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大量饮酒及乙醇戒断后对大鼠脑内维生素C 水平的影响
田华刘富杨宏艳宋娟苑家鑫张奇柏青杨崔红霞
0
()
摘要:
目的 研究大量饮酒4 d及乙醇戒断后大鼠脑脊液中及各重要脑区内维生素C(VC)水平 及钠依赖性VC 转运体2(SVCT2)蛋白水平的变化。方法 30 只健康Wistar 大鼠随机分为5 组,对照组 (A 组)、大量饮酒4 d(B 组)、大量饮酒4 d 后戒断1 d(C 组)、大量饮酒4 d 后戒断2 d(D组)、大量饮酒 4 d后戒断7 d(E组),每组6只。B、C、D、E组大鼠大量灌胃给予乙醇4 d,乙醇浓度为25% W/V,每8小时 灌胃1次,连续4 d;A组给予等体积蒸馏水。采用Y迷宫实验评价大鼠的空间工作记忆能力;以高效液相- 电化学法(HPLC-ECD)检测大鼠脑脊液及前额叶皮质、顶叶皮质、颞叶皮质、海马脑区内细胞内VC 含量, 免疫印迹法检测大鼠各脑区内SVCT2蛋白水平的变化。结果 (1)Y迷宫实验: C、D组大鼠自发交替反应 率分别为(24.64±15.11)% 和(41.48±13.01)%,均显著低于A 组[(75.47±8.61)%]; E 组大鼠的自发交替 反应率恢复接近正常[(61.64±11.51)%],与A 组差异无统计学意义(P> 0.05)。(2)脑脊液内VC 含量:B 组大鼠脑脊液内VC 含量为(204.54±25.51)μmol/L,显著高于A 组[(145.57±18.98)μmol/L], 而C、D 组 大鼠脑脊液内VC 水平[分别为(90.24±15.45)、(86.93±14.53)μmol/L]明显低于A、B 组,差异均有统计 学意义(均P< 0.001);E 组大鼠脑脊液中VC 含量[(135.80±17.16) μmol/L]与A 组间差异无统计学意义 (P> 0.05)。(3)各脑区内VC 水平:B 组前额叶皮质、顶叶皮质、海马脑区脑组织匀浆(细胞内)VC 水 平变化趋势相同[分别为(1.18±0.13)、(1.14±0.12)、(1.20±0.20)μmol/g],均显著低于A 组[分别为 (1.64±0.11)、(1.62±0.13)、(2.06±0.27)μmol/g];C 组在乙醇戒断后,前额叶皮质、顶叶皮质、海马脑区 细胞内VC 水平[分别为(1.20±0.29)、(1.05±0.06)、(1.21±0.15)μmol/g]有所恢复,但仍明显低于A 组 (均P< 0.01);C组大鼠颞叶皮质细胞内VC水平[(1.37±0.04)μmol/g]显著高于顶叶皮质(P<0.05);D组 各脑区细胞内VC水平均有回升趋势,前额叶皮质细胞内VC水平显著高于顶叶皮质[(1.63±0.24)μmol/g 比(1.26±0.16)μmol/g,P<0.05];E组前额叶皮质、顶叶皮质、颞叶皮质、海马区细胞内VC水平[分别为 (1.72±0.19)、(1.43±0.22)、(1.67±0.19)、(1.86±0.22)μmol/g]均较B组显著升高(均P<0.01),逐渐恢复至正 常水平,海马区VC水平显著高于顶叶皮质(P<0.05)。(4)Western blot结果显示,与A组比较,B、C、D组大 鼠前额叶皮质、顶叶皮质及C、D、E组大鼠海马区的SVCT2蛋白均显著升高(均P<0.05)。结论 大量饮 酒4 d可致大鼠脑损伤,乙醇干预后大鼠脑脊液中及各脑区脑组织匀浆中VC水平呈现负相关的变化过程, 此过程中SVCT2蛋白表达上调,这将有助于脑脊液内的VC被转运至各脑区神经元内发挥抗氧化作用。
关键词:  维生素C  维生素C 转运体2  饮酒  大脑皮层  海马
DOI:10.3969/j.issn.1009-6574.2021.06.002
基金项目:黑龙江省自然科学基金资助项目(H2017079);国家自然科学基金青年项目(81803757)
Effects of binge drinking ethanol and ethanol withdrawal on vitamin C levels in rat brain
Tian Hua, Liu Fu, Yang Hongyan, Song Juan, Yuan Jiaxin, Zhang Qi, Bai Qingyang, Cui Hongxia
()
Abstract:
Objective To study the changes of vitamin C( VC) levels and sodium vitamin C transporter 2( SVCT2) protein levels in cerebrospinal fluid( CSF) and important brain regions of rats after 4 days of binge drinking and ethanol withdrawal. Methods A total of 30 healthy wistar rats were randomly divided into 5 groups: control group( Group A), binge drinking for 4 days( Group B), withdrawal 1 day after 4-day binge drinking( Group C), withdrawal 2 days after 4-day binge drinking( Group D), withdrawal 7 days after 4-day binge drinking( group E). There were 6 rats in each group. Rats in group B, C, D and E were given ethanol by gavage for 4 days at a concentration of 25% W/V and gavage once every 8 hours for 4 days. Group A was given equal-volume distilled water. Y maze experiment was used to evaluate the spatial working memory of rats. The concentration of intracellular VC in cerebrospinal fluid, prefrontal cortex, parietal cortex, temporal cortex and hippocampus of rats in each group was detected by HPLC combined with electrochemical detector( HPLC-ECD), and the protein level of SVCT2 in each brain area of rats was detected by Western blotting. Results In the Y maze experiment, the rate of spontaneous response alternations of rats in group C and D was( 24.64±11.51)% and( 41.48±13.01)%, respectively, which were significantly lower than that in group A( 75.47±8.61)%. The rate of spontaneous response alternations of rats in group E( 61.64±15.11)% was close to normal, and there is no statistically significant difference between group E and group A( P< 0.05). The content of VC in CSF of rats in group B was( 204.54±25.51) μmol/L, which was significantly higher than that in group A( 145.57±18.98) μmol/L. The CSF VC levels in group C and group D were( 90.24±15.45)μmol/L and( 86.93±14.53) μmol/L, respectively, lower than those in group A and group B, and the differences were statistically significant( P < 0.001). The CSF VC level of group E was (135.80±17.16)μmol/L. There was no significant difference in the content of VC in CSF between group A and group E( P< 0.05). The VC levels in brain tissue homogenates( intracellular) in prefrontal cortex, parietal cortex and hippocampus showed the same trend in group B, which were( 1.18±0.13) μmol/g,( 1.14±0.12) μmol/g, (1.20±0.20) μmol/g, respectively, and were all significantly lower than those in group A[ (1.64±0.11) μmol/g, (1.62±0.13) μmol/g,( 2.06±0.27) μmol/g]. The VC levels in brain tissue homogenates( intracellular) in prefrontal cortex, parietal cortex and hippocampus of group C after ethanol withdrawal[ (1.20±0.29) μmol/g, (1.05±0.06) μmol/g,( 1.21±0.15) μmol/g, respectively]recovered to some extent, but still significantly lower than those of group A( P< 0.01). In group C, the level of VC in temporal cortex[ (1.37±0.04) μmol/g] was significantly higher than that in parietal cortex( P < 0.05). In group D, the level of VC in all brain regions increased, and the level of VC in prefrontal cortex was significantly higher than that in parietal cortex [(1.63±0.24) μmol/g vs( 1.26±0.16) μmol/g, P < 0.05]. The levels of VC in prefrontal cortex, parietal cortex, temporal cortex and hippocampus in group E were( 1.72±0.19) μmol/g,( 1.43±0.22) μmol/g, (1.67±0.19) μmol/g and( 1.86±0.22)μmol/g, respectively, significantly higher than those in group B( all P < 0.01), and gradually returned to normal level. The level of VC in hippocampus was significantly higher than that in parietal cortex( P<0.05). Western blot results showed that compared with group A, the protein of SVCT2 in prefrontal cortex, parietal cortex of group B, group C and group D, and in hippocampus of group C, group D and group E was significantly increased( P<0.05). Conclusions Binge drinking for 4 days can cause brain damage in rats. The VC level in CSF and brain tissue homogenates of each brain region present a negative correlation dynamic change process after ethanol treatment. During ethanol treatment process, the expression of SVCT2 protein is up-regulated, which will help the VC in the CSF to be transported to the neurons in each brain region to play an antioxidant role.
Key words:  Vitamin C  Vitamin C transporter 2  Alcohol drinking  Cerebral cortex  Hippocampus

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