【作者】 许月芳；

【导师】 李锦；

【作者基本信息】 中国人民解放军军事医学科学院 ， 药理学， 2008， 硕士

【摘要】 到目前为止,阿片类药物依然是其它药物无法替代的强效镇痛药,然而其强大的致躯体依赖和精神依赖潜能导致其在社会上的滥用。阿片依赖的病理生理学基础是机体长期接触阿片类物质产生的代偿性适应,涉及许多神经递质及其受体系统的改变。谷氨酸NMDA受体作用系统是阿片受体系统外最重要的阿片功能调节系统之一。谷氨酸是中枢神经系统中重要的兴奋性神经递质,在突触传递和神经元可塑性方面具有非常重要的作用。NMDA受体是一个重要的离子型谷氨酸受体,该受体是一种配体门控型阳离子通道,由基本亚基NR1和至少一个NR2调节亚基组成异聚体。NMDA受体的功能特性主要由组成异聚体的NR2亚基的特异性决定。编码NR2的基因有4种,即NR2A、NR2B、NR2C和NR2D。大量研究表明NR2B亚基直接参与了药物依赖的形成、发展和维持等全部病理生理过程,与阿片依赖关系十分密切。本研究旨在筛选和评价具有抗阿片依赖作用的新型选择性NR2B亚基拮抗剂,为阿片类药物脱毒和防复吸提供潜在的先导化合物。本课题以哌啶类化合物为先导结构,经过结构改造,合成了一系列候选化合物。由于激活NMDA受体后会导致钙离子大量内流,能够引发膜电流,因此我们在表达NMDA受体(NR1A/NR2B)的爪蟾卵母细胞上,利用双电极电压钳技术对化合物阻断NMDA受体通道电流作用进行初步筛选。此外,大量研究表明NR2B拮抗剂有一定的外周镇痛作用,因此我们采用醋酸扭体镇痛实验对候选化合物进行了镇痛活性初筛。结果显示Y-IP5等化合物阻断NMDA受体通道电流作用较强,Y-IP9、Y-IP10等化合物具有显著的镇痛作用。初步构效关系分析如下:1)当以哌嗪环代替哌啶环时,化合物保留镇痛活性,但通道电流阻断作用显著减弱;2)当以烷基胺如(美)金刚胺、苏胺等取代哌嗪环或哌啶环时,镇痛活性和通道电流阻断作用均显著减弱;3)当哌嗪环1位N原子和芳基之间的连接链为丙酰基时镇痛活性较好;连接链为丙酰基或乙酰胺基时通道电流阻断作用较强;4)当哌嗪环1位取代基上的芳基为苯并噁唑酮或苯并噻唑酮时,有显著的镇痛活性或通道电流阻断作用,进一步说明以杂环或苯酚的生物等排体代替苯酚基团的可行性。进一步选择在小鼠醋酸扭体模型上镇痛作用显著的化合物Y-IP9和Y-IP10和对NMDA受体通道电流阻断作用较强的化合物Y-IP5进行了药效学评价。Y-IP9、Y-IP10均具有显著的镇痛作用,在小鼠醋酸扭体模型上镇痛ED50值分别为3.3 mg·kg-1和3.8 mg·kg-1;对大鼠糖尿病性神经源性痛也都表现出显著的镇痛作用;Y-IP9、Y-IP10与大鼠脑膜制备阿片受体无亲和力,表明其镇痛作用不是通过阿片受体介导的。Y-IP9、Y-IP10具有一定的抗阿片依赖作用,均能显著抑制吗啡诱导小鼠条件性位置偏爱(CPP)的形成,而Y-IP10对吗啡所致小鼠躯体依赖的表达也有显著的抑制作用。Y-IP9、Y-IP10均能显著抑制小鼠的自发活动。Y-IP5阻断NMDA受体通道电流作用强,浓度为10μM时电流抑制率达到84.4%。该化合物能一定程度上对抗吗啡耐受。Y-IP5在(2.5-10.0 mg·kg-1)剂量范围内不影响小鼠的自发活动,但能剂量依赖性地抑制吗啡急性处理引起的活动增强。Y-IP5显著抑制小鼠吗啡躯体依赖的形成,并能显著抑制吗啡所致小鼠行为敏化及CPP的形成;表明其具有明显的抗阿片躯体和精神依赖潜能。综上所述,Y-IP5表现出明显的抗阿片耐受和依赖的作用,有可能作为一个潜在的防治阿片类药物依赖的先导化合物,其药理学作用值得进一步深入研究。更多还原

【Abstract】 Opioids have long been used for the treatment of moderate to severe pain. Despite their strong antinociceptive effects, the use of opioids in the treatment of pain is restricted by their physical and psychological dependence. In addition, opioids induce drug addiction among people. The pathophysiological mechanism of opioid dependence is related to the adaptation of many neurotransmitters and their receptors after chronic opioid treatment. Many studies demonstrated that glutamate-NMDA receptor system was an important modulatory system that has effect on the pharmacological actions of opioids.Glutamate is a major excitatory neurotransmitter in mammal central nervous system. NMDA receptor is an important ionotropic glutamate receptor, which is a ligand-gated positive ion channel composed by one NR1 subunit and at least one NR2 subunit. There are 4 genes encoding the NR2 subunit, including NR2A, NR2B, NR2C and NR2D. The property of NMDA receptor depended on the NR2 subunit which formed the heteromeric. NMDA receptor, especially NR2B subunit, plays important role in the pathophysiological process of drug dependence. Therefore, the main purpose of this investigation is to screen and evaluate novel selective NR2B antagonist that has inhibitory effect on opioid dependence, hopefully we can find potential new drugs for the prevention and therapy of opioid dependence and relapse.4-substituted piperdines were chosen as the lead structure. A series of novel compounds were designed and synthesized in this study. Because activation of NMDA receptors results in calcium influx which evokes membrane current. Xenopus oocytes expressing NMDA receptors (NR1A/NR2B) were established in this test. Two-electrode voltage clamp experiment was used to screen these compounds which had inhibitory effect on membrane current evoked by NMDA receptor channel in Xenopus oocytes. In addition, many studies revealed that this kind of antagonist had peripheral analgesic activity. Mice acetic acid writhing test was also used to screen these compounds. Y-IP5 blocked membrane current evoked by NMDA receptor channel significantly. Y-IP9 and Y-IP10 had relatively strong analgesic activity. Based on the results of preliminary screening, the structure-activity relationship was analyzed as follow:1) When the piperdine ring of lead structure was replaced by piperazine ring, the analgesic activity was retained,but the ability to block membrane current was attenuated. 2) When the piperazine ring or piperdine ring was replaced by bulky amino groups such as memantine, both the analgesic activity and the ability to block membrane current were attenuated significantly. 3) When the linker between 1-site of piperazine ring and aryl was ethylenecarbonyl, the analgesic activity and the ability to block membrane current were strong. 4) When the aryl group was 2-benzoxazolone-6-yl or 2-benzothioazolone-yl, the analgesic activity and the ability to block membrane current were retained or enhanced.Among these compounds, Y-IP9 and Y-IP10 had relatively strong analgesic activity. Y-IP5 blocked membrane current significantly. The pharmacological properties of these three compounds were investigated thereafter.Y-IP9 and Y-IP10 showed potent antinociceptive effects with ED50 of 3.31 mg·kg-1 and 3.80 mg·kg-1 respectively in the mice acetic acid writhing test. In rat diabetic neuropathy pain model these two compounds also showed antinociceptive effects. Y-IP9 and Y-IP10 bound opioid receptors with low affinity, which implicated their analgesic action were not related to activation of opioid receptor. Both Y-IP9 and Y-IP10 inhibited the development of morphine-induced conditioned place preference in mice. Y-IP10 also inhibited the expression of physical dependence in morphine-dependent mice. In addition, both Y-IP9 and Y-IP10 inhibited locomotor activity in mice. Y-IP5 blocked membrane current evoked by NMDA receptor channel with the inhibition rate of 84.4%. In chronic morphine tolerance model, Y-IP5 inhibited the development of morphine-induced tolerance. Y-IP5 didn’t influence locomotor activity in mice, but inhibited acute morphine-induced hyperactivity. Y-IP5 inhibited the development of morphine-induced physical dependence. Furthermore, Y-IP5 inhibited the development of morphine-induced behavioral sensitization and conditioned place preference in mice.In conclusion, Y-IP5 was found to have inhibitory effect on tolerance to and dependence on opioid, it could be taken as a potential leading compound in prevention and therapy of opioid dependence. Further investigation still needs to do to evaluate the possible effect of Y-IP5 on opioid dependence.更多还原