基于PI3K/Akt通路研究右美托咪定对癫痫持续状态大鼠海马神经元自噬的调节作用

目的 探讨右美托咪定(Dexmedetomidine,Dex)对癫痫持续状态(Status epilepticus,SE)大鼠海马神经元自噬的调节作用及对磷脂酰肌醇-3-羟激酶/蛋白激酶B(PI3K/Akt)信号通路的影响。方法 采用戊四氮(PTZ)点燃制备SE大鼠模型,随机分为模型组(M组)、Dex低剂量(Dex-L)组、Dex高剂量(Dex-H)组、Dex-H +LY294002(Dex-H+LY)组,另取正常SD大鼠为对照组(NC组),每组各15只; Dex-L组、Dex-H组分别腹腔注射Dex 25、100 μg/kg; Dex-H+LY组脑室注射5 μL PI3K抑制剂LY294002+腹腔注射Dex 100 μg/kg,NC组、M组大鼠腹腔注射等量生理盐水; 观察大鼠行为学表现并进行脑电图描记; 原位末端标记法(TUNEL)检测各组大鼠海马神经元凋亡情况; 免疫印迹(WB)检测各组大鼠海马组织中LC3、PI3K、p-PI3K、Akt、p-Akt蛋白表达水平。结果 NC组大鼠脑电图无异常放电现象,主要以α、β波为主; M组大鼠出现大量阵发性棘波、高幅尖波、棘慢复合波、尖慢复合波等; Dex-L组、Dex-M组大鼠癫痫发作减少或波幅降低; Dex-H+LY组大鼠癫痫样放电较Dex-H组明显增加。与NC组比较,M组大鼠海马神经元凋亡细胞数、海马LC3-Ⅱ蛋白表达显著增加,海马LC3-Ⅰ,p-PI3K/PI3K,p-Akt/Akt蛋白表达水平显著降低(P<0.05); 与M组比较,Dex-L组、Dex-H组大鼠海马神经元凋亡细胞数、海马LC3-Ⅱ蛋白表达依次减少(P<0.05),海马LC3-Ⅰ,p-PI3K/PI3K,p-Akt/Akt蛋白表达水平依次增高(P<0.05); 与Dex-H组比较,Dex-H+LY组大鼠海马神经元凋亡细胞数、海马LC3-Ⅱ蛋白表达显著增加,海马LC3-Ⅰ,p-PI3K/PI3K,p-Akt/Akt蛋白表达水平显著降低(P<0.05)。结论 Dex可能通过促进PI3K/Akt信号通路激活来抑制SE大鼠海马神经元过度自噬,减轻神经元凋亡,发挥抗惊厥及脑保护作用。     

α-细辛醚对氯化锂-匹罗卡品致痫大鼠PI3K/ Akt /mTOR通路的调节作用

目的 探讨α- 细辛醚对癫痫大鼠学习记忆的影响及分子机制。方法 SD 大鼠随机分成 生理盐水组、癫痫组、α-细辛醚组,采用氯化锂-匹罗卡品注射诱导大鼠癫痫造模,α-细辛醚组造模 前用α-细辛醚干预28 d。水迷宫检测大鼠空间学习记忆能力,Western blot方法检测大鼠海马组织PI3K、 Akt、mTOR蛋白表达,采用RT-PCR技术检测大鼠海马组织PI3K、Akt、mTOR 的 mRNA表达。结果 水 迷宫实验结果显示α-细辛醚能够改善大鼠的学习记忆能力;Western blot 结果显示,癫痫组海马组织 PI3K、p-Akt、mTOR 蛋白表达较生理盐水组降低（P＜ 0. 05） ,α-细辛醚组海马组织的各蛋白表达较癫 痫组增加（P＜ 0. 05）。RT-PCR 结果显示,癫痫组海马组织PI3K、p-Akt、mTOR mRNA表达水平与生理盐 水组比较显著降低（P＜ 0. 05）,α-细辛醚组海马组织PI3K、p-Akt 的mRNA表达水平与癫痫组比较显著 增高（P＜ 0. 05） 。结论 α-细辛醚能够改善氯化锂-匹罗卡品致痫大鼠学习记忆,其分子机制可能与 上调PI3K/Akt /mTOR 表达有关。     

PI3K/Akt信号通路在癫痫鼠海马神经细胞中的作用及葡萄籽原花青素影响

目的探讨PI3K/Akt信号通路在戊四氮(PTZ)诱导癫痫大鼠海马神经细胞凋亡中的作用及葡萄籽原花靑素(GSPE)对其影响。方法根据是否给予PI3K/Akt信号通路特异性抑制剂LY294002,将大鼠随机分为5组,每组10只:正常对照组、癫痫模型组(PTZ组)、LY294002+PTZ组、GSPE+LY294002+PTZ组、GSPE+PTZ组。采用流式细胞学检测海马组织线粒体膜电位(ΔΨm)水平,Western blot法检测海马组织p-Akt、Caspase 3蛋白的表达变化,电镜观察线粒体的超微结构,以明确GSPE预处理对上述指标的影响。结果与PTZ组相比,LY294002+PTZ组中Caspase 3表达水平显著升高。与LY294002+PTZ组相比,GSPE+PTZ组中p-Akt水平明显升高、Caspase 3蛋白水平明显下降(P0.05),相应海马组织中线粒体ΔΨm的水平升高(P0.05)及超微结构有所改善;而GSPE+LY294002+PTZ组中上述指标则差异无统计学意义。结论 GSPE干预可抑制癫痫鼠海马神经细胞的凋亡并保护线粒体的功能,其机制可能与激活PI3K/Akt信号通路抑制Caspase 3的表达有关。     

脑缺血再灌注后STIM1通过内质网应激促进小胶质/巨噬细胞M1型活化的作用研究

目的探讨基质相互作用分子1(STIM1)对脑缺血再灌注损伤后小胶质/巨噬细胞M1型活化的影响及机制。方法 (1)动物实验:采用随机数字表法将20只雄性C57BL/6J小鼠分为假手术组、大脑中动脉缺血再灌注(MCAO/R)组、MCAO/R+si-Ctrl组及MCAO/R+si-STIM1组, 后3组小鼠建立MCAO/R模型, MCAO/R+si-Ctrl组及MCAO/R+si-STIM1组小鼠分别转染空载体对照病毒和STIM1基因敲除慢病毒。观察STIM1转染效率及各组小鼠中小胶质/巨噬细胞M1型活化标记物分化抗原86(CD86)的表达情况。(2)细胞实验:将原代小胶质细胞分为Ctrl组、氧糖剥夺/复氧(OGD/R)组、OGD/R+si-Ctrl组、OGD/R+si-STIM1组、OGD/R+溶媒组及OGD/R+4-苯基丁酸(4-PBA)组。后5组细胞构建OGD/R模型, OGD/R+si-Ctrl组、OGD/R+si-STIM1组细胞分别转染空载体对照病毒和STIM1基因敲除慢病毒;OGD/R+4-PBA组细胞在OGD/R造模前24 h使用1 mmol/L预处理1 h以抑制内质网应激(...     

氯化锂通过Akt/GSK-3β通路抑制苯丙胺诱导的大鼠行为敏化

目的 探讨氯化锂对苯丙胺诱导的大鼠行为敏化的影响及蛋白激酶B（Akt）/ 糖原合成激 酶-3β（GSK-3β）通路在其中的作用。方法 40 只雄性SD 大鼠随机分为空白对照组（S-S 组,n=13）、苯 丙胺敏化组（S-A 组,n=13）和氯化锂预处理苯丙胺敏化组（L-A组,n=14）,S-S 组大鼠接受连续5 d的生 理盐水（10 ml/kg）预处理+ 生理盐水（10 ml/kg）腹腔注射; S-A 组大鼠接受连续5 d 的生理盐水（10 ml/kg） 预处理+苯丙胺（1.5 mg/kg）腹腔注射;L-A组大鼠接受5 d的氯化锂（100 mg/kg）预处理+苯丙胺（1.5 mg/kg） 腹腔注射,之后停药6 d,第12 天时每组随机选取6 只大鼠接受低剂量苯丙胺腹腔注射。采用自发活 动视频分析记录系统记录3 组大鼠每组随机6 只分别在用药第1 天和第12 天时150 min 内的自发活 动。第12 天时在未接受行为学测试的3 组大鼠中每组随机6 只,采用Western blot 检测其前额叶皮质磷 酸化Akt（p-Akt）/Akt、磷酸化GSK-3β（p-GSK-3β）/GSK-3β表达水平。结果 在用药第1 天时,S-A组 大鼠的自发活动总距离［（26 826.50±5 987.96） cm］明显高于S-S 组［（1 861.50±612.49） cm］和L-A 组 ［（7 599.00±4 778.14） cm］,差异均有统计学意义（均P＜0.05）。在第12天时,苯丙胺激发实验中S-A组大 鼠自发活动总运动距离［（43 823.83±5 831.88） cm］明显高于S-S 组［（14 274.50±4 724.98） cm］和L-A组 ［（17 823.50±4 313.64） cm］,差异均有统计学意义（均P ＜ 0.05）。S-A 组大鼠前额叶皮质p-Akt/Akt 比 值和p-GSK-3β/GSK-3β比值均低于S-S 组（均P ＜ 0.05）,而L-A 组大鼠前额叶皮质p-Akt/Akt 比值和 p-GSK-3β/GSK-3β比值高于S-A组（均P＜0.05）。结论 氯化锂可以抑制苯丙胺诱导的大鼠行为敏化, 其机制可能是通过Akt/GSK-3β 通路介导。     

SB-399885阻断5-羟色胺6受体/雷帕霉素靶蛋白 途径对精神分裂症大鼠认知和记忆障碍的改善作用

目的 研究 5- 羟色胺 6 受体（HTR6）拮抗剂 SB-399885 通过阻断 HTR6 / 雷帕霉素靶蛋 白（mTOR）途径对精神分裂症大鼠认知和记忆障碍的作用。方法 将 40 只 SD 大鼠随机分为 4 组：空 白对照组、精神分裂症模型组（SZ 组）、SZ+SB-399885 组（10 mg/kg）、阳性对照组（SZ+ 利培酮,0.1 mg/kg）, 每组各 10 只大鼠。采用 MK-801 建立 SZ 大鼠模型。新物体辨别实验检测大鼠视觉识别记忆,Morris 水 迷宫实验检测大鼠认知能力,被动回避实验检测大鼠学习记忆能力,乙酰胆碱酯酶（AChE）活性测定 试剂盒检测大鼠海马和大脑皮质 AChE 活性,TUNEL 染色法检测大鼠海马 CA1 区神经细胞凋亡情况, Western blot 检测大鼠海马 HTR6 / mTOR 途径相关蛋白的表达。 结果 5-HT6 受体拮抗剂 SB-399885 和 利培酮均能显著改善 SZ 大鼠视觉识别记忆障碍、认知障碍和学习记忆障碍（均P＜0.05）。与SZ组比较, SZ+SB-399885组大鼠海马AChE活性［（0.008±0.001）μmol/（min·mg）］、神经细胞凋亡率［（21.75±4.45）%］、 HTR6 蛋白表达（0.56±0.10）和 mTOR 活性（0.41±0.05）均显著降低（均P＜ 0.05）;阳性对照组大鼠海马 和大脑皮质 AChE 活性显著降低（均P＜ 0.05）,大鼠海马神经细胞凋亡率［（19.28±5.22）%］、HTR6 蛋 白表达（0.40±0.10）和 mTOR 活性（0.33±0.05）均显著降低（均P＜ 0.05）。结论 5-HT6 受体拮抗剂 SB- 399885 可能通过阻断 HTR6 / mTOR 途径改善精神分裂症大鼠认知和记忆障碍。     

VEGF介导PI3/K信号通路保护脊髓运动神经元的实验研究

目的建立脊髓前角运动神经元兴奋性氨基酸损伤的模型,以研究不同浓度血管内皮生长因子(Vascular endothelial growth factor,VEGF)干预及磷脂酰肌醇3-激酶(Phosphatidyl-inositol3-kinase,PI3/K)抑制剂对其活力及PI3/K表达的影响。方法体外培养新生SD大鼠脊髓运动神经元,并建立对照组、VEGF组、谷氨酸(Glutamate,Glu)组及PI3/K组,通过MTT法检测各组运动神经元活力,观察各组大鼠脊髓运动神经元的数量,采用免疫荧光方法及酶标记免疫吸附测定法检测PI3/K的表达水平。结果筛选出适宜建立兴奋性氨基酸损伤模型的Glu浓度为5 umol/ml。在Glu干预下VEGF0.1 ug/ml组的细胞活力较VEGF1 ug/ml组显著降低(P<0.05);VEGF1 ug/ml加PI3/K组的细胞活力较VEGF1 ug/ml组显著降低(P<0.05);通过免疫荧光法及ELISA法检测运动神经元的PI3/K表达时也有相似的发现。结论 VEGF可改善由Glu所造成的脊髓运动神经元损伤,增加损伤神经元PI3/K的表达,且在一定浓度范围内存在量效关系。通过LY294002抑制VEGF增加细胞PI3/K表达的作用,推测VEGF对运动神经元的保护作用可能部分通过PI3/K信号通路完成。     

依达拉奉调控PI3K/Akt信号通路抑制七氟烷致海马神经元凋亡

目的 探讨依达拉奉对七氟烷致海马神经元凋亡及对PI3K/Akt信号通路影响.方法 将80只成年昆明小鼠随机分为正常组、七氟烷组、七氟烷+依达拉奉组及LY294002+七氟烷+依达拉奉组,每组20只.正常组:小鼠不做处理;七氟烷组:小鼠每天给予1h吸入1.5％七氟烷;七氟烷+依达拉奉组:小鼠每天尾静脉注射依达拉奉3 mg...     

环磷酸腺苷反应元件结合蛋白与认知功能障碍相关疾病研究进展

目的 基于磷酯酰激醇 3- 激酶 / 蛋白激酶 B（PI3K/Akt）通路探讨氯吡格雷对脑缺血再 灌注损伤大鼠的神经保护作用。方法 建立脑缺血再灌注大鼠模型，随机分为模型组、氯吡格雷组、 LY294002（PI3K 抑制剂）组、氯吡格雷 +LY294002 组，每组 12 只，另取 12 只 SD 大鼠设为假手术组。分组 处理后，所有大鼠进行神经功能缺损评分并尾静脉取血，处死大鼠，HE 染色检测各组大鼠神经元病理 情况；三苯基氯化四氮唑（TTC）染色检测各组大鼠脑组织梗死面积；ELISA 检测血清中中枢神经特异性 蛋白（S100β）、神经元特异性烯醇化酶（NSE）、白细胞介素 -6（IL-6）、肿瘤坏死因子 -α（TNF-α）水平；蛋 白免疫印迹法检测脑组织中 PI3K/Akt 通路蛋白表达情况。结果 与假手术组相比，模型组大鼠脑组织 神经元出现坏死、核收缩变小等病理变化，神经功能缺损评分、脑梗死面积、血清中 S100β、NSE、IL-6 及 TNF-α 水平均明显升高（P＜ 0.05），脑组织中 p-PI3K/PI3K、p-Akt/Akt 明显降低（P＜ 0.05）；与模型组相 比，氯吡格雷组大鼠神经元病理损伤减轻，神经功能缺损评分、脑梗死面积、血清中 S100β、NSE、IL-6 及 TNF-α 水平均降低（P＜ 0.05），脑组织中 p-PI3K/PI3K、p-Akt/Akt 升高（P＜ 0.05）；LY294002 组大鼠神 经元病理损伤加重，神经功能缺损评分、脑梗死面积、血清中 S100β、NSE、IL-6 及 TNF-α水平均升高 （P＜ 0.05），脑组织中 p-PI3K/PI3K、p-Akt/Akt 降低（P＜ 0.05）。与 LY294002 组相比，氯吡格雷 +LY294002 组大鼠神经元病理损伤减轻，神经功能缺损评分、脑梗死面积、血清中 S100β、NSE、IL-6 及 TNF-α 水 平均降低（P＜ 0.05），脑组织中 p-PI3K/PI3K、p-Akt/Akt 升高（P＜ 0.05）。与氯吡格雷组相比，氯吡格雷 + LY294002 组大鼠神经元病理损伤加重，神经功能缺损评分、脑梗死面积、血清中 S100β、NSE、IL-6 及 TNF-α 水平均升高（P＜ 0.05），脑组织中 p-PI3K/PI3K、p-Akt/Akt 降低（P＜ 0.05）。结论 氯吡格雷可通 过激活 PI3K/Akt 通路减轻大鼠脑缺血再灌注损伤，保护脑组织。     

基于PI3K/Akt通路探讨氯吡格雷对脑缺血再灌注损伤大鼠的神经保护作用

目的基于磷酯酰激醇3-激酶/蛋白激酶B(PI3K/Akt)通路探讨氯吡格雷对脑缺血再灌注损伤大鼠的神经保护作用.方法建立脑缺血再灌注大鼠模型,随机分为模型组、氯吡格雷组、LY294002(PI3K抑制剂)组、氯吡格雷+LY294002组,每组12只,另取12只SD大鼠设为假手术组.分组处理后,所有大鼠进行神经功能缺损评分并尾静脉取血,处死大鼠,HE染色检测各组大鼠神经元病理情况;三苯基氯化四氮唑(TTC)染色检测各组大鼠脑组织梗死面积;ELISA检测血清中中枢神经特异性蛋白(S100β)、神经元特异性烯醇化酶(NSE)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)水平;蛋白免疫印迹法检测脑组织中PI3K/Akt通路蛋白表达情况.结果与假手术组相比,模型组大鼠脑组织神经元出现坏死、核收缩变小等病理变化,神经功能缺损评分、脑梗死面积、血清中S100β、NSE、IL-6及TNF-α水平均明显升高(P<0.05),脑组织中p-PI3K/PI3K、p-Akt/Akt明显降低(P<0.05);与模型组相比,氯吡格雷组大鼠神经元病理损伤减轻,神经功能缺损评分、脑梗死面积、血清中S100β、NSE、IL-6及TNF-α水平均降低(P<0.05),脑组织中p-PI3K/PI3K、p-Akt/Akt升高(P<0.05);LY294002组大鼠神经元病理损伤加重,神经功能缺损评分、脑梗死面积、血清中S100β、NSE、IL-6及TNF-α水平均升高(P<0.05),脑组织中p-PI3K/PI3K、p-Akt/Akt降低(P<0.05).与LY294002组相比,氯吡格雷+LY294002组大鼠神经元病理损伤减轻,神经功能缺损评分、脑梗死面积、血清中S100β、NSE、IL-6及TNF-α水平均降低(P<0.05),脑组织中p-PI3K/PI3K、p-Akt/Akt升高(P<0.05).与氯吡格雷组相比,氯吡格雷+LY294002组大鼠神经元病理损伤加重,神经功能缺损评分、脑梗死面积、血清中S100β、NSE、IL-6及TNF-α水平均升高(P<0.05),脑组织中p-PI3K/PI3K、p-Akt/Akt降低(P<0.05).结论氯吡格雷可通过激活PI3K/Akt通路减轻大鼠脑缺血再灌注损伤,保护脑组织.     

银杏内酯 K 通过 PI3K/Akt/mTOR 通路促进 血管生成改善缺血性卒中

目的 描述综合医院具有自杀倾向患者的特征,并分析此类患者发生自杀行为的危险因 素。方法 回顾性连续纳入南方医科大学南方医院 2012 年 10 月至 2017 年 10 月的住院患者中具有自杀 倾向的病例共 680 例,根据是否发生自杀行为分为自杀倾向组（有自杀倾向无自杀行为,527 例）和自杀 行为组（有自杀倾向亦有自杀行为,153 例）。收集他们的性别、年龄、婚姻状况、户籍、入院科室、自杀诱 因、自杀方式、精神疾患、躯体疾患及显著的精神症状等资料。采用单因素分析比较两组间的社会人口 学和临床特征差异,采用二分类 Logistic 回归分析研究自杀行为的危险因素。结果 自杀倾向组主要为 女性［66.6%（351例）］、已婚［77.4%（458例）］、心理科［74.4%（392例）］、抑郁症［66.2%（349例）］。自杀行为 组中,男性和女性的自杀方式差异有统计学意义（χ2 =12.489,P=0.014）,选择跳楼方式的男性较多,药物 及割脉方式的女性更多。Logistic回归分析结果表明,入住重症医学科（OR=7.844,95%CI：2.240～27.475, P=0.001）、婚恋受挫（OR=3.646,95%CI：1.217～10.917,P=0.021）,肿瘤（OR=4.620,95%CI：1.552～13.755, P=0.006）,双相情感障碍（OR=3.734,95%CI：1.157～12.052,P=0.028）是自杀行为的危险因素。结论 具 有自杀倾向的患者中,入住重症医学科、双相情感障碍、肿瘤、婚恋挫折是发生自杀行为的危险因素,而 且两性的自杀方式有所不同,需要临床高度重视,并采取针对性预防措施。     

H2S protects hippocampal neurons from anoxia-reoxygenation through cAMP-mediated PI3K/Akt/p70S6K cell-survival signaling pathways

The study aims to investigate the effect of hydrogen sulfide (H₂S) on the phosphatidylinositol 3-kinase (PI3K)/Akt/p70 ribosomal S6 kinase (p70S6K) signal transduction pathway after oxygen glucose deprivation/reoxygenation (OGD/R) in the rat hippocampus. Newborn Wister rats were decapitated under anesthesia, and hippocampal tissue was dissected. Cells were plated at 1.0 × 10⁵ cells/mL on polylysine-treated 96-well and 6-well plates. After 7 days in culture, cells were randomly assigned to six groups: control, OGD/R, sodium hydrosulfide (NaHS) following OGD/R, NaHS/triciribine following OGD/R, NaHS/rapamycin following OGD/R, and NaHS/triciribine/rapamycin following OGD/R. Neuronal purity and cell viability were assessed in each group, as well as apoptosis and expression of cyclic adenosine 3′, 5′-monophosphate (cAMP), PI3K, Akt, and p70S6K. NaHS enhanced cAMP concentration and expression of PI3K, Akt, and p70S6K. In addition, neuronal viability was increased and apoptotic neuronal numbers decreased (P < 0.01). Triciribine inhibited Akt and p70S6K, as well as decreased cell survival and viability compared with the NaHS group (P < 0.05 or P < 0.01). Rapamycin resulted in decreased p70S6K expression and neuronal viability, as well as increased number of apoptotic neurons compared with the NaHS group (P < 0.05 or P < 0.01). H₂S acted via cAMP-mediated PI3K/Akt/p70S6K signal transduction pathways to inhibit hippocampal neuronal apoptosis and protect neurons from OGD/R-induced injury.     

Screening performance of K6/K10 and other screening instruments for mood and anxiety disorders in Japan

Aims: This study aimed to establish the screening performance and optimal cut‐off points for the Japanese version of Kessler (K)6, K10 and the Depression and Suicide Screen (DSS). Methods: A self‐report questionnaire including K6, K10 and DSS, as well as the Center for Epidemiologic Studies – Depression Scale (CES‐D), was administered to a random sample of community residents in Japan (non‐cases, n = 147) and psychiatric outpatients diagnosed with mood or anxiety disorders according to DSM‐IV (cases, n = 17). A receiver–operator characteristics (ROC) curve was drawn to estimate the area under the curve (AUC), the sensitivity, and specificity with the optimal cut‐off points for K6, K10, and DSS, which were then compared with those of CES‐D. The community sample was also asked to rate each measure on a scale from ‘very easy’ to ‘very hard’ to use. Results: K6 and K10 showed a high AUC (0.93–0.94), which was comparable to that of CES‐D (0.95), but DSS showed a significantly smaller AUC (0.89) than CES‐D (P < 0.05). The optimal cut‐off points were estimated as 4/5 for K6, 9/10 for K10, and 1/2 for DSS. The sensitivity of these three scales was similar, but the specificity was lower for DSS than for the other two. K6, K10 and DSS were rated as being ‘very easy’ or ‘easy to use’ significantly more than CES‐D (P < 0.01). Conclusion: The screening performance of the Japanese versions of K6 and K10 was comparable with that of CES‐D, and better than that of DDS. K6/K10, particularly K6, might have an advantage, even over the CES‐D, because of its similar screening performance and better acceptability.     

Characterization of electrogenic Na/K pump in rat neostriatal neurons

The electrogenic Na/K pump current (Ip) was studied in the dissociated neostriatal neurons of the rat by using the nystatin-perforated patch recording mode. The Ip was activated by external K⁺ in a concentration-dependent manner with an EC₅₀ of 0.7 mM at a holding potential (V_H) of −40 mV. Other monovalent cations also caused Ip and the order of potency was Tl⁺>K⁺, Rb⁺>NH₄⁺, Cs⁺>>>Li⁺. The Ip decreased with membrane hyperpolarization in an external solution containing 150 mM Na⁺, while the Ip did not show such voltage dependency without external Na⁺. Ouabain showed a steady-state inhibition of Ip in a concentration- and temperature-dependent manner at a V_H of −40 mV. The IC₅₀ values at 20 and 30°C were 7.1×10⁻⁶ and 1.3×10⁻⁶ M, respectively. The decay of Ip after adding ouabain well fitted with a single exponential function. At a V_H of −40 Mv, the association (k₊₁) and dissociation (k₋₁) rate constants estimated from the time constant of the current decay at 20°C were 4.0×10² s⁻¹ M⁻¹ and 6.3×10⁻³ s⁻¹, respectively. At 30°C, k₊₁ increased to 2.8×10³ s⁻¹ M⁻¹ while k₋₁ showed no such change with a value of 1.8×10⁻³ s⁻¹. A continuous Na⁺ influx was demonstrated by both the Na⁺-dependent leakage current and tetrodotoxin-sensitive Na⁺ current, which resulted in the continuous activation of the Na/K pump. It was thus concluded that the Na/K pump activity was well-maintained in the dissociated rat neostriatal neurons with distinct functional properties and that the activity of the pump was tightly connected with Na⁺ influxes.     

Desensitization kinetics of a K acetylcholine response in Aplysia

We have studied the process of acetylcholine receptor desensitization on Aplysia medial pleural neurons under voltage clamp conditions. Acetylcholine, applied by microperfusion, elicits a biphasic response on these neurons, a rapid component which reverses polarity at about −60 mV and is Cl-dependent, and a slower component which reverses at about −85 mV and is K-dependent. Both components show desensitization, and the present study focuses on the K-dependent component, which could be isolated by maintaining membrane potential at the Cl equilibrium potential or by blocking the Cl component pharmacologically. K-dependent acetylcholine responses on these neurons varied in regard to time to peak of response and rate of desensitization. While the rising phase of the response was always fitted by a single exponential process, times to peak were divided somewhat arbitrarily into three broad groups of fast (<3 s), medium (3–6 s) and slow (>6 s.) Desensitization of fast responses was best described by two exponential processes plus a constant, medium responses by a double exponential, and slow responses by single exponential plus a constant. The apparent dissociation constant of acetylcholine was 17.3 ± 1.6 μM. The best fit of responses for a given cell remained constant over a range of acetylcholine doses, but the kinetics of both fast and slow components accelerated with dose and depolarization. The fast component of desensitization was very temperature dependent. In neurons where it was present it was abolished by cooling, while in neurons with no fast component at room temperature it would appear with warming. The time constant of the fast component varied inversely with temperature. The time constant of the slow component was maximal at 22–24 °C, and fell on either side of this temperature. These results suggest that receptor desensitization for acetylcholine K responses is, like Na-dependent responses, composed of two independent processes. When responses to the acetylcholine agonists, carbachol and arecoline, were compared to those of acetylcholine on fast-type neurons, the times to peak varied in the order acetylcholine < carbachol < arecoline. The carbachol response was best fitted by two exponential functions, while arecoline was best fitted by a single exponential plus a constant.     

Neuroglobin Protects Rats from Sepsis-Associated Encephalopathy via a PI3K/Akt/Bax-Dependent Mechanism

The pathogenesis of autism spectrum disorders (ASD) is not completely understood, but there is evidence of associations with altered immune responses. The aim of this study was to determine the serum levels of various cytokines in children with ASD and in healthy controls, in order to determine their role in ASD and its diagnostic subgroups. Sixty-five ASD patients were enrolled from an epidemiological survey in Norway, of which 30 were diagnosed with childhood autism, 16 with Asperger syndrome, 12 with atypical autism, 1 with Rett syndrome, and 6 with another ASD diagnosis. The serum levels of 12 cytokines were measured in all of the patients and in 30 healthy children. The cytokine levels did not differ significantly between the ASD group and the healthy controls. However, the interleukin-8 (IL-8) level was significantly higher (6.82 vs 4.58 pg/ml, p?=?0.017) while that of IL-10 was significantly lower (2.24 vs 6.49 pg/ml, p?=?0.009) in patients with childhood autism than in controls. Furthermore, the IL-8 level was significantly higher in childhood autism than in Asperger syndrome (6.82 vs 4.05 pg/ml, p?=?0.013). Our study shows that the cytokine profile of children diagnosed with ASD, regardless of the subdiagnosis, does not differ from healthy controls. However, differentiation into different diagnostic subgroups reveals significantly different levels of IL-8 and IL-10. This indicates that different mechanisms may underlie the different ASD subdiagnoses. Future research into the pathophysiological mechanisms of ASD should pay more attention to the different subdiagnoses of ASD.     

Neuregulin signaling through a PI3K/Akt/Bad pathway in Schwann cell survival

beta-Neuregulin (betaNRG) is a potent Schwann cell survival factor that binds to and activates a heterodimeric ErbB2/ErbB3 receptor complex. We found that NRG receptor signaling rapidly activated phosphoinositide 3-kinase (PI3K) in serum-starved Schwann cells, while PI3K inhibitors markedly exacerbated apoptosis and completely blocked NRG-mediated rescue. NRG also rapidly signaled the phosphorylation of mitogen-activated protein kinase (MAPK) and the serine/threonine kinase Akt. The activation of Akt and MAPK in parallel pathways downstream from PI3K resulted in the phosphorylation of Bad at different serine residues. PI3K inhibitors that blocked NRG-mediated rescue also blocked the phosphorylation of Akt, MAPK, and Bad. However, selective inhibition of MEK-dependent Bad phosphorylation downstream from PI3K had no effect on NRG-mediated survival. Conversely, ectopic expression of wild-type Akt not only enhanced Bad phosphorylation but also enhanced autocrine- and NRG-mediated Schwann cell survival. Taken together, these results demonstrate that NRG receptor signaling through a PI3K/Akt/Bad pathway functions in Schwann cell survival.     

Astrocytes protect oligodendrocyte precursor cells via MEK/ERK and PI3K/Akt signaling

Accumulating evidence suggest that trophic coupling among different cell types in the brain is required to maintain normal CNS function. Here we show that astrocytes secrete soluble factors that can be oligodendrocyte‐supportive. Oligodendrocyte precursor cells (OPCs) and astrocytes were prepared from neonatal rat brain and cultured separately. We conducted cell culture medium‐transfer experiments to examine whether astrocytes secrete OPC‐protective factors. Conditioned media from astrocytes protected OPCs against H₂O₂‐induced oxidative stress, starvation, and oxygen‐glucose deprivation. This protective effect may be mediated in part via ERK and Akt signaling pathways. Astrocyte‐conditioned media upregulated the phosphorylation levels of ERK and Akt in OPC cultures. Blockade of ERK or Akt signaling with U0126 or LY294002 cancelled the OPC‐protective effects of astrocyte‐conditioned media. Taken together, these data suggest that astrocytes are an important source for oligodendrocyte‐supportive factors. Coupling between these two major glial components in brain may be vital for sustaining white matter homeostasis. © 2009 Wiley‐Liss, Inc.     

Peripheral antinociceptive effect of pertussis toxin: activation of the arginine/NO/cGMP/PKG/ ATP-sensitive K channel pathway

The aim of the present study was to determine the effect of pertussis toxin (PTX) on inflammatory hypernociception measured by the rat paw pressure test and to elucidate the mechanism involved in this effect. In this test, prostaglandin E(2) (PGE(2)) administered subcutaneously induces hypernociception via a mechanism associated with neuronal cAMP increase. Local intraplantar pre-treatment (30 min before), and post-treatment (5 min after) with PTX (600 ng/paw1, in 100 microL) reduced hypernociception induced by prostaglandin E(2) (100 ng/paw, in 100 microL, intraplantar). Furthermore, local intraplantar pre-treatment (30 min before) with PTX (600 ng/paw, in 100 microL) reduced hypernociception induced by DbcAMP, a stable analogue of cAMP (100 microg/paw, in 100 microL, intraplantar), which indicates that PTX may have an effect other than just G(i)/G(0) inhibition. PTX-induced analgesia was blocked by selective inhibitors of nitric oxide synthase (L-NMMA), guanylyl cyclase (ODQ), protein kinase G (KT5823) and ATP-sensitive K(+) channel (Kir6) blockers (glybenclamide and tolbutamide). In addition, PTX was shown to induce nitric oxide (NO) production in cultured neurons of the dorsal root ganglia. In conclusion, this study shows a peripheral antinociceptive effect of pertussis toxin, resulting from the activation of the arginine/NO/cGMP/PKG/ATP-sensitive K(+) channel pathway.