Objective: To investigate the effect of long-term ethanol consumption on learning-memory functions in mice and the mechanisms involved.
Methods: Thirty male C57BL6/J mice were randomly assigned to 3 groups, with 10 mice in each group. The three groups included a control group in which the mice were given water ad libitum for 30 days, a long-term ethanol consumption group, or the EtOH group, in which the mice were given 6% (volume fraction) ethanol ad libitum for 30 days, and a long-term alcoholism group, or the EtOH+G group, in which the mice were given 5% (volume fraction) ethanol ad libitum for 30 days plus intermittent intragastric gavage of 20% ethanol at 3.5 g per kilogram body mass once every three days. After 30 days, the learning-memory functions of the mice were evaluated. At the conclusion of the experiment, the brain tissue of the mice was collected in order to examine the oxygen consumption rate (OCR) of mitochondria, the levels of pan-acetylation and protein oxidative stress in the hippocampal tissue, and the expression of sirtuin-3 (SIRT3) in hippocampus.
Results: Morris water maze test showed that, compared with those of the control group, the times of crossing the platform and the percentage of platform time in the EtOH group and the EtOH+G group were both lower, and the EtOH+G group had the lowest results ( P<0.05). Western blot results showed that long-term ethanol intake increased the levels of protein oxidative stress and pan-acetylation in the hippocampal tissue and down-regulated SIRT3 expression of hippocampal mitochondria. The results of mitochondrial complex Ⅱ respiration showed that the brain mitochondrial 3-state respiration in the EtOH group and the EtOH+G group was lower than that in the control group ( P<0.05). Compared that with the control group, the mitochondrial maximum respiration in EtOH+G group was decreased ( P<0.05).
Conclusions: Both long-term ethanol consumption and long-term alcoholism can reduce learning-memory functions and long-term alcoholism has the greater impact of the two. The potential mechanism may involve the down-regulation of the expression of SIRT3 protein in the hippocampus, which results in an increased level of pan-acetylation and enhanced expression of oxidative stress protein in the hippocampus, affects the mitochondrial functions of the brain, inhibits the oxidative phosphorylation capacity of mitochondrial complex Ⅱ, reduces the ATP energy supply of the brain tissue, and thus affects the learning-memory function.
目的: 探讨长期酒精摄入对小鼠学习记忆功能的影响及相关机制。
方法: 实验将30只雄性C57BL6/J小鼠随机分为3组，每组10只。对照（Control）组：30 d内自由饮水。长期饮酒（EtOH）组：30 d内自由饮体积分数6%的酒精。长期酗酒（EtOH+G）组：30 d内自由饮体积分数5%的酒精，并且每3 d一次的间歇性灌胃，小鼠按每次每公斤体质量灌胃3.5 g体积分数20%的酒精。30 d后，评估小鼠的学习记忆功能。实验完成后，提取脑组织，检测脑线粒体耗氧率、海马体泛乙酰化水平和蛋白氧化应激水平、海马体sirtuin-3（SIRT3）的表达。
结果: Morris水迷宫实验显示，与Control组相比，EtOH组和EtOH+G组穿越平台次数、平台时间百分比均降低，且EtOH+G组最低（P<0.05）。Western blot结果显示，长期酒精摄入可使海马组织泛乙酰化水平和蛋白氧化应激水平升高，而海马线粒体SIRT3表达降低。线粒体complex Ⅱ呼吸能力结果显示，与Control组相比，EtOH组和EtOH+G组脑线粒体3态呼吸均降低（P<0.05）；与Control组相比，EtOH+G组线粒体最大呼吸值降低（P<0.05）。
结论: 长期饮酒和长期酗酒均可降低学习记忆功能，且长期酗酒的影响更大，其机制可能是通过下调海马体SIRT3表达，引起海马体中泛乙酰化水平升高和蛋白氧化应激表达增强，影响脑线粒体功能，抑制线粒体complex Ⅱ氧化磷酸化产能，减少脑组织的ATP供能，从而影响学习记忆功能。
Keywords: Ethanol; Learning-memory function; Mitochondria; SIRT3.
Copyright© by Editorial Board of Journal of Sichuan University (Medical Sciences).