不同剂量6-OHDA对小鼠多巴胺能神经元及行为的影响

Effects of different doses of 6-OHDA on dopaminergic neurons and be⁃haviors in mice

目的:探讨多巴胺能神经元数量与动物运动行为之间的关系,为基于6-羟基多巴胺(6-OHDA)注射的、不同病程阶段及不同类型运动障碍的帕金森病(PD)小鼠造模提供参考依据。方法:在C57BL/6雄性小鼠单侧黑质致密部(SNc)注射不同剂量(3 g/L、6 g/L和12 g/L)6-OHDA诱导病变,构建不同损伤程度的PD小鼠,从而模拟人类帕金森病的不同进展阶段(早期、中期和晚期)。在术后第14天,通过转棒实验、爬杆测试、平衡木穿越实验、旷场测试、圆筒实验及步态分析评估小鼠模型的行为缺陷。此外,通过免疫荧光染色定量检测SNc中酪氨酸羟化酶(TH)阳性神经元数量,以及纹状体尾壳核(CPu)中TH+的多巴胺能神经元末梢的密度,以评估小鼠脑组织损伤情况。结果:与对照组相比,注射6-OHDA后,高剂量组和中剂量组的SNc多巴胺能神经元数量显著减少(P<0.05),且损伤呈剂量依赖性(Spearman相关,P<0.01)。同时,高剂量组(P<0.01)和中剂量组(P<0.05)的CPu多巴胺能神经末梢也显著减少。行为学测试显示,高剂量组小鼠的运动协调性和后肢平衡能力严重受损,表现为转棒测试时间缩短、步态异常及圆筒测试中前肢使用不对称等。而中剂量组和低剂量组的小鼠肢体协调性轻度下降,但自主运动能力和步态指标未受明显影响。结论:(1)SNc多巴胺能神经元损伤程度呈剂量依赖效应,即剂量越高,损伤程度越严重;同时,运动障碍与多巴胺能神经元损伤程度并不同步,仅高剂量组才出现明显的运动障碍;(2)基于高剂量6-OHDA的小鼠模型可作为研究PD及运动障碍的有效动物模型;(3)基于低剂量和中剂量6-OHDA的小鼠模型可用于研究PD运动症状出现前期的发病机制。

AIM:To investigate the relationship between the number of dopaminergic neurons and the locomo-tor behavior of animals,and to provide a reference basis for the modeling of mice with different stages of Parkinson disease(PD)and different types of locomotor deficits based on 6-hydroxydopamine(6-OHDA)injection.METHODS:We in-duced lesions in the substantia nigra pars compacta(SNc)by administering various doses of 6-OHDA(3 g/L,6 g/L,and 12 g/L)to create PD mouse models with differing degrees of injury,thereby mimicking the various stages of PD progression observed in patients(early,moderate and advanced stages).On the 14th day post-surgery,we evaluated the behavioral deficits of the mouse models using the rotarod test,pole test,beam traversal test,open field test,and gait analysis.Fur-thermore,the quantification of tyrosine hydroxylase(TH)-positive neurons within the SNc and TH-stained dopaminergic terminals in the corpus striatum caudate-putamen(CPu)was conducted utilizing immunofluorescence staining techniques to assess brain tissue damage.RESULTS:Compared to the control group,the number of dopaminergic neurons in the SNc was significantly reduced in both the high-dose group(P<0.05)and the medium-dose group(P<0.05)following 6-OHDA injection,demonstrating a dose-dependent effect(Spearman correlation,P<0.01).Similarly,the dopaminergic terminals in the CPu were significantly diminished in the high-dose group(P<0.01)and the medium-dose group(P<0.05).Behavioral tests revealed that mice in the high-dose group exhibited severe impairments in motor coordination and hindlimb balance,as evidenced by reduced rotarod test times,gait abnormalities,and asymmetrical forelimb use in the cylinder test.In contrast,mice in the medium-and low-dose groups displayed only mild declines in limb coordination,while their autonomous motor abilities and gait indices remained largely unaffected.CONCLUSION:The results reveal a dose-dependent effect on dopamine neuron damage,with higher doses causing the severest damage.Unexpectedly,signifi-cant locomotion impairments were only manifested in the high-dose group.This suggests that a mouse model induced by higher 6-OHDA dose is effective for studying PD and associated dyskinesia.Conversely,animal models with low to medi-um doses can be useful for exploring the early stages of PD locomotion symptoms.