免疫荧光法显示贝居腹盾吸虫肌肉神经组织的实验研究

Experimental Study on the Display of Muscle and Nerve Tissue of Aspidogaster conchicola by Immunofluorescence

从背角无齿蚌(Anodonta woodiana)围心腔内检获自然感染的贝居腹盾吸虫(Aspidogaster conchicola),根据虫体内睾丸、卵巢、子宫及卵黄腺4种生殖器官的发育程度,将所获虫体分为幼虫组和成虫组,所有虫体用4%甲醛固定后,分别采用鬼笔环肽染色法和乙酰化微管蛋白免疫组织化学法显示贝居腹盾吸虫成虫和幼虫的肌肉纤维和神经纤维,使用激光共聚焦显微镜观察。结果显示,鬼笔环肽荧光染色的贝居腹盾吸虫成虫较幼虫肌肉组织更为发达,口周围、盾盘附着器、子宫末段及阴茎囊肌肉纤维分布较为密集。乙酰化微管蛋白荧光染色的贝居腹盾吸虫成虫较幼虫神经网络更为复杂,在部分生殖器官如子宫末段、阴茎囊和睾丸部位存在神经纤维。双重染色结果显示,该吸虫不同器官上的肌肉纤维分布与神经纤维的分布相互吻合。该结果提示,贝居腹盾吸虫可能通过神经系统对其肌肉活动进行调控。

[Objectives]The study aims to understand the regulation of the Aspidogaster conchicola nervous system on muscles.[Methods]The naturally infected Aspidogaster conchicola were collected from the pericardium of Anodonta woodiana.According to the development degree of the testis,ovary,uterus,vitelline glands,and other reproductive organs in the insect body under the microscope,the insect body was divided into larva group and adult group.After being fixed with 4%formalin,the muscle fibers and nerve fibers of the larva and adult were observed by phalloidin staining and acetylated tubulin immunohistochemistry,respectively,and were observed by confocal laser microscopy.[Results]Through phalloidin fluorescent staining,we found that the muscle tissue of adult was more developed than that of larvae,and there are two different types of muscle fibers distributed in the body wall,the outer layer is longitudinal muscle fibers,the inner layer is annular muscle fibers,and the muscle fibers are reticulate(Fig.2a),and the muscle fibers of oral sucker(Fig.2b),uterus and cirrus pouch(Fig.2f),and adhesive disc(Fig.2g)was more densely distributed.Fluorescence staining of acetylated tubulin showed that the neural network of adults was more complex than that of larvae,and some nerve fibers of reproductive organs could be seen(Fig.4b,c).The results of double staining showed that the distribution of muscle fibers and nerve fibers on different organs of the fluke were consistent with each other(Fig.5).[Conclusion]The trematode may regulate its muscle activity through the neural network.