉注射LPS后4 h时处死大鼠,观察肺组织病理学结果 ,计算肺组织湿/干重(W/D)比;测定肺组织髓过氧化物酶(MPO)活性和丙二醛(MDA)、一氧化氮(NO)含量;采用Western blot法测定肺组织诱导型一氧化氮合酶(iNOS)和硝基酪氨酸(NT)的表达.结果 与C组相比,LPS组和EPO+LPs组肺组织W/D比、MPO活性、MDA和NO含量升高,iNOS和NT表达上调(P<0.01);与LPS组相比,EPO+LPS组肺组织W/D比、MPO活性、MDA和NO含量降低,iNOS和NT表达下调(P<0.01).结论 EPO预先给药可减轻大鼠内毒素性急性肺损伤,与其下调iNOS表达,减少NO生成有关.     

Subhypnotic doses of propofol relieve pruritus induced by epidural and intrathecal morphine.

We investigated the efficacy of subhypnotic doses of propofol for spinal morphine-induced pruritus in a prospective, randomized, double-blind, placebo-controlled study. Fifty patients, ASA physical status 1-3, with spinal morphine-induced pruritus were allocated to receive either 1 ml propofol (10 mg) or 1 ml placebo (Intralipid) intravenously after gynecologic, orthopedic, thoracic, or gastrointestinal surgery. In the absence of a positive response, a second drug treatment was given 5 min later. The persistence of pruritus 5 min after the second treatment dose was considered a treatment failure. All failures then received, in an open fashion, a supplementary dose of propofol (10 mg) and were reevaluated 5 min later. Both groups were well matched. The success rate was significantly greater in the propofol group (84%) than in the placebo (16%) group (P less than 0.05). Ninety percent of the treatment failures in the placebo group were successfully treated by a supplementary dose of 10 mg propofol. Eight percent of the patients (4% in each group) were resistant to all treatments, including naloxone 0.08 mg intravenously. Three patients had a slight increase in sedation in the propofol group versus none in control (not significant). The beneficial effect of treatment was longer than 60 min in 85% of patients in the propofol group and in 100% of the controls (not significant). These results suggest that propofol in a subhypnotic dose is an efficient drug treatment for spinal morphine-induced pruritus. At the dose administered (10 mg), side effects were rare and minor.