进一步改良Sihler's肌肉神经染色法显示SD大鼠皮肤神经

施权; 魏在荣; 杨胜波; 金文虎; 李海; 王达利; 王波

doi:10.13418/j.issn.1001-165x.2018.04.024

中国临床解剖学杂志 ›› 2018, Vol. 36 ›› Issue (4) : 461-464. DOI: 10.13418/j.issn.1001-165x.2018.04.024

技术方法

进一步改良Sihler's肌肉神经染色法显示SD大鼠皮肤神经

施权，　魏在荣，　杨胜波，　金文虎，　李海，　王达利，　王波

作者信息 +

Displaying the nerve distribution of SD rat skin by a further modified Sihler's intramuscular nerve staining method

SHI Quan, WEI Zai-rong, YANG Sheng-bo, JIN Wen-hu, LI Hai, WANG Da-li, WANG Bo

Author information +

文章历史 +

摘要

目的　探讨进一步改良Sihler's肌内神经染色法在显示SD大鼠皮肤神经分布中的应用。方法　选取成年SD大鼠6只处死后紧贴肌肉表面分别于背侧及腹侧切取皮肤，最终获得背侧皮肤6块，腹侧皮肤6块，经10%甲醛溶液固定1月，无水乙醇脱脂3 d，并于0.25%的胶原酶Ⅰ和Ⅲ溶液中消化3 d后，经进一步改良的Sihler's肌内神经染色法染色SD大鼠皮肤。结果　12块SD大鼠皮肤经染色后，除2块皮肤染色效果不佳外，其余10块皮肤均获得了良好的效果。在除色素的过程中，表皮常常脱落，即使不脱落，平铺的皮肤上也不能分界表皮与真皮，无法看见表皮内神经分支，且在解剖过程中剔除皮下脂肪时，部分皮下脂肪难以剔除干净，故肉眼可见部分皮下脂肪内明显的神经分布。透过X线阅片灯肉眼即可见皮肤呈现淡紫色，而神经为黑色，各区域神经分布不均，直径、形态及颜色深浅也不相同。甚至可看见神经呈现扭曲、打结的现象。结论　经进一步改良的Sihler's肌内神经染色法染色SD大鼠皮肤，能够清晰显示肉眼可见的皮肤神经分布，这些结果将对于SD大鼠皮肤内各区域神经纤维密度的测定及动物模型中皮瓣移植后神经再生情况的研究具有一定意义。

Abstract

Objective　To investigate the application of the further modified Sihler's intramuscular nerve staining method in displaying the nerve distribution of SD rat skin.　Methods　After six adult SD rats were selected and sacrificed, the skin was harvested against muscle surface on the dorsal and ventral side. Finally, 6 pieces of the dorsal skin and 6 pieces of the ventral skin were obtained. After fixation in 10% formaldehyde solution for 1 month, the skins were defatted by anhydrous ethanol for 3 days, and digested in 0.25% collagenaseⅠand Ⅲ solution for 3 days. After these steps, the skins were stained using the further modified Sihler's intramuscular nerve staining method.　Results　A total of 12 pieces of SD rat skin were stained, among which 10 pieces obtained a good staining result. In the process of removing the pigment, the epidermis often fell off and even if it did not fall off, a boundary between the epidermis and dermis could not be found on the tiled skin and the nerve branches in the epidermis could not be seen; it was difficult to remove partial subcutaneous fat completely in the anatomical process. Therefore, obvious nerve distribution in partial subcutaneous fat was visible to naked eyes. By virtue of X-Ray film viewer, it could be seen by naked eyes that the skin was light purple in color, and the nerves were black in color and had an uneven distribution with different shape and diameter. It could even be seen that the nerves were twisted and knotted.　Conclusion　Skin nerve distribution visible to naked eyes can be clearly displayed after SD rat skin is stained by the further modified Sihler 's intramuscular nerve staining method. These results have a certain significance for the determination of nerve fiber density in each region of SD rat skin and the study of nerve regeneration after skin flap transplantation in animal models.

导出引用

施权，　魏在荣，　杨胜波，　金文虎，　李海，　王达利，　王波. 进一步改良Sihler's肌肉神经染色法显示SD大鼠皮肤神经[J]. 中国临床解剖学杂志. 2018, 36(4): 461-464 https://doi.org/10.13418/j.issn.1001-165x.2018.04.024

SHI Quan, WEI Zai-rong, YANG Sheng-bo, JIN Wen-hu, LI Hai, WANG Da-li, WANG Bo. Displaying the nerve distribution of SD rat skin by a further modified Sihler's intramuscular nerve staining method[J]. Chinese Journal of Clinical Anatomy. 2018, 36(4): 461-464 https://doi.org/10.13418/j.issn.1001-165x.2018.04.024

参考文献

[1] Sihler C.Ueber muskelspindeln und intramuskuläre nervenendigungen bei schlangen und röschen[J].Arch Mikrosk Anat,1895,46:709-723.
[2] Sihler C. Die Muskelspindeln Kerne und Lage der motorischen Nervenendigungen Zugleich em Nachtrag zuder Arbeit: Ueber Muskelspindeln etc Dieses Archiv Bd. XXXXVI, pg.709. Archiv Mikrosk Anat,1900, 56:334-354.
[3] Won SY,Yang HM,Woo HS,et al. Neuroanastomosis and the innervation territory of the mental nerve[J].Clin Anat,2014, 27(4):598-602.
[4] Zur KB, Mu L, Sanders I. Distribution pattern of the human lingual nerve[J].Clin Anat, 2004,17(2):88-92.
[5] Liu AT, Yu DZ, Chen G, et al. Profiling of innervations of mimetic muscles in fresh human cadavers using a modified Sihler's technique[J].Muscle Nerve, 2010, 42(1):88-94.
[6] Shiozaki K, Miida K,Tanaka R, et al. A method for the staining of intraosseous nerve fibers using Sihler's staining technique[J].Biotech Histochem, 2013, 88(6):290-294.
[7] Won SY, Choi DY, Lee JG, et al. Intramuscular communicating branches in the flexor digitorum profundus: dissection and Sihler's staining[J].Surg Radiol Anat, 2010, 32(3):285-289.
[8] 于大志, 党瑞山, 曹学成, 等. 人小腿骨骼肌肌内神经染色及其亚肌部划分[J]. 解剖学杂志, 2015, 38(5): 579-583.
[9] Van Acker N, Ragé M, Sluydts E, et al. Automated PGP9.5 immunofluorescence staining :a valuable tool in the assessment of small fiber neuropathy [J]? BMC Res Notes, 2016, 9(1):280.
[10]Frahm KS, Mørch CD, Grill WM, et al. Activation of peripheral nerve fibers by electrical stimulation in the sole of the foot[J]. BMC Neurosci, 2013, 14(l):116.
[11]Reynders A, Mantilleri A, Malapert P, et al. Transcriptional profiling of cutaneous mrgprd free nerve endings and CLTMRs[J].Cell Rep, 2015, 10(6):1007-1019.
[12]赵良仲, 刘世宏, 王当憨. 固定皮肤褪色试剂研究[J].解剖学报,1982,13(1):109-112.
[13]Won SY, Kim DH, Yang HM, et al. Clinical and anatomical approach using Sihler's staining technique (whole mount nerve stain)[J].Anat Cell Biol, 2011, 44(1):1-7.
[14]Liem RS, Rokx JT, van Willigen JD. Histochemical detection of horseradish peroxidase activity in the rat by the vibratome-paraplast method[J]. Acta Anat (Basel),1988,132(2):164-168.
[15]Mu L, Sanders I. Sihler's whole mount nerve staining technique: a review[J]. Biotech Histochem, 2010, 85(1):19-42.
[16]Chen G, Jiang H, Liu AT, et al. Neurovascular details about forearm muscles: applications in their clinical use in functional muscular transfer[J]. Surg Radiol Anat, 2010, 32(1):3-8.
[17]Lee SJ, Lim AY, Lim IJ, et al. Innervation of the face studied using modi?cations to Sihler's technique in a primate model[J].Plast Reconstr Surg, 2008,121(4):1188-1205.
[18]Wong MT, Lim AY, Coninck CD, et al. Functional units within the latissimus dorsi muscle based on Sihler technique[J].Ann Plast Surg, 2007, 59(2):152-155.
[19] 刘安堂，江华，赵耀忠, 等. 面神经颊支和下颌缘支的解剖学研究及应用[J].中华整形外科杂志, 2007, 23(5):434-437.
[20]Lim AY, Pereira BP, Kumar VP, et al. Intramuscular innervation of upper-limb skeletal muscles[J].Muscle Nerve, 2004, 29(4):523-530.
[21]Gupta R, Nelson SD, Baker J, et al. The innervation of the triangular fibrocartilage complex: nitric acid maceration rediscovered[J].Plast Reconstr Surg, 2001, 107(1):135-139.