基于KCC2-GABAA受体通路探讨恢刺“阳陵泉”穴抗卒中后痉挛效应的机制研究

Needling techniques are essentialfor the effect of acupoints. Applying appropriate needling technique according to the disease and syndrome is the key point to improve the therapeutic effect of acupuncture. Waggle needling is a classical needling technique originated from Neijing and it is specific for treating muscular and sinew spasm. It shows confirmed clinical effect for post-stroke spasticity. However, the biological mechanismsunderlying this action are unclear. K+-Cl- cotransporter 2 (KCC2), which is essential for the maintenance of chloride homeostasis, plays an important role in modulating the strength of γ-aminobutyric acid (GABA)mediated synaptic inhibition and then relieving spasticity. Whether it is involved in acupuncture’s effect of anti-spasticity is unclear. This study plans to apply waggle needling technique to GB34 of rats with post-stroke spasticity to evaluate its effect on the excitability of dynamoneurein vivo. The changes of KCC2, GABAA and GABA in spinal cord, brain stem and cortex will be observedin vitro. The specific role of KCC2 will be investigated by employing KCC2 antagonism and agonist respectively. The purpose of this study is to investigate the possible target spots of waggle needling’s anti-spasticity effect based on KCC2-GABAA receptor pathway, which modulates GABA inhibition effect, to explore the scientific connotation underlying this action, and to provide more reliable experimental evidence for the clinical application of waggle needling.

毫针刺法是影响经穴效应的重要因素。恢刺是《内经》中主治筋肉拘挛的经典刺法。大量临床实践证实恢刺对卒中后痉挛疗效肯定，但其抗痉挛效应的机制尚不清楚。KCC2调控的神经元Cl-稳态机制是GABA发挥抑制性递质作用从而缓解痉挛的前提和关键环节，但其是否参与了针刺抗痉挛机制尚未见报道。本研究采用技术成熟的线栓法制作卒中后痉挛大鼠模型，以恢刺法针刺筋会“阳陵泉”穴，以行为学、肌张力电生理学评价为效应指标活体观察脊髓运动神经元的兴奋性，离体观察脊髓、脑干、皮层中KCC2、GABAA、GABA的变化，从整体及分子水平揭示恢刺抗痉挛的机制；经鞘内注射KCC2抑制剂及激动剂活体观察KCC2对针刺引起的肌张力变化的特异性作用；从KCC2-GABAA受体通路调控GABA抑制效率的角度探寻恢刺法抗痉挛的可能途径，揭示“恢刺恢筋急”的科学内涵，丰富刺法理论，为临床制定卒中后痉挛的最佳刺法方案提供实验依据。

脑卒中后肌痉挛是导致患者残疾、阻碍患者康复的关键因素。恢刺是主治筋肉拘挛的经典刺法,对卒中后痉挛临床疗效肯定。现代医学认为，KCC2调控的神经元Cl-稳态机制是GABA发挥抑制性递质作用从而缓解痉挛的前提和关键环节。本研究通过改进的线栓法闭塞大脑中动脉制作卒中后痉挛模型大鼠，采用恢刺法及常规针法针刺卒中后痉挛大鼠“阳陵泉”穴，运用行为学、形态学、电生理学、分子生物学等多种现代技术观察了恢刺法对卒中后痉挛的效应，发现恢刺法可减轻卒中后痉挛大鼠的神经功能损伤和肌痉挛状态，增强模型大鼠大脑皮层缺血区、脑干及脊髓中GABA、KCC2和GABAAγ2的含量及表达，且其效应可被KCC2抑制剂所消除、被KCC2激动剂所模拟。本研究证实了通过KCC2-GABAA受体通路调控GABA抑制效率是恢刺法改善卒中后痉挛的可能靶点和途径之一，揭示了“恢刺恢筋急”的科学内涵，丰富了刺法理论，为临床制定卒中后痉挛的最佳刺法方案提供了实验依据。