Polysomal and total RNA in controls and after 10 min global brain ischemia and 8 hours of reperfusion. Rattus norvegicus

Complete global brain ischemia (CGBI) and reperfusion occur following resuscitation from cardiac arrest. Different brain neurons are selectively vulnerable to CGBI: pyramidal neurons of hippocampal CA3 survive 10 min CGBI but those of CA1 die at 3 days following 10 min CGBI. CA3 neurons are expected to have more robust stress responses and repair responses than CA1 neurons. We used microarrays to compared total and polysome-bound mRNAs in CA1 and CA3 at 8 hr reperfusion after 10 min CGBI in Long Evans male rats to ascertain differences in total vs polysome-bound gene expression. Overall design: Male Long Evans rats were subjected to (1) sham operation (non-ischemic control, NIC) or normothermic CGBI of 10 min followed by 8 hr reperfusion (8R). Hippocampal CA1 and CA3 were dissected. n = 5 CA1 or CA3 were pooled to give a single replicate and there were 3 or 4 replicates per group. Post-mitochondrial supernatant (PMS) was prepared. Twenty percent of PMS was TRIzol extracted to give total RNA. The remainder was run on a 20% sucrose pad to isolate polysome pellets, which were also TRIzol extracted to give polysome RNA. Total and polysome RNA were then run on Affymetrix Rat Gene 2.0 microarrays.

心脏骤停复苏后，可发生完全性全脑缺血（Complete global brain ischemia, CGBI）及再灌注事件。不同脑神经元对CGBI具有选择性易损性：海马CA3区的锥体神经元可耐受10分钟的CGBI，而CA1区锥体神经元则会在缺血10分钟后的第3天发生死亡。相较于CA1区神经元，CA3区神经元被认为拥有更为强大的应激反应与修复反应能力。本研究使用微阵列（microarray）技术，对雄性Long Evans大鼠在10分钟CGBI后再灌注8小时的海马CA1与CA3区组织中，总mRNA与多聚体结合mRNA的表达水平进行比较，以明确总mRNA与多聚体结合mRNA的基因表达差异。

实验整体设计如下：
雄性Long Evans大鼠被分为两组：（1）假手术组（非缺血对照，NIC）；（2）10分钟常温完全性全脑缺血后再灌注8小时组（记为8R组）。分离获取海马CA1与CA3区组织。将每5份CA1或CA3区组织样本合并为一个生物学重复，每组设置3至4个生物学重复。制备线粒体后上清液（post-mitochondrial supernatant, PMS）：取20%的PMS使用TRIzol试剂提取总RNA；剩余PMS经20%蔗糖垫离心分离得到多聚体沉淀，同样使用TRIzol试剂提取多聚体RNA。将总RNA与多聚体RNA分别在Affymetrix Rat Gene 2.0微阵列芯片上进行表达谱检测。