不同浓度七氟醚可双向调节大鼠学习记忆及海马ARC蛋白表达

Sevoflurane Inhalation at Different Dosage Regulate Hippocampal ARC Protein Expression and Memory Formation Bidirectionally

背景：术中知晓和术后认知功能障碍是困扰麻醉医师的棘手问题。以往多认为与全身麻醉药物对神经系统的影响有关。但近来不断有实验研究证实．低浓度吸入麻醉气体在特定环境下反而会兴奋大脑功能．甚至产生脑保护的作用。显然全麻药物对大脑认知功能的影响机制仍未明瞭。全身麻醉药物七氟醚已广泛应用于各类临床手术．其对哺乳类动物中枢神经系统是否及如何产生影响是我们一直在探讨的课题。细胞骨架蛋白ARC可在大脑海马结构大量表达．已证实可依据其表达情况来考察神经元的活性．以及突触可塑性的变化。目前认为其表达程度可作为检测学习记忆形成的指标。近来有学者发现，较低浓度的七氟醚可引发中枢神经兴奋现象．并猜测亚麻醉剂量对于中枢神经系统具潜在保护作用。为进一步证实这一现象．本研究拟结合ARC蛋白和抑制性逃避（inhibitory avoidance．IA）这一行为学实验来进行深入探讨。方法：250－300g雄性SD大鼠随机分为3组：空白组，极低浓度七氟醚吸入组（0．11％SEV）和低浓度七氟醚吸入组（0．3％SEV）。依据分组依次将大鼠安置于连接有低流量（0．5L／min）闭合环路吸入麻醉系统的特制容器内，分别给予吸入空气，0．11％SEV（0．05MAC）和0．3％SEV（0．14MAC）各45min。结束后马上进行单次IA训练（0．4mA,2s），24小时后进行IA记忆（潜伏期）检测。另取一批大鼠在IA初次训练后45min处死取材．以western-blotting方法检测脑内海马ARC蛋白的表达水平．以real—timePCR方法检测ARC的mRNA转录水平。结果：与吸入空气组相比。0．11％sEV组的24小时IA记忆潜伏期明显延长，而0．3％SEV组的24小时IA记忆潜伏期缩短：相应地．0．1196SEV组的海马ARC蛋白表达增加．而0．30％SEV组ARC蛋白的表达减少。但ARC的mRNA水平却始终无显著差异。结论：吸入不同浓度的七氟醚能对大鼠的学习记忆能力产生双相作用．这种现象伴有海马蛋白ARC表达的相应改变．却没有相应mRNA水平的改变。提示全麻药物七氟醚对于记忆的双向调节存在即可早期基因如NARC对神经元突触可塑性的作用．但这种干预可能体现在转录后水平。

Background： Central nervous system （CNS） disorder such as intra-operative awareness and post operative cognitive dysfunction are major concerns of anesthesiologists, general anesthetics have been discovered to have controversial impact on CNS. Studies showed that low dose of inhalational gas may excite the brain function in specialized environment, this is much different from their negative effect at clinical dosage and therefore the exact modulating mechanism is still unclear. Sevoflurane is widely applied in routine operation room work, so how this agent affect the CNS is becoming a popular topic. Activity-regulated cytoskeleton protein （Arc） can be fastly expressed in hippocampus, and evidence has shown that it is a powerful factor for investigating the activity of neuronal work, even the modulation of synaptic plasticity. Recently, researchers have discovered that lower dose sevoflurane would have neuronal excitatory effect in CNS, and thus proposing that certain subanesthetic dose of sevoflurane can display a memory enhancing phenomenon. To further investigate it, we conducted the following study by associating Arc protein expression and ＂inhibitory avoidance＂ （IA） behavioral training. Methods：250-300g, male SD rats were randomly assigned into three groups： sham group, 0.11% SEV group and 0.3% SEV group. Anesthesia was given by target dosage of sevofiurane for 45min and IA training （0.4mA, 2s） was given to every subject immediately after inhalation. The memory retention latency was observed 24hs after. Another serial of rats were killed for hippocampal tissue after first IA training, both western-blotting and real-time PCR for Arc mRNA level were applied. Results： Statistical difference of the 24-h inhibitory avoidance memory retention performance was compared among groups. 0.11% SEV group displayed a significant elevation of memory retention while 0.3% SEV group showed descendent of memory retention, both compared with the sham （air） group. PCR analysis of relative Arc mRNA levels showed that subanesthetic doses of sevoflurane inhalation did not change Arc transcription level, when compared to sham group. 0.11% sevofiurane significantly increased Arc protein in the hippocampus, while 0.3% sevofiurane reversed this phenomenon and even suppress the Arc expression （＊ P 〈 0.05, compared with the sham group）. Conclusion： Inhalation of different subanesthetic dosage of sevotflurane have bidirectional regulation on rat＇s learning and memory function. Arc protein level varies according to this regulation while Arc mRNA level keeps still. This demonstrates that hippocampal ARC protein expression influences the sevoflurane induced bidirectional regulation of memory, potentially in a posttranslational level.