中下焦俞募穴与相应脏腑特异性联系通路的荧光双标法研究

【作者】 童晨光；

【导师】 谷世喆；

【作者基本信息】 北京中医药大学 ， 针灸学， 2003， 硕士

【摘要】 本课题采用荧光素双标记法研究中下焦俞募穴与相应脏腑的形态学联系通路。研究分四部分。第一部分是文献研究，通过对古今文献的查阅整理发现：（1）经络功能的整体性和多样性决定了其结构的复杂性，经络的功能绝不是某种单一的因素所能实现的，因而也不可能把它的物质基础想得过分简单。（2）许多实验证据都说明神经是经络的重要组成部分之一。（3）经穴---脏腑相关是经络学说的核心内容之一，是指导中医诊断和治疗的重要理论基础，是沟通经络基础和临床的纽带，所以也是当前经络研究的重点内容之一。（4）许多学者的研究表明，经穴---脏腑相关与神经节段有着密切关系。（5）研究表明，来自体表和内脏的信息可在脊神经节及从脊髓到大脑皮层的各级中枢发生汇聚。（6）俞募穴与脏腑有着极为密切的关系，不论古今临床都大量应用俞募穴诊断和治疗疾病，俞募配穴亦是有效的常用配穴法。俞募穴的分布与神经节段有密切关系。（7）目前对气街的研究很少。气街的分布是横贯脏腑经络，前后相接，按横向的形式将脏腑与其在体表的相应部位紧密联系在一起。气街的存在，为俞募穴与脏腑的联系及其临床应用提供了经络学理论依据。（8）通过对古今应用俞募穴情况的对比，发现现代一些俞募穴的使用在逐渐萎缩。由此便产生了从形态学研究俞募穴与相应脏腑神经联系途径的实验构想。这一形态学结果很可能就是临床应用俞募穴治疗脏腑病的理论基础；是气街的实质部位；是穴位功能具有相对特异性的重要依据；还可能解释一些经络现象。第二部分是实验研究，实验对中下焦除三焦俞、石门以外的8对俞募穴及两个非穴点进行研究，分背俞穴---脏腑组、腹募穴---脏腑组和非穴点---胃组。采用荧光素双标记法，取健康成年Wister大鼠，将PI（碘化丙定）按组分别注入背俞穴、腹募穴或非穴点，存活36小时后，分别于相应脏腑包膜下注入Bb（双苯甲亚胺），继续存活12小时，再次麻醉，经心脏和升主动脉灌注固定，根据脏腑位置取相应脊神经节若干，置入含15%蔗糖的磷酸缓冲液中，直至完全下沉。用恒冷箱切片机连续切片，片厚40微米，收集每个脊神经节的所有切片，直接融裱于载玻片上，室温快速吹干，即刻在荧光显微镜下观察，计数荧光素标记细胞，有选择摄片。结果在对肝俞募穴、脾俞募穴、胃俞募穴、大肠俞募穴、小肠俞募穴以及肾俞募穴与相应脏腑联系的研究中，实验都比较成功，分别在相同或邻近神经节段的脊神经节内观察到PI单标记、Bb单标记以及PI-Bb双标记细胞的存在。各组标记细胞均呈圆形或椭圆形，大小不一，多为中小型。PI单标记、Bb单标记细胞和PI-Bb双标记细胞之间的形态大小未见明显差异，但其数量有所不同。至于胆俞募穴与胆和膀胱俞募穴与膀胱联系的研究，则由于不同原因出现一些问题，但这不会影响整体结论的得出。第三部分是分析与讨论，根据实验结果可以得出以下结论：（1）中下焦俞募穴与其相应脏腑之间存在特异性联系通路，此通路就是相同或相近节段的传入神经，并在脊神经节进行整合。（2）这种广泛存在的俞募穴---脊神经节---脏腑的联系途径是俞募穴与<WP=4>其相应脏腑相关的形态学基础之一，也是中医学气街（腹气街）的实质部位之一。（3）脊神经汇聚神经元和周围突分支的存在为穴位特异性调节的外周机制和穴位功能的相对特异性提供了形态学依据。（4）现代一些俞募穴的使用在逐渐萎缩的原因，很大程度上是因为对俞募穴与相应脏腑之间特异性通路的认识程度不够。本课题为临床运用气街理论治疗相关脏腑病提供了可靠的理论和实验依据。第四部分是展望，对今后的研究提出了一些看法。更多还原

【Abstract】 This subject explores into the nervous connection between back-shu and front-mu points and the corresponding internal organs. Fluorescent double-labeling method was used in the study. The research divides into four parts.The first part is literature review. Through statistics and analysis of ancient and modern literature, it was found that (1) The totalite and variety of the function of channels have determined the complexity of its structure, the function of channels cannot realize by a certain single factor, therefore can not think of its material base too simple. (2) A lot of experimental evidences all prove that the nerve is one of important composition of channels. (3) It is one of the key content of the theory of channels that the acupoints is correlated with internal organs, this is the important theoretical foundation of guiding Chinese medicine to diagnose and treat, it is the tie between channels foundation and clinic, so it is also one of important content of channels study at present. (4) A lot of scholars’ researches indicate that the correlation between acupoints and internal organs has close relation with nerve supply. (5) Some researches indicate that the information coming from the body surface and viscera can be assembled at all levels of centers from spinal ganglion to cerebral cortex. (6) Back-shu and front-mu points have extremely close relation with internal organs. No matter ancient time or today doctors always use back-shu and front-mu points to diagnose and treat diseases. It has close relation between the distribution of back-shu and front-mu points and nerve supply. (7) There is little study on "Qijie" at present. The theory of "Qijie" is the foundation of using back-shu and front-mu points. (8)Through the comparison between ancient time and today, it was found that today some back-shu and front-mu points are rarely used at clinic.Therefore the experiment ideas come into my mind, which is to study the nervous connection between back-shu and front-mu points and the corresponding internal organs from morphology point of view. This morphology result is probably the theoretical foundation of using back-shu and front-mu points to treat internal organs diseases, the essence position of "Qijie", the important basis of the relative specificity of the points. It also can explain some channels phenomena.<WP=6>The second part is experiment study. The experiment studies 8 pairs of back-shu and front-mu points and two non-acupoints. The experiment divided into 3 groups: back-shu points --internal organs, front-mu points--internal organs and non-acupoints--stomach. Fluorescent double-labeling method was used in the experiment and use health adult Wister rats to study. PI was separately injected into back-shu points, front-mu points or non-acupoints. The animals survived for 36 hours, and then Bb was pushed into corresponding internal organs. For 12 more hours survival, 4% paraformaldehyde solution in phosphate buffer was perfused through heart, then the DRG were taken, and 40um frozen sections were made for observation. The result is that 6 pairs succeed and other two pairs have some problems. In the same or near Spinal ganglion can observe single-labeling and double-labeling cells. The third part is conclusion. According to the experimental results the following conclusions can be made: (1) The specificity connection path exists between back-shu and front-mu points and corresponding internal organs. These paths are afferent nerves, and assemble at spine ganglion. (2) There exist neurons in the spinal ganglion that control viscera and body surface. This not only provides further explanation to the mechanism of the referred pain, but also indicates that the sensational impulse of acupuncture exert its effect on the functions, activities and sensations of the internal organs through afferent axes at the branches. (3) This study has provided morphological basis for the application of back-shu and front-mu points in the clinical practice. (4) This study also indicates that "Qi更多还原