摘要目的:寻找一种新型、直观地观察大鼠背根神经节（DRG）感觉神经纤维走行的顺行示踪方法。方法:2022年7月-2023年8月，中山大学附属第一医院整形外科构建带神经元特异性启动子及红色荧光蛋白的腺相关病毒（AAV- syp-mCherry）,同时随机选择3只健康1月龄雌性SD大鼠,体质量180~220 g,通过无菌手术将病毒注射入大鼠第4腰髓（L 4）~第6腰髓（L 6）背根神经节远端脊神经中。饲养1个月后,取注射侧L 4~L 6脊髓组织、DRG、坐骨神经及足底皮肤进行神经丝蛋白（NF）和微管关联蛋白2（MAP2）免疫荧光染色,用共聚焦显微镜观察平均光密度（AOD）从而分析腺相关病毒（AAV）阳性神经纤维的分布特征。 结果:注射病毒示踪剂1个月后,在注射侧L 4~L 6脊髓后角（AOD为0.5419±0.0245）、DRG（AOD为0.4917±0.0087）、坐骨神经（AOD为0.6237±0.0634）、足底皮肤的真皮层及皮肤附属器（AOD为0.4943±0.0165）中可见红色标记物,红色荧光聚集区与NF阳性神经纤维或MAP2阳性神经元共标记,提示AAV- syp-mCherry能通过感觉神经内轴突的顺行运输到远隔靶器官（皮肤）处,并长时间显影。 结论:顺行示踪法能够很好地显示初级感觉神经元中枢突和外周突的完整形貌,可作为一种观察周围感觉神经纤维路径的有效策略。

abstractsObjective:To discover a novel and intuitive anterograde tracing method for observing the course of sensory nerve fibers in dorsal root ganglia (DRG).Methods:From July 2022 to August 2023, studies were conducted in the Department of Plastic Surgery of the First Affiliated Hospital of Sun Yat-sen University. Three healthy 1-month old female SD rats, weighed approximately 180-220 g, were randomly selected. An adeno-associated virus carrying a neuron-specific promoter and red fluorescent protein (AAV- syp-mCherry) was prepared. The virus was injected into the spinal roots outside L 4-L 6 DRG through sterile surgery. After 1 month of breeding, L 4-L 6 spinal cord tissue of immunofluorescent staining of the side injected with viral tracer, DRG, sciatic nerve, and plantar skin of the SD rats were collected for immunofluorescent staining of neurofilament protein (NF) and microtubule associated protein 2 (MAP2). Distribution characteristics of AAV-positive nerve fibers were observed using a confocal microscope by average optical density (AOD) . Results:One month after injection of the viral tracer, red markers were visible in the dorsal horn of the L 4-L 6 spinal cord (AOD=0.5419±0.0245), DRG (AOD=0.4917±0.0087), sciatic nerve (AOD=0.6237±0.0634), dermis of plantar skin and skin appendages on the injected side (AOD=0.4943±0.0165). The red fluorescent clusters were found co-labeled with NF-positive nerve fibers or MAP2-positive neurons, it suggested that AAV- syp-mCherry could be transported anterogradely within sensory axons to distant target organs (skin) and offered a long-term image. Conclusion:The anterograde tracing method by adeno-associated virus can effectively show the complete trace of the central and peripheral processes of primary sensory neurons, and can serve as an effective methodology in observation of the pathways of peripheral sensory nerve fibers.