摘要目的:探讨左旋肉碱(L-carnitine，LCAR)对慢性脑低灌注大鼠认知功能障碍的保护作用和机制。方法:90只3~4月龄雄性SPF级SD大鼠按照随机数字表法分为4组：假手术对照组(SHAM组， n＝15)、假手术左旋肉碱组(LCAR组， n＝25)、双侧颈总动脉结扎组(2VO组， n＝25)、双侧颈总动脉结扎+左旋肉碱组(2VO+LCAR组， n＝25)。采用双侧颈总动脉结扎法建立慢性脑低灌注模型，假手术组只分离血管，不结扎；LCAR组和2VO+LCAR组在手术后按照300 mg·kg -1·d -1的剂量连续给予左旋肉碱腹腔注射给药，共注射30 d。左旋肉碱干预30 d后，用Morris水迷宫实验检测大鼠的空间学习和记忆能力，用化学发光法检测海马组织ATP水平，通过透射电镜观察海马神经元的线粒体结构和突触结构，对线粒体损伤程度进行评分，计数和测算囊泡密度，用Western blot法检测海马组织的N-甲基-D-天冬氨酸受体功能亚单位(N-methyl-D-aspartate receptor subunit 2A or 2B，NR2A/B)、突触后致密蛋白95(postsynaptic density 95，PSD95)水平，用免疫荧光观察转录因子环磷酸腺苷反应元件结合蛋白(cAMP response element-binding protein，CREB)在脑组织中的分布和表达水平。采用SPSS 16.0软件进行统计分析，大鼠水迷宫7 d重复学习训练的逃避潜伏期数据采用重复测量方差分析，其他多组间数据比较采用单因素方差分析，进一步两两比较采用Dunnett's t检验。 结果:(1)Morris水迷宫实验结果显示，4组大鼠逃避潜伏期的分组与时间的交互作用不显著( F＝1.4， P>0.05)，但时间主效应和组别主效应均显著( F＝21.6，15.2，均 P<0.05)。在第3~7天的平台位置学习训练中，2VO组大鼠逃避潜伏期长于SHAM组和2VO+LCAR组(均 P<0.05)。大鼠短期记忆检测结果显示，2VO组大鼠逃避潜伏期长于SHAM组和2VO+LCAR组(均 P<0.05)；2VO大鼠的平台区域穿梭次数和滞留时间均少于SHAM组和2VO+LCAR组(均 P<0.05)。(2)海马组织ATP水平结果显示，4组大鼠海马组织的ATP绝对水平和相对水平均差异有统计学意义( F＝14.6，13.2，均 P<0.05)。2VO组大鼠海马组织ATP水平低于SHAM组和2VO+LCAR组(均 P<0.05)。电镜观察线粒体形态显示，2VO组大鼠海马组织线粒体损伤的Flameng评分[(2.82±0.17)分]高于SHAM组[(0.25±0.07)分]和2VO+LCAR组[(1.76±0.09)分](均 P<0.05)。(3)2VO组大鼠海马神经元突触囊泡密度[(289.09±22.41)个/μm 2]显著低于SHAM组[(497.49±28.89)个/μm 2]和2VO+LCAR组[(401.23±45.09)个/μm 2](均 P<0.01)。Western blot结果显示，2VO海马组织突触蛋白NR2A/B、PSD95和CREB相对水平低于SHAM组和2VO+LCAR组(均 P<0.05)。免疫荧光结果显示，2VO海马亚区和皮质CREB表达相对水平低于SHAM组和2VO+LCAR组(均 P<0.01)。 结论:左旋肉碱治疗能够改善慢性脑低灌注大鼠空间学习和记忆功能障碍，可能与促进ATP生成和保护线粒体形态，以及促进突触囊泡合成和突触蛋白表达有关。

abstractsObjective:To explore the protective effects and mechanisms of L-carnitine (LCAR) on cognitive dysfunction in chronic cerebral hypoperfusion rats.Methods:Totally 90 SD male rats (SPF class) aged 3-4 months were divided into four groups according to random number talbe: sham operated control group (SHAM group, n=15), sham operated with L-carnitine treatment group (LCAR group, n=25), 2-vessel occlusion group (2VO group, n=25), and 2-vessel occlusion with L-carnitine treatment group (2VO+ LCAR group, n=25). The chronic cerebral hypoperfusion model was established by bilateral common carotid artery ligation, and the carotid arteries from SHAM group and LCAR group were only separated without ligation.L-carnitine was administered intraperitoneally (300 mg·kg -1·d -1) for 30 days after surgery in the LCAR and 2VO+ LCAR groups.After 30 days of L-carnitine intervention, Morris water maze was performed to test the spatial cognitive function of the rats, the ATP level of hippocampal tissue was detected by chemiluminescence, the mitochondrial structure and synaptic structure of hippocampal neurons were observed by transmission electron microscopy, the degree of mitochondrial damage was scored, the vesicle density was counted and measured, the level of N-methyl-D-aspartate receptor subunit 2A or 2B(NR2A/B) and postsynaptic density 95(PSD95) in hippocampal tissue were detected by Western blot.The expression and distribution levels of transcription factor cAMP response element-binding protein(CREB) in brain tissues were observed by immunofluorescence.SPSS 16.0 software was used for statistical analysis.The escape latency data of repeated learning training in Morris water maze was conducted by repetitive measurement ANOVA, while other data were adopted by one-way ANOVA, and Dunnett's t test was used for further pairwise comparison. Results:(1)Morris water maze results showed that the time and group interaction of escape latency was not significant among the 4 groups of rats ( F=1.4, P>0.05), but the time main effect and group main effect were significant( F=21.6, 15.2, both P<0.05). Morris water maze results showed that platform position learning from 3rd to 7th day, the escape latencies in 2VO group were longer than those in SHAM group and 2VO+ LCAR group (all P<0.05). The results of short-term memory showed that the escape latency in 2VO group was longer than those in SHAM group and 2VO+ LCAR group (all P<0.05). Meanwhile, the retention time and crossing times in the platform area of 2VO group were less than those in SHAM group and 2VO+ LCAR group (all P<0.05). (2) The absolute and relative levels of ATP in hippocampus showed that the difference among the 4 groups were statistically significant ( F=14.6, 13.2, both P<0.05). ATP level of hippocampus in 2VO group was lower than those in SHAM group and 2VO+ LCAR group (both P<0.05). Electron microscopic observation of mitochondrial morphology showed that the Flameng score of mitochondrial damage in the hippocampus of rats in 2VO group (2.82±0.17) was higher than those in SHAM group (0.25±0.07) and 2VO+ LCAR group (1.76±0.09) (both P<0.05). (3) The density of synaptic vesicles in the hippocampus of rats in 2VO group ((289.09±22.41)/μm 2)was lower than those in SHAM group ((497.49±28.89)/μm 2)and 2VO+ LCAR group ((401.23±45.09)/μm 2) (both P<0.01). Western blot results showed that the relative levels of synaptic proteins NR2A/B, PSD95 and CREB in 2VO group were lower than those in SHAM group and 2VO+ LCAR group (all P<0.05). Immunofluorescence results showed that the relative level of CREB expression in hippocampal subregions and cortex in 2VO group was lower than those in SHAM group and 2VO+ LCAR group (both P<0.01). Conclusion:L-carnitine can improve spatial learning and memory dysfunction in rats with chronic cerebral hypoperfusion, which are related with promoting ATP production and protecting mitochondrial morphology, and promoting synaptic vesicle synthesis and synaptic protein expression.