乌头汤调控海马BDNF/Trk B通路以缓解神经病理性疼痛的痛共情绪症状

Wutoutang Modulation Hippocampal BDNF/TrkB Pathway to Relieve Pain-emotion Co-curation

目的:探索乌头汤对海马组织内脑源性神经营养因子/原肌球蛋白受体激酶B(BDNF/Trk B)通路的调控作用,以明确乌头汤抑制神经病理性疼痛(NP)达到痛与情绪共治的初步机制。方法:将小鼠随机分为假手术组(sham),脊神经结扎组(SNL),乌头汤组(乌头汤),乌头汤-ANA12拮抗剂组(乌头汤-ANA12),普瑞巴林(PGB)组和盐酸氟西汀(FLU)组6组。所有小鼠针对海马安装给药套管,除假手术组暴露但不结扎神经外,其余组均进行脊神经结扎。术后1~21 d,各给药组分别以126 mg·kg^-1乌头汤水煎液,25 mg·kg^-1PGB,3 mg·kg^-1FLU灌胃给药,sham组及SNL组以等体积生理盐水灌胃,1次/d。术后10~16 d进行海马套管给药,其中乌头汤-ANA12组给予Trk B受体拮抗剂ANA12(50 nmol·L-1),其余组给予等体积生理盐水。采用Von Frey法检测小鼠的机械痛阈值,采用旷场和悬尾实验分别检测小鼠的焦虑和抑郁症状,采用高尔基染色法检测小鼠海马CA3区椎体神经元的病理性改变。结果:与假手术组比较,SNL模型组小鼠机械痛阈值显著降低,旷场中心区停留时间显著降低,悬尾不动时间显著增加(P<0.01),海马神经元树突分支数明显减少(P<0.05,P<0.01);与SNL模型组比较,术后9~21 d乌头汤组小鼠的机械痛阈值显著增高,旷场中央区停留时间显著增高,悬尾不动时间显著降低,海马CA3区椎体神经元顶部和底部树突分支数显著增高(P<0.01);与SNL模型组比较,乌头汤-ANA12组的机械痛阈值在术后9 d显著增高(P<0.01),但在通过套管进行ANA12干预后,乌头汤-ANA12组的机械痛阈值持续降低,在术后14~21 d与SNL组比较无统计学差异。乌头汤-ANA12组小鼠中心场停留时间在术后9 d较SNL模型组显著增高(P<0.01),但在小鼠海马进行ANA12干预后,乌头汤-ANA12组的中心场停留时间持续降低,在术后21 d降至与SNL组无统计学差异;乌头汤-ANA12组小鼠悬尾不动时间在术后21 d与SNL组比较无统计学差异;海马高尔基染色显示,乌头汤-ANA12组海马CA3区椎体神经元顶部和底部树突分支数与SNL组比较无统计学差异。结论:乌头汤能缓解SNL导致的海马CA3区椎体神经元损伤及痛共情绪障碍症状,相关作用可能与其对海马BDNF/Trk B通路的调控有关。

Objective:To explore the mediating effect of Wutoutang(WTT)on brain-derived neurotrophic factor/tropomyosin receptor kinase B(BDNF/Trk B)pathway in hippocampus and to clarify the mechanism of therapeutic action of WTD on pain-emotion comorbidity by inhibiting neuropathic pain(NP)preliminarily.Method:The mice were divided into sham group,spinal cord ligation(SNL)group,Wutoutang(WTT)group,Wutoutang-ANA12 antagonist(WTT-ANA12)group,pregabalin(PGB)group,Fluoxetine Hydrochloride(FLU)group randomly.Mice were fixed with the drug delivery cannula for hippocampal CA3.The L5 spinal cord of mice were tightly ligated but sham group(only exposed).During the 10-16 thday after surgery,WTT,WTT-ANA12 groups were gavaged with 126 mg·kg^-1 WTT,PGB and FLU groups were respectively given25 mg·kg^-1 PGB and 3 mg·kg^-1 FLU,sham and SNL groups were given the physiological saline once a day.Then,50 nmol·L-1 ANA12 were given to the hippoicampal CA3 of the WTT-ANA12 mice by drug delivery cannula,and physiological saline were given to the others on the 10-16 thday after surgery.Mechanical pain were detected by Von Frey tests,anxiety and depression behaviors were separately detected by the open field and the tail tailing experiments,while the morphology of CA3 pyramidal neurons were qualified by the Golgi-staining.Result:Compared with sham group,significant decreases of the mechanical pain thresholds,decreases of the duration time in the open field,as well as the increases of the no-struggling time during the tail-suspension were detected in the SNL mice(P<0.01).In addition,as illustrated by the Golgi-staining,the atrophy of hippocampal pyramidal neurons were found in SNL mice as compared with sham(P<0.05,P<0.01).On the contrary,as compared to the SNL,significant increases of the mechanical pain thresholds,increases of the duration time in the open field,the decreases of the no-struggling time during the tail-suspension(P<0.01),as well as the morphological improvements of the hippocampal CA3 pyramidal neurons were detected in the WTT mice.Furthermore,after 7 d hippocampal injections,There is no significant distinction of the mechanical pain thresholds,the duration time in the open field,the no-struggling time during the tail-suspension,as well as the atrophy of hippocampal neurons were detected in the WTT-ANA12 groups as compared with SNL.Conclusion:The data suggested that the effective inhibition of WTT on SNL-induced vertebral neuron injury in hippocampus CA3 and pain-emotion disorder,which might attribute to it’s regulation of BDNF/Trk B pathway in hippocampus.