生物素葡聚糖胺示踪观察脊髓腹侧损伤大鼠模型下行传导通路的改变

摘要

目的 观察大鼠脊髓腹侧损伤后下行传导通路的改变.方法 清洁级雌性SD大鼠28只,随机分为实验对照组(6只)和脊髓腹侧损伤15、30、50 g 3个小组(7、7、8只),使用标准化脊髓损伤动物模型实验装置在T9～T10水平压迫脊髓腹侧制备动物模型,对照组大鼠仅咬除椎弓、横突,不损伤脊髓.术后12h,所有大鼠立体定向开颅,生物素葡聚糖胺(BDA)注射于大脑皮层,并采用改良的Tarlov分级评分、斜板实验及BBB评分评价实验动物的运动功能.BDA注射2周后取出损伤节段以下脊髓组织,行BDA染色成像.统计学方法采用双因素方差分析或秩和检验.结果 脊髓损伤组大鼠改良的Tarlov分级评分、斜板实验及BBB评分均明显低于对照组(P＜0.05)；增加砝码质量,使损伤组大鼠受到的腹侧向上提拉力量增大,双下肢改良的Tarlov分级评分逐渐下降,鼠斜板实验临界角逐渐下降,BBB评分呈降低趋势；BDA神经示踪后免疫组织化学显色后阳性的神经元呈棕褐色深染,脊髓损伤组较对照组明显减少,且随着损伤外力的加大,阳性神经元数量明显减少.结论 脊髓腹侧受损后大脑中枢下行传导通路神经传导异常程度越重,其造成的功能障碍程度也越重.%Objective To determine the changes in descending neuronal pathways in rat models with ventral spinal cord injury. Methods Twenty-eight clean-grade female Sprague-Dawley rats were randomly allocated into Sham group (n=6) and 15 g (n=6-8), 30 g (n=6-8) and 50 g ventral spinal cord injury group (n=6-8), respectively. The animal models were constructed by compressing the ventral spinal cord at T9～T10 via a standardized instrument for testing spinal cord injury. In Sham group, both the vertebral arch and horizontal axon, but not the spinal cord, were removed. At hour 12 postoperatively, all rats were subjected to three-dimensional directed craniotomy followed by biotin dextran amine (BDA) injection in the cortex. Modified Tarlov grading, ramping exercise and BBB grading were employed for assessment of the exercise capacity. At week 2, all rats were sacrificed for extraction of spinal cord tissues inferiorly and further staining of BDA. Two-way ANOVA or rank sum test was adopted for statistical analysis. Results Rats as having spinal cord injury yielded significantly lower modified Tarlov grading, ramping exercise and BBB scores compared with control group (P＜0.05). Increased mass of the weight resulted in stronger ventral pull in rats with spinal cord injury. This was associated with a gradual decline in modified Tarlov grading for lower extremities and critical angle during ramp exercise and a decreasing BBB score. The neurons positively stained in immuno-histochemistry assay were brown in appearance as a result of neural tracer BDA. The spinal cord injury group, but not control group, was found to have considerably decreased number of BDA attached neurons. Additionally, stronger pull was related to a greater loss of positively stained neurons. Conclusion Increased severity of ventral spinal cord injury is associated with more significant dysfunction in cerebral descending neuronal pathways.