Jamal M, Ameno K, Ameno S, Morishita J, Wang W, Kumihashi M, Ikuo U, Miki T, Ijiri I
Department of Forensic Medicine, Faculty of Medicine, Kagawa University, 1750-1, Ikenobe, Miki, Kita, Kagawa 761-0793, Japan.
Neuroscience. 2007 Jan 5;144(1):232-8. doi: 10.1016/j.neuroscience.2006.08.066. Epub 2006 Oct 11.
Our previous microdialysis study demonstrated that both ethanol (EtOH) and acetaldehyde (ACe) decrease in vivo acetylcholine (ACh) release in the medial frontal cortex of freely moving rats. To better understand the mechanisms of EtOH and ACe's effects on the cholinergic system in the brain, choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) expression was examined at 40 and 240 min after a dose of EtOH (1 g/kg) in the rat frontal cortex and hippocampus. The control group was treated with 0.9% saline, and other groups received EtOH or cyanamide (CY, 50 mg/kg, a potent aldehyde dehydrogenase inhibitor) and 60 min later by EtOH intraperitoneally. Reverse-transcription polymerase chain reaction (RT-PCR) analysis revealed that ChAT mRNA levels were decreased by 72.8% and 71.6% in the EtOH and CY+EtOH groups, respectively, at 40 min after EtOH injection compared with saline in the frontal cortex. The hippocampal ChAT levels were reduced by 76.5% and 53.0% in the EtOH and CY+EtOH groups, respectively, at this time. CY+EtOH-induced depletion in ChAT mRNA levels was markedly higher than EtOH in the hippocampus. A similar decrease pattern of ChAT was observed at protein levels as determined by Western blot, but the reduced ChAT levels were significantly higher in the CY+EtOH group as compared with the EtOH group both in the frontal cortex and hippocampus. At 240 min after EtOH injection, the EtOH group had no effect on ChAT at mRNA levels, as compared with saline, whereas CY+EtOH group induced a significant decrease in ChAT mRNA expression to 62.0% and 65.5% in the frontal cortex and hippocampus, respectively. These data were consistent with the results of the Western blot analysis. AChE expression at mRNA levels was not changed at either 40 or 240 min after EtOH dosing in either of these groups in the frontal cortex and hippocampus. Within 40 and 240 min, a statistically significant difference in ChAT expression at mRNA and protein levels was found in the EtOH and CY+EtOH groups both in the frontal cortex and hippocampus. The data obtained from this study demonstrate that EtOH and ACe concentrations decreased ChAT expression at 40 and 240 min after EtOH administration in the frontal cortex and hippocampus, and this result suggests that reduced ChAT expression is strongly related to a decrease in ACh release in the rat brain.
我们之前的微透析研究表明，乙醇（EtOH）和乙醛（ACe）均可降低自由活动大鼠内侧前额叶皮质中体内乙酰胆碱（ACh）的释放。为了更好地理解EtOH和ACe对大脑胆碱能系统作用的机制，在给大鼠腹腔注射一剂EtOH（1 g/kg）后的40分钟和240分钟，检测了前额叶皮质和海马中胆碱乙酰转移酶（ChAT）和乙酰胆碱酯酶（AChE）的表达。对照组用0.9%生理盐水处理，其他组接受EtOH或氰胺（CY，50 mg/kg，一种有效的乙醛脱氢酶抑制剂），60分钟后腹腔注射EtOH。逆转录聚合酶链反应（RT-PCR）分析显示，与前额叶皮质中的生理盐水组相比，在注射EtOH后40分钟，EtOH组和CY + EtOH组的ChAT mRNA水平分别降低了72.8%和71.6%。此时，EtOH组和CY + EtOH组海马中的ChAT水平分别降低了76.5%和53.0%。CY + EtOH诱导的ChAT mRNA水平的降低在海马中明显高于EtOH组。通过蛋白质印迹法测定，在蛋白质水平上观察到ChAT有类似的降低模式，但在额叶皮质和海马中，CY + EtOH组ChAT水平的降低明显高于EtOH组。在注射EtOH后240分钟，与生理盐水组相比，EtOH组在mRNA水平上对ChAT没有影响，而CY + EtOH组在前额叶皮质和海马中分别导致ChAT mRNA表达显著降低至62.0%和65.5%。这些数据与蛋白质印迹分析的结果一致。在额叶皮质和海马的这些组中，在给EtOH后40分钟或240分钟，AChE在mRNA水平上的表达均未改变。在40分钟和240分钟内，在额叶皮质和海马的EtOH组和CY + EtOH组中，ChAT在mRNA和蛋白质水平上的表达均存在统计学上的显著差异。从本研究中获得的数据表明，在给EtOH后40分钟和240分钟，EtOH和ACe浓度降低了前额叶皮质和海马中ChAT的表达，这一结果表明ChAT表达的降低与大鼠脑中ACh释放的减少密切相关。
