慢性脑低灌注致大鼠持续性学习记忆受损及海马神经血管单元改变

Chronic cerebral hypoperfusion facilitates lasting learning and memory impairment and induces hippocampal neurovascular unit dysfunction in rats

目的观察在大鼠慢性脑低灌注(chronic cerebral hypoperfusion,CCH)模型中海马神经血管单元变化的特点,探讨海马神经血管单元变化与CCH致大鼠学习记忆受损的关系。方法取220~250 g SPF级8周龄SD雄性大鼠44只,按随机数字表法分为CCH组(n=24)和对照组(n=20)。CCH组通过永久性结扎双侧颈总动脉建立CCH模型,对照组不结扎。各组大鼠分别于术后4、12周通过旷场实验、物体辨别实验和Morris水迷宫实验检测自主探索行为反应和学习记忆功能,双重免疫荧光标记检测大鼠海马星形胶质细胞足突水通道蛋白-4(aquaporin-4,AQP4)和血管基底膜CollagenⅣ的共表达,透射电镜观察大鼠海马神经血管单元超微结构改变。结果与对照组比较,4周和12周CCH组大鼠:(1)物体辨别指数明显下降[(0.86±0.14)vs(0.71±0.16),P<0.05;(0.83±0.19)vs(0.64±0.10),P<0.05],定位航行及空间探索逃避潜伏期均显著延长(P<0.05);(2)海马星形胶质细胞足突AQP4[(1.94±0.80)vs(1.10±0.41),P<0.05;(2.05±0.51)vs(0.96±0.50),P<0.01]、血管基底膜CollagenⅣ[(6.90±2.42)vs(4.25±1.01),P<0.05;(6.63±2.20)vs(3.66±1.22),P<0.01]表达显著降低,二者的共表达比例[(0.94±0.02)vs(0.91±0.01),P<0.01;(0.91±0.01)vs(0.89±0.01),P<0.01]均明显减少;(3)超微结构显示海马星形胶质细胞足突与血管基底膜明显解离,胶质血管连接及神经血管单元明显破坏。结论慢性脑低灌注所致大鼠持续性学习记忆功能受损可能与海马神经血管单元的破坏有关。

Objective To investigate the characteristics of changes in hippocampal neurovascular unit in rats after chronic cerebral hypoperfusion（ CCH）,and explore the relationship of the changes of the unit with learning and memory impairment induced by CCH. Methods A total of 44 male adult SD rats（ 220 ~250 g） were randomly divided into： CCH group（ n = 24） and control group（ n = 20）. CCH was induced by permanent ligation of the bilateral carotid artery. Open field test,object recognition test and Morris water maze test were used to evaluate exploratory behavior and learning and memorial function at 4 and 12 weeks after the operation for each group of rats. Then double immunofluorescence labeling was used to observe the coexpression of aquaporin-4（ AQP4） in astrocytic endfeet and vascular basement membrane of collagen Ⅳ in the hippocampus. Furthermore, transmission electron microscopy was used to observe the ultrastructure of neurovascular unit in the hippocampus. Results Compared with the control group,in CCH groups in 4 and12 weeks after the operation,（1） the object discrimination index was decreased clearly（ 0. 86 ± 0. 14 vs 0. 71 ±0. 16,P〈0. 05; 0. 83 ± 0. 19 vs 0. 64 ± 0. 10,P〈0. 05）,the escape latencies of place navigation test and spatial probe test were prolonged notably（ P〈0. 05）;（2） the expression of AQP4 in astrocytic endfeet（ 1. 94 ± 0. 80 vs 1. 10 ± 0. 41,P〈0. 05; 2. 05 ± 0. 51 vs 0. 96 ± 0. 50,P〈0. 01）,that of collagen Ⅳ in vascular basement membrane（ 6. 90 ± 2. 42 vs 4. 25 ± 1. 01,P〈0. 05; 6. 63 ± 2. 20 vs 3. 66 ± 1. 22,P〈0. 01）,and the co-expression ratio of them（ 0. 94 ± 0. 02 vs 0. 91 ± 0. 01,P〈0. 01; 0. 91 ± 0. 01 vs 0. 89 ±0. 01,P〈0. 01） were all decreased obviously;（3） the ultrastructure of neurovascular unit displayed that astrocytic endfeet were separated from the vascular basement membrane, and gliovascular interface and neurovascular unit was damaged in the hippocampus. Conclusion CCH facilitates the lasting learning and memory impairment,which may be associated with neurovascular unit dysfunction.