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Application of adeno-associated virus on thalamic striatum pathway in rats
- HE Tianqi, LI Min, WANG Xuenan, WANG Yanan, LI Yuchuan, SUN Shuang, ZHAO Hailong, WANG Hao, CHEN Dadian, ZHU Meijia, WANG Xiaojun, WANG Min, LI Xiuhua
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Journal of Shandong University (Health Sciences). 2020, 58(3):
65-74.
doi:10.6040/j.issn.1671-7554.0.2019.1369
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Objective To investigate the neuronal projection of adeno-associated virus(AAV2/9-GFP)in parafascicular nucleus-striatum pathway and its effect on tissues. Methods A total of 136 adult male Wistar rats were randomly divided into parafascicular nucleus(PF)group (n=64), dorsolateral striatum(DLS)group(n=64), PF control group(n=4)and DLS control group (n=4)to conduct stereotactic localization experiments. Based on stereotactic localization experiments, behavior experiments were performed in PF behavior group(n=12)and behavior control group(n=4). After stereotactic localization experiments, the neuron counting experiments were conducted in PF counting group (n=16), 山 东 大 学 学 报 (医 学 版)58卷3期 -何天齐,等.腺相关病毒在大鼠丘脑纹状体通路中的应用 \=- DLS counting group(n=16), PF counting control group(n=4)and DLS counting control group(n=4). After different doses of AAV2/9-GFP were injected into the PF group and DLS group for different durations, the transfection efficiency was analyzed, and the optimal dose and transfection period were screened. The transfected neurons and neuronal projection pathways between the two nuclei were observed. The behavioral changes of rats and number and morphology of neurons were analyzed. SPSS Statistics 21.0 software was used for statistical analysis. Results With the increase of transfection doses and time, the PF optical density value increased (Fdoses=55.617, P<0.001; Ftime=65.288, P<0.001), and the transfection area increased (Fdoses=202.443, P<0.001; Ftime=224.064, P<0.001), showing an upward trend. With the increase of transfection doses and time, the DLS optical density value increased (Fdoses=117.062, P<0.001; Ftime=155.792, P<0.001), and transfection area increased (Fdoses=240.422, P<0.001; Ftime=304.571, P<0.001), showing an upward trend. The optimal dose of PF for viral transfection was 0.4 μL and the optimal duration was 4 weeks. The optimal dose of DLS transfection was 0.6 μL and the optimal duration was 4 weeks. When AAV2/9-GFP was injected into PF, it was projected to the dorsolateral striatum, motor cortex and insular lobe. When AAV2/9-GFP was injected to DLS, it was projected to the motor cortex, hypothalamic nucleus, parafascicular nucleus and substantia nigra. AAV2/9-GFP had damage to neurons, which was characterized by a decreased number of neurons (P<0.05), irregular morphology, enlarged cell body, and shallower staining of nucleoli and Nissl bodies. Conclusion There is a close neuronal projection relationship between PF, DLS, motor cortex, hypothalamic nucleus and substantia nigra. AAV2/9-GFP is an effective neural tracer and harmful to neurons, so it should be cautiously used in clinical practice.
