随着竞技体育的快速发展和比赛的日趋激烈，运动员在训练和比赛过程中面临的运动性疲劳问题越来越突出，并已成为制约竞赛成绩进一步提高的重要因素，为此，深入研究运动疲劳产生的相关机制，探索延缓运动疲劳产生或加速运动疲劳消除的有效方法或手段已经成为各国体育工作者普遍关心和亟待解决的重要理论和现实问题。目前，大量人体和动物实验均已证实，基底神经节相关核团电活动异常是导致运动疲劳的重要因素之一，其中，纹状体作为基底神经节主要的信息输入核团在运动疲劳调控中的作用尤为突出。本实验室前期利用电生理学、免疫组织化学、神经化学等多种研究技术初步证实，纹状体神经元异常兴奋是导致运动疲劳的重要因素，并推测其机制与皮层-纹状体Glu通路的过度传导有关。正常状态下，皮层-纹状体Glu通路传递受内源性大麻素系统（endocannabinoid system, eCBs）的调控，纹状体eCBs功能降低会导致皮层-纹状体Glu通路传递效能异常增加，并最终导致运动功能障碍。由此推测，运动疲劳状态下纹状体神经元异常兴奋可能与eCBs的调节功能降低之后皮层-纹状体Glu通路的传递效能异常增强有关。因此，本研究试图通过探讨运动疲劳状态下大鼠纹状体eCBs的功能变化，及其对皮层-纹状体Glu通路传递效能的调节作用，进一步揭示运动疲劳的中枢机制。这对于探索有效的抗运动性疲劳的方法和手段具有重要意义。 研究目的： 以纹状体eCBs为切入点，通过观察运动疲劳后大鼠纹状体投射神经元和中间神经元的兴奋性变化、皮层-纹状体Glu通路传递效能、eCBs中关键性调节蛋白RGS4和CB1受体蛋白表达，以及药物干预CB1受体对大鼠皮层-纹状体Glu通路传递效能和运动能力的影响，探讨eCBs介导皮层-纹状体Glu通路传递效能参与运动疲劳调控，以期从神经通路的角度阐释运动疲劳的可能机制。 研究方法： 选用成年雄性Wistar大鼠为研究对象，适应性饲养3天后随机分为安静对照组（Control）、1天运动疲劳组（1FG）、3天运动疲劳组（3FG）和7天运动疲劳组（7FG）。Control组在安静状态下，运动组根据实验室前期建立的大鼠运动疲劳模型完成建模后，采用玻璃微电极胞外记录的电生理学技术，分析运动疲劳后大鼠纹状体中等棘状神经元（medium spiny neurons, MSNs）、快放电中间神经元（Fast-spiking, FS）和大胆碱能中间神经元（Large aspiny-like neurons, LANs）的兴奋性变化，进一步证实实验室前期得到的纹状体神经元电活动改变参与运动疲劳调控的结论；采用刺激诱发电生理学技术，观察电刺激大鼠初级运动皮层（primary motor cortex, M1）对背外侧纹状体中等棘状神经元（MSNs）兴奋性的影响，评定运动疲劳后大鼠皮层-纹状体Glu通路传递效能的变化；采用免疫组化和免疫印迹技术，观察运动疲劳后大鼠纹状体eCBs中的主要受体蛋白CB1和关键性调节蛋白RGS4的表达，评价运动疲劳后大鼠纹状体eCBs的功能状态；采用侧脑室微量注射+刺激诱发电生理学技术，观察侧脑室微量注射CB1受体激动剂WIN 55,212-2（2mM, 5μL）对纹状体MSNs诱发放电活动的影响，证实 eCBs参与运动疲劳大鼠皮层-纹状体Glu通路传递效能变化的调节过程；采用侧脑室微量注射的方法，观察侧脑室微量注射CB1受体激动剂WIN 55,212-2对大鼠运动能力的影响，探讨CB1受体激动或类似物在抗运动性疲劳中的作用以及实际应用价值。 研究结果： （1）玻璃微电极胞外记录的电生理实验结果显示：1FG组大鼠纹状体MSNs、LANs和FS的兴奋性与Control组相比没有显著性变化（P＞0.05）；3FG和7FG组大鼠纹状体MSNs和FS的放电频率与Control组相比极显著性增加（P＜0.01），爆发式放电的神经元数量、爆发式放电中动作电位的个数、爆发式放电的持续时间和爆发率与Control组相比显著或极显著性增加（P＜0.05或P＜0.01），爆发指数极显著性降低（P＜0.01）；1FG和3FG组大鼠纹状体LANs的放电频率和爆发式放电的相关指标与Control组相比都没有显著性变化（P＞0.05）；7FG组大鼠纹状体LANs的放电频率与Control组相比极显著性增加（P＜0.01），但是爆发式放电的相关指标都没有显著性变化（P＞0.05）。 （2）刺激诱发电生理实验结果显示：在大鼠运动皮层（M1）给予200μA的电刺激后Control组、1FG、3FG和7FG组大鼠纹状体MSNs的放电频率与刺激前相比都出现显著或极显著性增加（P＜0.05或P＜0.01），3FG和7FG组大鼠纹状体MSNs放电频率增加的幅度极显著高于Control组（P＜0.01），诱发放电的潜伏期显著和极显著低于Control组（P＜0.05和P＜0.01），1FG组MSNs诱发放电频率增加的幅度和潜伏期与Control组相比没有显著性变化（P＞0.05）；当各组大鼠纹状体MSNs的诱发放电频率增加的幅度相同时，3FG和7FG组大鼠运动皮层所需要的刺激强度与Control组相比显著和极显著降低（P＜0.05和P＜0.01），1FG组大鼠皮层刺激强度与Control组相比没有显著性变化（P＞0.05）；在大鼠运动皮层（M1）给予200μA的电刺激后，1FG、3FG和7FG组大鼠纹状体MSNs的爆发式放电持续时间与Control组相比显著或极显著延长（P＜0.05或P＜0.01），爆发式放电中动作电位的个数极显著性增加（P＜0.01），爆发式放电中的峰峰间隔在3FG和7FG组中极显著缩短（P＜0.01），1FG组的峰峰间隔与Control组相比没有显著性变化（P＞0.05），7FG组MSNs的爆发指数与Control组相比极显著增加（P＜0.01），1FG和3FG的爆发指数与Control组相比没有显著性变化（P＞0.05）；200μA的电流刺激可以诱发Control组和1FG组大鼠纹状体MSNs的放电频率出现显著和极显著性增加（P＜0.05和 P＜0.01），但是在3FG和7FG组中，仅需160μA的电流刺激即可诱发大鼠纹状体MSNs的放电频率出现显著和极显著性增加（P＜0.05和 P＜0.01）。 （3）免疫组化和免疫印迹实验结果显示：1FG组大鼠纹状体CB1受体和RGS4蛋白表达与Control组相比都没有显著性变化（P＞0.05）；3FG组和7FG组大鼠纹状体CB1受体和RGS4表达与Control组相比都出现极显著性增加（P＜0.01）。 （4）侧脑室微量注射+刺激诱发电生理实验结果显示：侧脑室微量注射CB1受体激动剂WIN 55,212-2（2mM, 5μl）后1h，Control组大鼠纹状体MSNs的诱发放电频率与注射前相比没有显著性变化（P＞0.05）；1FG组大鼠纹状体MSNs的诱发放电频率在注射后10～20min时出现降低趋势，但未见显著性差异（P＞0.05）；3FG和7FG组大鼠纹状体MSNs的诱发放电频率在注射后10～20min时极显著性降低（P＜0.01），至注射后50～60min时基本恢复至注射前水平。 （5）运动疲劳实验结果显示：侧脑室微量注射WIN 55,212-2后7FG组大鼠运动至力竭的时间较aCSF注射组显著延长（P＜0.05），运动距离显著增加（P＜0.05）；1FG组和3FG组大鼠运动至力竭的时间和运动距离与aCSF注射组相比有延长趋势，但未见显著性变化（P＞0.05）。 研究结论： （1）运动疲劳可显著增加大鼠纹状体投射神经元和中间神经元的兴奋性，尤其是以MSNs和FS的兴奋性增强最为明显，研究结果进一步证实纹状体投射神经元和中间神经元共同参与了运动疲劳调控，并且具有明显的累积效应。 （2）重复性运动疲劳可以显著增强大鼠皮层-纹状体Glu通路的传递效能，但是一次性运动疲劳对其影响不明显，研究结果表明皮层-纹状体Glu通路传递效能异常增强参与了运动疲劳调控，并且还是引起重复性运动疲劳后大鼠纹状体神经元兴奋性增加的主要因素。 （3）重复性运动疲劳大鼠皮层-纹状体Glu通路传递效能异常增强与纹状体eCBs中主要受体蛋白CB1和关键性调节蛋白RGS4过表达后eCBs的调节功能降低有关。提高运动疲劳大鼠纹状体eCBs功能可以抑制皮层-纹状体Glu通路的传导效能，并在一定程度上起到延缓运动疲劳的作用，证实了eCBs介导大鼠皮层-纹状体Glu通路传导效能异常增强参与了运动性中枢疲劳调控，同时纹状体eCBs有望成为运动中枢疲劳调控作用的新靶点。

With the rapid development of competitive sports and the increasingly fierce competition, athletes' sports fatigue problems are becoming more and more prominent in the course of training and competition, and gradually become an important factor restricting the further improvement of competition performance,so, exploring effective methods or means to delay or accelerate fatigue elimination has become an important theoretical and practical problem that sports workers all over the world were generally concerned about and needed to be solved urgently. At present, a large number of human and animal experiments have confirmed that the change of electrical activity in basal ganglion associated nuclei was one of the important factors leading to exercise-induced fatigue.Striatum, as the main information input nucleus in basal ganglion, played an important role in the regulation of exercise-induced fatigue.Using electrophysiology, immunohistochemistry, neurochemistry and other research techniques in the early stage,our laboratory has fully confirmed that abnormal excitation of neurons in the striatum was an important factor leading to exercise-induced fatigue,and speculated that its mechanism was related to the excessive transmission of corticostriatal glutamatergic pathway.Under normal conditions, corticostriatal glutamatergic pathway is regulated by endocannabinoid system (eCBs),and the decreased eCBs function in striatum will lead to abnormal transmission efficiency of corticostriatal glutamatergic pathway, and ultimately resulte in motor dysfunction.Thus conjecture, abnormal activation of striatal neurons under exercise-induced fatigue may be related to the abnormal transmission efficiency of corticostriatal glutamatergic pathway after the decrease of eCBs regulatory function.Therefore, to investigate the functional changes of eCBs in striatum of rats under exercise-induced fatigue,and its regulatory effect on the transmission efficiency of corticostriatal Glupathway in rat is of great significance to further reveal the central mechanism of exercise-induced fatigue and to explore effective methods and means of anti-exercise fatigue. Objectives: eCBs in striatum as the breakthrough point in this paper.The excitability changes of striatal projection neurons and interneuron after exercise fatigue in rats, the transmission efficiency of the cortex-striatal Glu pathway, the expression of key regulatory protein RGS4 and CB1 receptor in eCBs,and the effect of drug intervention on CB1 receptor on the transmission efficiency of corticostriatal Glu pathway andexercise ability in rat were observed.To explore the transmission efficiency of cortex-striatal Glu pathway mediated by eCBs through changing the excitability of striatal neurons involved in the regulation of motor fatigue, so as to explain the possible mechanism of motor fatigue from the perspective of neural pathways. Methods: Adult male Wistar rats were randomly divided into control group (Control) and 1-day fatigue group (1 FG), 3-day fatigue group (3FG) and 7-day fatigue group (7FG) after 3 days rearing. Control group in quiet state and the exercise group completed the model according to the model of exercise-induced fatigue established in the early stage of the laboratory. The Medium spiny neurons (MSNs), Fast-spiking (FSs) and Large aspiny-like neurons (LANs)in the striatum of rats after exercise-induced fatigue were analyzed by electrophysiological technique of extracellular recording with glass microelectrode in order to further confirm the conclusion that the changes of striatal neuronal electrical activity were involved in the regulation of sports fatigue. The stimulation-induced electrophysiological technique was used to observe the effects of electrical stimulation of primary motor cortex(M1)on the excitability of MSNs in dorsolateral striatum in order to evaluate the changes in the transmission efficiency of corticostriatal Glu pathway after exercise-induced fatigue. Immunohistochemistry and immunohistochemistry were used to observe the expression of main receptor protein CB1 and key regulatory protein RGS4 in eCBs of rat’s striatum in order to assess the functional status of eCBs in rats with exercise-induced fatigue. Microinjection of lateral ventricle plus the stimulation-induced electrophysiological technique was used to observe the effect of microinjection of CB1 receptor agonist WIN 55,212-2 (2mM,5μL) on the evoked discharges in MSNs of striatum in order to confirm that eCBs was involved in the regulation of changes in the transmission efficiency ofcorticostriatal Glu pathway Glu pathway in rats with exercise-induced fatigue.Using the method of microinjection of the lateral ventricle, the effect of microinjection of CB1 receptor agonist WIN 55,212-2 (2mM,5μL) on the motor ability of rats was observed in order to dicuss the role of eCBs in anti-fatigue and its practical application value. Results: (1)The results of electrophysiological experiments recorded by glass microelectrode showed that the excitability of MSNs,LANs and FS in the striatum of rats in 1FG was not significantly different from that of Control group(P＞0.05).The firing rate of MSNs and FS in striatum of rats in 3FG and 7FG was significantly higher than that of Control group(P＜0.01), The number of “bursty”neurons, the number of action potential in burst, the burst duration, and the burst rate in striatum of rats in 3FG and 7FG were significantly or significantly higher than those in Control group(P＜0.05 or P＜0.01) , the burst index was significantly reduced(P＜0.01). However, in 1FG and 3FG, there was no significant change in firing rate and burst parameters of LANs(P＞0.05).The firing rat of LANs in 7FG was significantly higher than that of Control group, but there was no significant change in the burst parameters. (2)The results of stimulation-induced electrophysiological experiment showed that the firing rate of MSNs in striatum of rats in Control group,1FG,3FG and 7FG was significantly increased compared with that before stimulation when the rats were treated with 200 μ A electrical stimulation in the motor cortex (M1)( P＜0.05 or P＜0.01). But, the amplitude of increase in firing rate of MSNs in 3FG and 7FG was significantly higher than that in Control group(P＜0.01)and the latency was significantly lower than that of Control group(P＜0.05 and P＜0.01). There was no significant change in the latency and the amplitude of increase in firing rate of MSNs evoked discharge in 1FG compared with Control group(P＞0.05). In addition, when the frequency of evoked discharge of striatum MSNs increased to the same amplitude in each group of rats, the intensity of stimulation required in the motor cortex in 3FG and 7FG was significantly and extremely significantly lower than that in the Control group(P＜0.05 and P＜0.01),however, there was no significant change in cortical stimulation intensity between 1FG group and Control group. After 200 μA electrical stimulation in the motor cortex of rats, compared to the Control group, the burst duration of MSNs in Control group,1FG ,3FG and 7FG was significantly or extremely prolonged(P＜0.05 or P＜0.01),and a significant increase in the number of action potentials in burst(P＜0.01). Interspike interval in burst in 3FG and 7FG was significantly shortened than that in the control group(P＜0.01).There was no significant change in interspike interval in burst between 1FG and Control group(P＞0.05). The burst index of MSNs in 7FG was significantly higher than that in Control group(P＜0.01). The burst indices of 1FG and 3FG had no significant change compared with Control group(P＞0.05). In the Control group, 200μA current stimulation was required to induce a significant increase in the frequency of MSNs discharge in the striatum of rats(P＜0.05). In the 1FG, 200μA current stimulation could induce a significant increase in the frequency of MSNs discharge in the striatum of rats(P＜0.01) .However, in the 3FG, only 160μ A current stimulation could induce a significant increase in the frequency of MSNs discharge in the striatum of rats(P＜0.05).In the 7FG, the frequency of MSNs discharge in the striatum was significantly increased by current stimulation of 160μA(P＜0.01). (3)The results of immunohistochemistry and Western blotting showed that there was no significant change in the expression of CB1 receptor and RGS4 protein in striatum of rats in 1FG compared with Control group(P＞0.05). But the expression of CB1 receptor and RGS4 in striatum was significantly increased in both FG and 7FG compared with Control group(P＜0.01). (4)The results of stimulation-induced electrophysiological experiment plus microinjection of lateral ventricle showed thatthere was no significant change in the frequency of MSNs evoked discharge activity in the control group at 1h after injection of WIN55N212-2(P＞0.05), that the induced discharge frequency of MSNs in the striatum of rats in group 1FG was decreased at 10～20 min after injection of WIN55N212-2, but no significant difference was found(P＞0.05), andthat the evoked discharge frequency of MSNs in 3FG and 7FG was significantly lower than that before injection at 10～20 min after injection of WIN55N212-2(P＜0.01),and it was basically restored to the pre-injection level by the time of 50~60min injection. (5) The experimental results of exercise fatigue showed that the time of exercise to fatigue in 7FG group after microinjection of WIN 55-212-2 in lateral ventricle was significantly longer than that in aCSF group, and the distance of movement was also significantly increased(P＜0.05).However, the time and distance from exercise to fatigue in 1FG and 3FG were not significantly different from those in aCSF injection group（P＞0.05）. Conclusions: (1)Repetitive exercise fatigue significantly increased the excitability of the projection neuron and interneurons in the striatum, especially the excitability of MSNs and FS.The results further demonstrated that both striatalprojection neurons and interneurons were involved in motor fatigue regulation and showed a significant cumulative effect. (2)Repetitive exercise fatigue could significantly enhance the transmission efficiency of the corticostriatal glutamatergic pathway in rats, but the effect of one-time exercise fatigue on its excitability was not obvious.This result hinted that the increased excitability of striatum neurons after repetitive exercise fatigue was related to the abnormal transmission efficiency of the cortex-striatal Glu pathway,and showed that the abnormal enhancement of transmission efficiency of corticostriatal glutamatergic pathway was involved in the regulation of exercise fatigue.. (3) The abnormal enhancement of the transmission efficiency of corticostriatal glutamatergic pathway in rats after repetitive exercise fatigue was associated with the decrease of the regulatory function of eCBs after overexpression of major receptor protein CB1 and key regulatory protein RGS4 in striatum eCBs.Increasing the eCBs function of striatum could inhibit the excessive activation of corticostriatal glutamatergic pathway and delay exercise fatigue to some extent.The results showed that the abnormal enhancement of transmission efficiency of corticostriatal glutamatergic pathway mediated by eCBs may participate in the regulation of exercise induced central fatigue, and eCBs may be a new target of anti-exercise central fatigue.