不同发育时期大鼠脑海马区胆固醇对发育与成熟的意义

背景胆固醇可使体外培养的神经元自身突触数量增加,囊泡数量及递质释放效能亦明显增高.目的建立大鼠脑海马组织中微量胆固醇的高效液相色谱检测方法,观察胆固醇在大鼠脑海马发育和成熟中的作用.设计重复测量实验.材料实验于2004-01/06在武汉大学人民医院神经科学研究室完成.选择健康雄性Wistar乳鼠(出生24 h内)、幼鼠(28～30 d,体质量40～60 g)和成鼠(90～100 d,体质量200～250 g)各10只.方法大鼠在麻醉状态下断头取海马称重,制备匀浆,离心,过滤后采用美国安捷伦公司反相色谱柱(ZORBAX XDB C18 4.6*250 mm 5 μm,Agilent),以乙腈-异丙醇(41)为流动相;流速0.8 mL/min;检测波长208nm.VARIAN Prostar 325紫外检测器和美国VARIAN公司的LCWORKSTATION V6.2色谱数据处理系统检测和处理色谱数据,获得胆固醇色谱图.样品中胆固醇含量(mg/g)=10×0.996 5×样品中胆固醇的峰面积×标准胆固醇的含量(g/L)÷标准胆固醇的峰面积主要观察指标①乳鼠、幼鼠和成鼠脑海马胆固醇含量与其色谱峰面积的关系.②乳鼠、幼鼠和成鼠脑海马胆固醇含量.结果30只大鼠均进入结果分析.①胆固醇含量在0.132 3～2.647 2 g/L时与其色谱峰面积呈良好的线性关系(r=0.999 93).②乳鼠、幼鼠和成鼠脑海马胆固醇含量3组样品的胆固醇含量分别为(2.92±0.03),(11.20±3.41),(12.91±1.25)mg/g,差别均有极显著性意义(P＜0.01).结论用高效液相色谱法检测大鼠脑海马组织中的微量胆固醇灵敏,特异性好,且分析时间短.在大鼠发育过程中,脑海马中胆固醇含量渐增,说明胆固醇对大鼠脑海马结构、功能的发育和成熟可能发挥了关键性的作用.

Effect of cholesterol on development and maturity in differently developing stage in rat hippocampus area

BACKGROUND: Cholesterol could induce the increase in the number ofsynapses per neuron, the increase in the number of the formation of synap-tic vesicles in vitro, and promoting evoked autaptic responses.OBJECTIVE: To establish high performance liquid chromatography(HPLC) method for determining trace cholesterol in rat hippocampus and tostudy the function of cholesterol during the stage of developing and matur-ing of rat hippocampus.DESIGN: Duplicated measured study.MATERIALS: The research was finished in the Department of Neurology,Remin Hospital of Wuhan University from January to June 2004. Respec-tively 10 male rats of the newborn (＜ 24 hours), the juvenile (28-30 days,weigh 40-60 g) and the adult (90-100 days, weigh 200-250 g) were selected.METHODS: Ten rats of each group were randomly taken and quicklykilled, and the brains were obtained. The Angelon Column of reversingphase high performance liquid chromatography (ZORBAX XDB C18, 4.6×15 cm, 5 μm, Agilent), the mobile phase consisted of acetonitrice-iso-propanol (4:1) at a flow-rate of 0.8 mL/minute, detected wave-length of208 nm, and 20 μL quantity of input were used. VARIAN Prostar 325 vio-let detector and LC WORKSTATION V6.2 color pattern data processing(VARIAN, USA) were used to obtain chromatigram ofcholesterol.terol concentration and its peak area in the hippocampus of mammal, juve-pus of mammal, juvenile and matured rats.tration was within 0.132 3-2.647 2 g/L which had a high linear relation-hippocampus of mammal, juvenile and matured rats: The cholesterol con-centration in each of the three samples was (2.92±0.03), (11.20±3.41),(12.91±1.25) mg/g respectively, and their difference being significant no-tably (P ＜0.01).CONCLUSION: This method has sufficient sensitivity, specificity andless separation time for determination of trace cholesterol in the hippocam-pus of rat. Cholesterol may play an important role during the stage of de-veloping and maturing of rat hippocampus.