托烷司琼通过激活大鼠脊髓α7nAChR减轻慢性神经病理性疼痛和p38MAPK表达

Tropisetron reduces chronic neuropathic pain and the expression of p38MAPK by activating α7nAChR in rat spinal cord

摘要:

目的： 探索鞘内注射托烷司琼是否通过激活α7烟碱乙酰胆碱受体（alpha 7 nicotinic acetylcholine receptor，α7nAChR）减轻坐骨神经分支选择性损伤（spared nerve injury，SNI）大鼠慢性神经病理性疼痛和p38丝裂原活化蛋白激酶（p38 mitogen-activated protein kinase，p38MAPK）表达。方法： 体质量180~220 g的成年雄性SD大鼠，鞘内置管和模型成功后，采用随机数字表法分为5组（n=12只/每组）：假手术组、神经损伤组、托烷司琼组、α7nAChR拮抗剂methyllycaconitine citrate组（MLA组）、MLA+托烷司琼组。假手术组仅暴露右侧坐骨神经，神经损伤组、托烷司琼组、MLA组、MLA+托烷司琼组均制备右侧坐骨神经损伤模型。模型建立后第14天分别鞘内给药，于不同时间点进行疼痛行为学观察和测试。给药1 h后处死大鼠取脊髓L4~6段，采用组织免疫荧光染色观察脊髓背角α7nAChR分布和表达情况，Western blot检测α7nAChR、p-p38和p38蛋白表达量变化。结果： 与给药前相比，托烷司琼组和MLA+托烷司琼组给药后机械缩爪阈值（paw mechanical withdrawal threshold，PMWT）和辐射热缩爪潜伏期（paw thermal withdrawal latency，PTWL）都明显升高，MLA+托烷司琼组均较托烷司琼组低。给药前后MLA组PMWT和PTWL均无明显变化；α7nAChR荧光结果显示，与假手术组阳性面积相比，神经损伤组明显降低。托烷司琼组和MLA+托烷司琼组阳性染色都明显较神经损伤组升高；Western blot结果发现托烷司琼组和MLA+托烷司琼组α7nAChR蛋白表达较神经损伤组明显增高，而p-p38蛋白表达水平与神经损伤组相比均明显降低。各组p38蛋白表达水平均无明显差异。结论： 鞘内注射托烷司琼可减轻SNI大鼠慢性神经病理性疼痛，α7nAChR拮抗剂MLA可一定程度阻断其疼痛减轻作用。其机制可能与托烷司琼选择性激活α7nAChR从而抑制p38MAPK信号通路有关。

Abstract:

Objective: To explore whether intrathecal injection of tropisetron alleviates chronic neuropathic pain and p38 mitogen-activated protein kinase (p38MAPK) expression in rats with spared nerve injury (SNI) by activating alpha 7 nicotinic acetylcholine receptor (α7nAChR) . Methods: Adult male SD rats weighing 180-220 g were randomly divided into 5 groups (n=12/each group): sham operation group, nerve injury group, tropisetron group, α7nAChR antagonist methyllycaconitine citrate group (MLA group), and MLA+tropisetron group. The right sciatic nerve was exposed only in the sham operation group, and the right sciatic nerve injury models were made in the nerve injury group, tropisetron group, MLA group and MLA+tropisetron group. On the 14th day after the establishment of the pain model, the drugs were administered intrathecally, and the pain behavior was observed and tested at different time. One hour after administration, the rats were sacrificed and the L4-6 segments of spinal cord were removed. The distribution and expression of α 7nAChR in spinal dorsal horn were observed by tissue immunofluorescence staining. Western blot was used to detect the changes of α7nAChR, p-p38 and p38 protein expression. Results: The paw mechanical withdrawal threshold (PMWT) and paw thermal withdrawal latency (PTWL) in tropisetron group and MLA+tropisetron group were significantly higher than those before administration, and that in MLA+tropisetron group was lower than that in tropisetron group. There was no significant change in PMWT and PTWL in MLA group before and after administration. The immunofluorescence results of α7nAChR showed that the positive area in the nerve injury group was significantly lower than that in the sham operation group. The positive staining in tropisetron group and MLA+tropisetron group was significantly higher than that in nerve injury group. Western blot results showed that the expression of α 7nAChR in tropisetron group and MLA+tropisetron group was significantly higher than that in nerve injury group, while the expression level of p-p38 protein in tropisetron group was significantly lower than that in nerve injury group. There was no significant difference in the expression level of p38 protein among all groups. Conclusion: Intrathecal injection of tropisetron can relieve chronic neuropathic pain in SNI rats, and α7nAChR antagonist MLA can block its pain-relieving effect to some extent. The mechanism may be related to the selective activation of α7nAChR and inhibition of p38MAPK signal pathway by tropisetron.