학술논문
吗啡预处理对小鼠海马脑片氧糖缺失/复氧复糖时PKCε膜转位的影响 / Effects of morphine preconditioning on oxygen-glucose deprivation and restoration induced-PKCε membrane translocation in hippocampal slices of mice
Document Type
Academic Journal
Source
中华麻醉学杂志 / CHINESE JOURNAL OF ANESTHESIOLOGY. 30(2):198-201
Subject
Language
Chinese
ISSN
0254-1416
Abstract
目的 探讨吗啡预处理对小鼠海马脑片氧糖缺失/复氧复糖(OGD/R)时蛋白激酶Cε(PKCε)膜转位的影响.方法 成年近交系BALB/C小鼠40只,体重18～22 g,雌雄不拘,制备海马脑片,将脑片置入无糖无氧的人工脑脊液中,培养20 min后再置于正常人工脑脊液(nACSF)中培养(复氧复糖)2h,以建立小鼠海马脑片OGD/R模型.脑片随机分为5组(n=40),对照组(C组):在nACSF中培养4 h;吗啡预处理组(M组):在含吗啡3μmol/L的nACSF中孵育30 min后用nACSF洗脱,随后制备OGD/R模型;纳洛酮+吗啡预处理组(N+M组):在含纳洛酮50 μmol/L的nACSF中孵育30 min后,加入吗啡至3 μmol/L,孵育30 min,随后制备OGD/R模型;纳洛酮组(N组):在含纳洛酮50μmol/L的nACSF中孵育1h,随后制备OGD/R模型;OGD/R组:制备OGD/R模型.于脑片OGD/R期间测定PKCε膜转位情况和神经元存活情况.结果 与C组比较,其余各组神经元存活率降低,膜相关蛋白中PKCε表达上调,胞溶蛋白中PKCε表达下调(P＜0.05或0.01);与OGD/R组比较,M组神经元存活率升高,膜相关蛋白中PKCε表达下调,胞溶蛋白中PKCε表达上调(P＜0.05),N+M组和N组上述指标差异无统计学意义(P＞0.05);与M组比较,N+M组神经元存活率降低,膜相关蛋白中PKCε表达上调,胞溶蛋白中PKCε表达下调(P＜0.05).结论 吗啡预处理诱导小鼠海马脑片缺血耐受的机制可能与抑制PKCε膜转位有关.
Objective To investigate the effects of morphine preconditioning on oxygen-glucose deprivation and restoration(OGD/R)-induced PKCε membrane translocation in hippocampal slices of mice.Methods Hippocampi were isolated from adult BALB/C mice of either sex weighing 18-22 g and sliced(400 μm thick)and incubated in normal artificial cerebro-spinal fluid(nACSF).The hippocampal slices were divided into5 groups(n = 40 each): group Ⅰ control(group C);group Ⅱ morphine preconditioning(group M);group Ⅲmorphine+naloxone(group N+M);group Ⅳ naloxone(group N)and group Ⅴ OGD/R.The slices were incubated in nACSF in the presence or absence of morphine 3.0 μmol/L for 30 min(group M and C).In group N+M,before being incubated with morphine the slices were incubated with naloxone 50μmol/L for 30 min.In group Ⅳ the slices were incubated with naloxone 50 μmol/L alone for 1 h.Slices were subjected to OGD for20 min followed by restoration of O2-glucose supply for 2 h in group Ⅱ-Ⅴ.The slices were homogenized,sonicated and centrifuged to separate the protein of particulate fraction from that of soluble fraction.SDS-PAGE Western blot was used to detect the PKCε membrane translocation in hippocampal slices.Results The PKCεmembrane translocation was significantly increased during OGD/R in OGD/R groups than in control group.Morphine preconditioning significantly decreased the PKCε membrane translocation during OGD/R.Naloxone completely blocked the inhibition of PKCε membrane translecation induced by morphine preconditioning but naloxone alone had no effect on PKCε membrane translocation induced by OGD/R.Conclusion Inhibition of PKCεmembrane translocation may be involved in the morphine preconditioning-induced cerebral ischemic tolerance.
Objective To investigate the effects of morphine preconditioning on oxygen-glucose deprivation and restoration(OGD/R)-induced PKCε membrane translocation in hippocampal slices of mice.Methods Hippocampi were isolated from adult BALB/C mice of either sex weighing 18-22 g and sliced(400 μm thick)and incubated in normal artificial cerebro-spinal fluid(nACSF).The hippocampal slices were divided into5 groups(n = 40 each): group Ⅰ control(group C);group Ⅱ morphine preconditioning(group M);group Ⅲmorphine+naloxone(group N+M);group Ⅳ naloxone(group N)and group Ⅴ OGD/R.The slices were incubated in nACSF in the presence or absence of morphine 3.0 μmol/L for 30 min(group M and C).In group N+M,before being incubated with morphine the slices were incubated with naloxone 50μmol/L for 30 min.In group Ⅳ the slices were incubated with naloxone 50 μmol/L alone for 1 h.Slices were subjected to OGD for20 min followed by restoration of O2-glucose supply for 2 h in group Ⅱ-Ⅴ.The slices were homogenized,sonicated and centrifuged to separate the protein of particulate fraction from that of soluble fraction.SDS-PAGE Western blot was used to detect the PKCε membrane translocation in hippocampal slices.Results The PKCεmembrane translocation was significantly increased during OGD/R in OGD/R groups than in control group.Morphine preconditioning significantly decreased the PKCε membrane translocation during OGD/R.Naloxone completely blocked the inhibition of PKCε membrane translecation induced by morphine preconditioning but naloxone alone had no effect on PKCε membrane translocation induced by OGD/R.Conclusion Inhibition of PKCεmembrane translocation may be involved in the morphine preconditioning-induced cerebral ischemic tolerance.