方法文章
This protocol describes the tibial neuroma transposition model, which entails a lesion of the tibial nerve with subsequent transposition of the proximal nerve end toward a subcutaneous pretibial or lateral position. Behavioral testing of neuroma pain and plantar hyperalgesia is quantified using Von Frey monofilaments.
The tibial neuroma transposition (TNT) is a rat model in which allodynia at the neuroma site (tibial nerve) can be independently evaluated from allodynia at the plantar surface of the hind paw innervated by the intact sural nerve. This TNT model is suitable to test therapies for neuroma pain, such as the potential superiority of certain surgical therapies that are already used in the clinic, or to evaluate new drugs and their effect on both pain modalities in the same animal. In this model, a distal lesion (neurotmesis) is made in the tibial nerve, and the proximal nerve end is transposed and fixed subcutaneously and pretibially to enable assessments of the neuroma site with a 15 g Von Frey monofilament. To assess allodynia over the sural nerve, Von Frey monofilaments can be used via the up-down method on the plantar lateral region of the hind paw. After cutting the tibial nerve, mechanical hypersensitivity develops at the neuroma site within 1 week after surgery and persists at least until 12 weeks after surgery. Allodynia at the sural innervated plantar surface develops within 3 weeks after surgery compared to the contralateral limb. At 12 weeks, a neuroma forms on the proximal end of the severed tibial nerve, indicated by dispersion and swirling of axons. For the TNT model surgery, multiple critical (micro)surgical steps need to be followed, and some surgery practice under terminal anesthesia is advised. Compared to other neuropathic pain models, such as the spared nerve injury model, allodynia over the neuroma site can be independently tested from sural nerve hypersensitivity in the TNT model. However, the neuroma site can be tested only in rats, not in mice. The tips and directions provided in this protocol can help research groups working on pain successfully implement the TNT model in their facility.
Every wound, varying from simple lacerations to whole limb amputation, is accompanied by varying degrees of peripheral nerve injury. Such nerve injury can result in the formation of a neuroma, a disorganized entanglement of sprouting nerve fibers. Neuromas become painful in 8%-30% of patients, severely impacting their quality of life1,2,3,4,5. After limb amputation, neuroma pain develops in 50% of patients6,7,8. Re....
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This research was performed in accordance with the IVD (Instantie voor Dierenwelzijn Utrecht) and the guidelines for animal research, project number AVD1150020198824.
1. Von Frey baseline measurements
2. Anesthesia and preparation
NOTE: This study was conducted on 15 male Sprague Dawley rats that were 12 weeks old.
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Assessment at the neuroma site showed increased sensitivity to the application of the 15 g von Frey monofilament. At baseline, rats typically responded to 10%-15% (± 13%) of the 25 applications of a 15 g monofilament. The response rate increased to 45%-50% (± 24%) 1 week after TNT surgery. On the contralateral side, the number of responses after surgery was similar to those at baseline (Figure 2A). Around 20% of the rats did not develop a painful neuroma; the response rate did not increase c.......
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Critical steps in the protocol
The TNT model involves cutting the tibial nerve and transposing it laterally and subcutaneously to a pretibial location to enable sensitivity testing of the neuroma, in addition to plantar hyperalgesia over the sural nerve. In the TNT model, it is key that the place of the neuroma is visible to the researchers. Therefore, an albino rat strain is preferred because subcutaneous sutures are easily visible through the skin and the color of the suture should preferably be da.......
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The authors report to have no conflict of interest. Although this research work was partly financed by Axogen, the company had no influence on the execution of the study and on the results.
We would like to thank Sabine Versteeg for assisting during microsurgery and Anja van der Sar and Trudy Oosterveld-Romijn from the Common Animal Laboratory (Gemeenschappelijk Dieren Laboratorium) for their help in preparing the microscope and surgical room and taking care of the animals.
This research was funded by Axogen.
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| 姓名 | 公司 | 目录编号 | 评论 |
|---|---|---|---|
| Aesthesio | Linton Instrumentation | 514007 till 514015 | 0.6 g 至 15 g 单丝 |
| Carprofen | 当地兽医药房 | n/a | 当地兽医药房制作卡洛芬稀释 |
| 液 棉签 | Nobamed | 974255 | |
| Electrocautery | Fine Science Tools | 18010-00 | |
| 乙醇 70% | Interchema BV | ||
| Ethilon 4.0 | 强生Johnson | 1854G | 重要提示：颜色应为蓝色或黑色 |
| Ethilon 8.0 | Johnson &Johnson | BV130-5 | |
| Isoflo，异氟醚 Zoetis | Dechra 兽医产品 | B506 | |
| 网状底笼 | StoeltingCo | 57816 和 57824 | |
| 微型镊子 | 精细科学工具 | 11251-35 | |
| 微型持针器 | 精细科学工具 | 12076-12 | |
| 微型剪刀 | 精细科学工具 | 15019-10 | |
| 微型镊子 | 精细科学工具 | 11254-20 | |
| NaCl 0.9% | Trademed | H7 1000-FRE | |
| 持针器 | 精细科学工具 | 12004-16 | |
| 眼科软膏 | 当地兽医药房 | n/a | 当地兽医药房制作眼药膏 |
| 手术刀 | 精细科学工具 | 10003-12 | |
| 剪刀 | 精细科学工具 | 14001-12 | |
| 立体手术显微镜 | Leica | A60 F | |
| 带孔无菌片 | Evercare OneMed | 1555-01 | |
| 手术刀片nr.15 | 精细科学工具 | 10015-00 | |
| 镊子 | 精细科学工具 | 11617-12 |
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