内侧前额叶皮层和眶额皮层在吗啡成瘾中的作用及机制探讨
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摘要
毒品滥用是威胁人类生存的重大社会问题。吗啡是我国广泛滥用的毒品之一，对其作用机制的研究有助于更好的预防和治疗成瘾性疾病。前额叶皮层作为大脑的高级中枢在药物依赖及成瘾过程中发挥重要作用，但相关机制研究仍然十分有限。本论文从行为学，神经化学，分子生物学等多个水平研究了前额叶皮层的两个亚区—内侧前额叶皮层和眶额皮层在吗啡成瘾中的作用并对其机制进行了初步探讨。
论文首先采用了条件性位置偏爱和行为敏化模型结合脑内兴奋毒性损毁技术，考察了内侧前额叶皮层和眶额皮层在吗啡引起的精神依赖中的作用。结果表明，内侧前额叶皮层损毁破坏了吗啡引起的小鼠条件性位置偏爱的形成，但对吗啡引燃的位置偏爱的重现无明显影响；而眶额皮层损毁不仅破坏了吗啡引起的小鼠条件性位置偏爱的形成，还能阻断吗啡引燃的位置偏爱的重现。
在行为敏化实验中，吗啡(2.5，5，10 mg/kg)剂量依赖性的引起小鼠自主活动的增加和行为敏化的形成。停止给药后小鼠的行为敏化逐渐消退，并在再次给予吗啡后重新表达。内侧前额叶皮层损毁能抑制急性吗啡引起的小鼠自主活动增加，阻断吗啡多次给药形成的小鼠行为敏化，但对行为敏化的表达无影响；而眶额皮层损毁不仅能抑制急性吗啡引起的小鼠自主活动增加，破坏吗啡多次给药形成的小鼠行为敏化，还能阻断吗啡行为敏化的表达。以上结果表明，内侧前额叶皮层和眶额皮层参与了吗啡的急性中枢作用，并在一定程度上影响吗啡的奖赏和复吸，但内侧前额叶皮层和眶额皮层的作用存在一定差异。
论文进一步采用免疫组织化学方法检测了吗啡急性给药，吗啡成瘾及成瘾后纳络酮催促戒断引起的内侧前额叶皮层和眶额皮层c-Fos蛋白的表达。结果显示，吗啡单次给药能够剂量依赖性的引起大鼠内侧前额叶皮层和眶额皮层Fos蛋白表达的增加，这种表达的增加在0.5—1小时最为明显，2小时后开始消退。吗啡成瘾后再次给药同样能够引起前额叶皮层和眶额皮层Fos蛋白表达的增加，应用纳络酮催促戒断后Fos蛋白的表达进一步增强。
大量研究支持药物成瘾与学习记忆过程密切相关，本文考察了内侧前额叶皮层和侧眶额皮层损毁对学习记忆功能的影响以探讨其参与吗啡作用的内在机制。结果发现，内侧前额叶皮层损毁破坏了小鼠的短时工作记忆，而眶额皮层损毁对小鼠的工作记忆功能无明显影响。
谷氨酸作为兴奋性氨基酸类神经递质在脑内广泛存在，除了中脑边缘多巴胺能系统，谷氨酸能系统在药物成瘾中也起到重要作用，此系统主要起源于前额叶皮层。本文采用脑内微透析结合高效液相技术研究了吗啡对内侧前额叶皮层和眶额皮层细胞外谷氨酸含量的影响。结果发现，吗啡单次给药能够显著降低大鼠内侧前额叶皮层细胞外谷氨酸含量，在给药后80—120分钟之内一过性的升高大鼠眶额皮层细胞外的谷氨酸含量；吗啡依赖大鼠内侧前额叶皮层和眶额皮层细胞外谷氨酸的基础含量显著降低，阿片受体阻断剂—纳络酮可翻转这一作用。提示，内侧前额叶皮层和眶额皮层参与吗啡的中枢作用，并可能与此部位的谷氨酸系统相关。
为了寻求和研制有效的中药戒毒新药，本文结合本研究室前期的研究结果，采用行为学及脑内微透析技术，进一步研究了拟人参皂苷F11(PF11)抗吗啡成瘾的作用机制。结果表明，单独应用PF11对小鼠的自主活动无显著影响，但能拮抗单次吗啡引起的小鼠自主活动增加和多次吗啡给药引起的小鼠行为敏化。单独应用PF11不能引起小鼠前额叶皮层细胞外谷氨酸水平的显著变化，但可拮抗吗啡急性给药引起的小鼠前额叶皮层细胞外谷氨酸含量的降低及吗啡依赖小鼠前额叶皮层细胞外谷氨酸基础水平的降低。
综上所述，前额叶皮层的两个主要亚区—内侧前额叶皮层和侧眶额皮层不同程度参与吗啡的急性作用，吗啡成瘾和戒断，这一作用可能与此部位的谷氨酸系统密切相关。拟人参皂苷F11能够拮抗吗啡引起的行为敏化和前额叶皮层细胞外谷氨酸含量的降低，提示其具有抗吗啡成瘾及依赖的作用。
Opiate abuse remains a major social problem in Asian countries. Opiateslike morphine are commonly used clinically to relieve pain and treat chronicdiarrhea. Repeated administration of morphine produces dependence andaddiction. However, the neurobiological mechanisms of morphine actions arestill not completely understood. A better understanding of the neuralmechanisms underlying morphine addiction could lead to more effectivetreatment strategies for addictive disorders. The prefrontal cortex hasimplicated in drug abuse in recent years. To further understand theneuromechanisms underlying morphine addiction, in the present study weinvestigated the pharmacological, biochemical and molecular alterations in thesubregions of the PFC, the medial prefrontal cortex (mPFC) and theorbofrontal cortex (OFC).
Firstly, we employed animal models of addiction coupled with excitotoxicbrain lesion technique to investigate the roles of the mPFC and OFC inmorphine actions. The results showed that both mPFC and OFC lesionsprevented the acquisition of morphine induced conditioned place preference(CPP), but only OFC lesions destroyed drug priming induced reinstatement ofCPP in mice.
In the behavioral sensitization tests, lesions of mPFC and OFC blockedthe hyperactivity induced by acute morphine and the induction of behavioralsensitization by repeated morphine. While only OFC lesions prevented theexpression of behavioral sensitization. These results suggested that the mPFCand OFC may involve in the acute morphine action and mediate, at leastpartially, morphine addiction and stress-induced relapse. This was also provedby the further studies using immunohistochemical methods that both acutemorphine treatment and naloxone-precipitated withdrawal significantly increased c-fos expression in mPFC and OFC.
With the consideration of learning-based addiction theories, we alsoexamined the effects of mPFC and OFC lesions on the process of learning andmemory using spontaneous Y maze in this study. The results showed mPFClesions impaired the working memory, while OFC lesions has little effect on theworking memory, indicating the involvement of mPFC in morphine addictionpartially by its role in memory process.
In addition to the mesocorticolimbic dopaminergic system, glutamate-mediated synaptic transmission also involves in the opiate abuse. The sourceof glutamate is shown to be glutamatergic afferents from the prefrontal cortex.In the present study, we investigated the effect of morphine on the extracellularlevels of glutamate in the mPFC and OFC in freely moving rats by using in vivomicrodialysis coupled with high performance liquid chromatography (HPLC).The results showed that acute morphine decreased the extracellular levels ofglutamate in the mPFC, while increased the extracellular levels of glutamate inthe OFC. Repeated morphine decreased the basal levels of glutamate in bothregions. Naloxone could reverse the decrease induced by repeated morphine,suggesting that the glutamatergic system in the two regions is involved in thecentral actions of morphine.
In the present study, we also investigate the effects of Pseudoginsenoside-F11 (PF11) , an ocotillol-type saponin existing in American ginseng, onmorphine induced behavioral sensitivity and alternations in glutamate levels inthe medial prefrontal cortex on the base of our previous studies. As the resultsshown PF11 antagonized morphine-induced behavioral sensitivity anddecrease of glutamate in the mPFC. Therefore, it is demonstrated that PF11may block morphine-induced behavioral sensitivity via its effect, at leastpartially, on the glutamatergic system in the mPFC and also suggest thatnatural products, such as ginseng, might be potential candidates for theprevention and treatment of the neurological disorders induced by morphineabuse.
In conclusion, the mPFC and OFC are differentially involved in the acutemorphine action and mediate, at least partially, morphine addiction and relapse.The mediation may relate to the glutamatergic system in these regions. Inaddition, PF11 might be a potential candidate for the prevention and treatmentof the neurological disorders induced by morphine.
论文首先采用了条件性位置偏爱和行为敏化模型结合脑内兴奋毒性损毁技术，考察了内侧前额叶皮层和眶额皮层在吗啡引起的精神依赖中的作用。结果表明，内侧前额叶皮层损毁破坏了吗啡引起的小鼠条件性位置偏爱的形成，但对吗啡引燃的位置偏爱的重现无明显影响；而眶额皮层损毁不仅破坏了吗啡引起的小鼠条件性位置偏爱的形成，还能阻断吗啡引燃的位置偏爱的重现。
在行为敏化实验中，吗啡(2.5，5，10 mg/kg)剂量依赖性的引起小鼠自主活动的增加和行为敏化的形成。停止给药后小鼠的行为敏化逐渐消退，并在再次给予吗啡后重新表达。内侧前额叶皮层损毁能抑制急性吗啡引起的小鼠自主活动增加，阻断吗啡多次给药形成的小鼠行为敏化，但对行为敏化的表达无影响；而眶额皮层损毁不仅能抑制急性吗啡引起的小鼠自主活动增加，破坏吗啡多次给药形成的小鼠行为敏化，还能阻断吗啡行为敏化的表达。以上结果表明，内侧前额叶皮层和眶额皮层参与了吗啡的急性中枢作用，并在一定程度上影响吗啡的奖赏和复吸，但内侧前额叶皮层和眶额皮层的作用存在一定差异。
论文进一步采用免疫组织化学方法检测了吗啡急性给药，吗啡成瘾及成瘾后纳络酮催促戒断引起的内侧前额叶皮层和眶额皮层c-Fos蛋白的表达。结果显示，吗啡单次给药能够剂量依赖性的引起大鼠内侧前额叶皮层和眶额皮层Fos蛋白表达的增加，这种表达的增加在0.5—1小时最为明显，2小时后开始消退。吗啡成瘾后再次给药同样能够引起前额叶皮层和眶额皮层Fos蛋白表达的增加，应用纳络酮催促戒断后Fos蛋白的表达进一步增强。
大量研究支持药物成瘾与学习记忆过程密切相关，本文考察了内侧前额叶皮层和侧眶额皮层损毁对学习记忆功能的影响以探讨其参与吗啡作用的内在机制。结果发现，内侧前额叶皮层损毁破坏了小鼠的短时工作记忆，而眶额皮层损毁对小鼠的工作记忆功能无明显影响。
谷氨酸作为兴奋性氨基酸类神经递质在脑内广泛存在，除了中脑边缘多巴胺能系统，谷氨酸能系统在药物成瘾中也起到重要作用，此系统主要起源于前额叶皮层。本文采用脑内微透析结合高效液相技术研究了吗啡对内侧前额叶皮层和眶额皮层细胞外谷氨酸含量的影响。结果发现，吗啡单次给药能够显著降低大鼠内侧前额叶皮层细胞外谷氨酸含量，在给药后80—120分钟之内一过性的升高大鼠眶额皮层细胞外的谷氨酸含量；吗啡依赖大鼠内侧前额叶皮层和眶额皮层细胞外谷氨酸的基础含量显著降低，阿片受体阻断剂—纳络酮可翻转这一作用。提示，内侧前额叶皮层和眶额皮层参与吗啡的中枢作用，并可能与此部位的谷氨酸系统相关。
为了寻求和研制有效的中药戒毒新药，本文结合本研究室前期的研究结果，采用行为学及脑内微透析技术，进一步研究了拟人参皂苷F11(PF11)抗吗啡成瘾的作用机制。结果表明，单独应用PF11对小鼠的自主活动无显著影响，但能拮抗单次吗啡引起的小鼠自主活动增加和多次吗啡给药引起的小鼠行为敏化。单独应用PF11不能引起小鼠前额叶皮层细胞外谷氨酸水平的显著变化，但可拮抗吗啡急性给药引起的小鼠前额叶皮层细胞外谷氨酸含量的降低及吗啡依赖小鼠前额叶皮层细胞外谷氨酸基础水平的降低。
综上所述，前额叶皮层的两个主要亚区—内侧前额叶皮层和侧眶额皮层不同程度参与吗啡的急性作用，吗啡成瘾和戒断，这一作用可能与此部位的谷氨酸系统密切相关。拟人参皂苷F11能够拮抗吗啡引起的行为敏化和前额叶皮层细胞外谷氨酸含量的降低，提示其具有抗吗啡成瘾及依赖的作用。
Opiate abuse remains a major social problem in Asian countries. Opiateslike morphine are commonly used clinically to relieve pain and treat chronicdiarrhea. Repeated administration of morphine produces dependence andaddiction. However, the neurobiological mechanisms of morphine actions arestill not completely understood. A better understanding of the neuralmechanisms underlying morphine addiction could lead to more effectivetreatment strategies for addictive disorders. The prefrontal cortex hasimplicated in drug abuse in recent years. To further understand theneuromechanisms underlying morphine addiction, in the present study weinvestigated the pharmacological, biochemical and molecular alterations in thesubregions of the PFC, the medial prefrontal cortex (mPFC) and theorbofrontal cortex (OFC).
Firstly, we employed animal models of addiction coupled with excitotoxicbrain lesion technique to investigate the roles of the mPFC and OFC inmorphine actions. The results showed that both mPFC and OFC lesionsprevented the acquisition of morphine induced conditioned place preference(CPP), but only OFC lesions destroyed drug priming induced reinstatement ofCPP in mice.
In the behavioral sensitization tests, lesions of mPFC and OFC blockedthe hyperactivity induced by acute morphine and the induction of behavioralsensitization by repeated morphine. While only OFC lesions prevented theexpression of behavioral sensitization. These results suggested that the mPFCand OFC may involve in the acute morphine action and mediate, at leastpartially, morphine addiction and stress-induced relapse. This was also provedby the further studies using immunohistochemical methods that both acutemorphine treatment and naloxone-precipitated withdrawal significantly increased c-fos expression in mPFC and OFC.
With the consideration of learning-based addiction theories, we alsoexamined the effects of mPFC and OFC lesions on the process of learning andmemory using spontaneous Y maze in this study. The results showed mPFClesions impaired the working memory, while OFC lesions has little effect on theworking memory, indicating the involvement of mPFC in morphine addictionpartially by its role in memory process.
In addition to the mesocorticolimbic dopaminergic system, glutamate-mediated synaptic transmission also involves in the opiate abuse. The sourceof glutamate is shown to be glutamatergic afferents from the prefrontal cortex.In the present study, we investigated the effect of morphine on the extracellularlevels of glutamate in the mPFC and OFC in freely moving rats by using in vivomicrodialysis coupled with high performance liquid chromatography (HPLC).The results showed that acute morphine decreased the extracellular levels ofglutamate in the mPFC, while increased the extracellular levels of glutamate inthe OFC. Repeated morphine decreased the basal levels of glutamate in bothregions. Naloxone could reverse the decrease induced by repeated morphine,suggesting that the glutamatergic system in the two regions is involved in thecentral actions of morphine.
In the present study, we also investigate the effects of Pseudoginsenoside-F11 (PF11) , an ocotillol-type saponin existing in American ginseng, onmorphine induced behavioral sensitivity and alternations in glutamate levels inthe medial prefrontal cortex on the base of our previous studies. As the resultsshown PF11 antagonized morphine-induced behavioral sensitivity anddecrease of glutamate in the mPFC. Therefore, it is demonstrated that PF11may block morphine-induced behavioral sensitivity via its effect, at leastpartially, on the glutamatergic system in the mPFC and also suggest thatnatural products, such as ginseng, might be potential candidates for theprevention and treatment of the neurological disorders induced by morphineabuse.
In conclusion, the mPFC and OFC are differentially involved in the acutemorphine action and mediate, at least partially, morphine addiction and relapse.The mediation may relate to the glutamatergic system in these regions. Inaddition, PF11 might be a potential candidate for the prevention and treatmentof the neurological disorders induced by morphine.
引文
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