Endocannabinoid brain mechanisms and addiction

内源性大麻素脑机制和成瘾

• 批准号: 8736746
• 负责人: Eliot Gardner
• 金额: $ 42.56万
• 依托单位: NATIONAL INSTITUTE ON DRUG ABUSE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8736746
• 关键词: AM 251; AM630; Addictive Behavior; Agonist; Animal Model; Animals; Attenuated; Behavior; Behavioral; Biochemical; Biological Assay; Brain; CNR1 gene; CNR2 gene; Cannabinoids; Cells; Chemicals; Cocaine; Cues; Dependence; Dopamine; Dose; Drug Addiction; Electrical Stimulation of the Brain; Endocannabinoids; Enzymes; Exposure to; Genes; Goals; High Pressure Liquid Chromatography; Immune system; Immunoblotting; Incentives; Intravenous; Knowledge; Laboratory Animals; Laboratory Rat; Life; Link; Locomotion; Measures; Mediating; Microdialysis; Microinjections; Midbrain structure; Motivation; Mus; Neuraxis; Neurons; Neurotransmitters; Nucleus Accumbens; Pharmaceutical Preparations; Polymerase Chain Reaction; Preparation; Proteins; Psychological reinforcement; RNA; Receptor Gene; Relapse; Reporting; Research; Research Personnel; Research Project Grants; Reverse Transcription; Rewards; Role; Sampling; Self Administration; Self-Administered; Slice; Stress; Synapses; System; TRPV1 gene; Techniques; Tetrahydrocannabinol; Time; Ventral Tegmental Area; Western Blotting; Wild Type Mouse; Work; addiction; cannabinoid receptor; capsazepine; craving; dopamine system; dopaminergic neuron; extracellular; in vivo; neurochemistry; overexpression; pre-clinical; preference; receptor; relating to nervous system; research study

During the present reporting period, very significant progress was made on this research project. The existence of cannabinoid CB2 receptors in the brain has been heretofore controversial. Most evidence has heretofore suggested that only CB1 cannabinoid receptors are found in brain and central nervous system while cannabinoid CB2 receptors are restricted to the body's periphery - primarily in the immune system. However, this view has been challenged by recent claims that CB2 receptors are present in the central nervous system and by recent claims that CB2 receptors modulate synaptic activity. Therefore, we used highly selective CB2 agonists and antagonists, combined with the use of CB1 and CB2 receptor gene-deleted mice, to study CB2 involvement in cocaine's behavioral and neurochemical effects. We found that the CB2 receptor-selective agonist JWH133 attenuates intravenous cocaine self-administration in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. This effect was abolished by the CB2 receptor-selective antagonist AM630. To confirm our findings, we also used the CB2-selective agonist GW405833 and found a similar inhibition of intravenous cocaine self-administration in wild-type mice. Under progressive-ratio reinforcement conditions, we found that JWH133 inhibits incentive motivation to self-administer cocaine, as evidenced by strong reductions in the progressive-ratio break-point. Similar effects were found when JWH133 was administered intra-nasally (for direct passage into the brain via the cribiform plate) or administered by direct intracerebral microinjections of JWH133 into the nucleus accumbens. Again, the effect was seen in wild-type but not in CB2 receptor gene-deleted mice. JWH133 by itself was found to have no reinforcing or aversive effects, as assessed by intravenous self-administration and by conditioned place preference/aversion experiments. Further, JWH133 inhibited cocaine-enhanced locomotion in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. JWH133 by itself had an inhibitory effect on locmotion, both with systemic administration and with intracerebral microinjection into the nucleus accumbens in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. The CB2 selective antagonist AM630 had a stimulatory effect on locomotion, both with systemic administration and with intracerebral microinjection into the nucleus accumbens in wild-type and CB1 gene-deleted mice, but not in CB2 gene-deleted mice. JWH133 by itself inhibited extracellular nucleus accumbens dopamine as measured by real-time in vivo brain microdialysis. JWH133 also inbited basal and cocaine-enhanced extracellular nucleus accumbens dopamine as measured by real-time in vivo brain microdialysis. This effect was blocked by the CB2-selective antagonist AM630. By itself, AM630 - microinjected intracerebrally into the nucleus accumbens - aumented basal extracellular nucleus accumbens dopamine. We conclude that CB2 cannabinoid receptors exist in the brain, that CB2 receptors functionally modulate the meso-accumbens dopamine system, and that CB2 receptors functionally modulate dopamine-mediated behaviors. In addition, we used the electrical brain-stimulation reward preclinical animal model to study the rewarding and/or aversive effects of several cannabinoids and the receptor mechanisms underlying these actions in laboratory rats. We found that the mixed CB1/CB2 cannabinoid agonists delta-9-tetrahydrocannabinol (THC) and WIN55212-2 produce biphasic effects on brain reward - low doses enhancing brain reward mechanism and high doses inhibiting them. On the other hand, the selective CB1 cannabinoid receptor agonist ACEA produces only brain-reward enhancement, while the selective CB2 receptor agonist produces only brain-reward inhibition. Further, the selective cannabinoid CB1 receptor antagonist AM251 selectively blocks the enhanced brain reward produced by low dose THC or WIN55212-2, while the selective cannabinoid CB2 receptor antagonist AM630 selectively blocks the brain reward inhibition produced by high dose THC or WIN55212-2. The TRPV1 antagonist capsazepine (posited by some researchers to act via a non-CB1, non-CB2 cannabinoid receptor) fails to alter THC- or WIN55212-2-induced changes in brain reward. In addition, the CB1 receptor selective antagonist AM251, but not the CB2 receptor selective antagonist AM630, blocks ACEA-enhanced brain reward. The CB2 receptor selective antagonist AM630, but not the CB1 receptor selective antagonist AM251, blocks JWH133-induced inhibition of brain reward. Intranasal JWH133 inhibits brain reward, an effect that is blocked by intranasal co-administration of the CB2 receptor selective antagonist AM630. We conclude that cannabinoid agonists produce biphasic effects on brain reward, with low doses enhancing and high doses inhibiting brain reward mechanisms. We further conclude that cannabinoid-induced reward enhancement is mediated by activation of brain CB1 receptors, while cannabinoid-induced inhibition of brain reward mechanisms is mediated by activation of CB2 receptors in the brain. These research findings suggest that brain CB1 and CB2 receptor-linked neural systems may functionally antagonize each other in a reciprocal mutually antatagonistic manner. Such mechanistic knowledge can aid in the search for new and effective pharmacotherapeutic compounds for the treatment of drug addiction and dependence. We have also extended this work to laboratory rats, to see if brain CB2 receptors act differently in a different mammalian species. We have also started electrophysiological studies of the effects of JWH133 on dopaminergic neuronal cell firing - in single dissociated ventral tegmental area dopamine neurons, in ventral tegmental area dopamine neurons in midbrain slice preparations, and in ventral tegmental area dopamine neurons in live anesthetized animals.

在本报告所述期间，这项研究项目取得了非常重大的进展。迄今为止，大麻素CB2受体在大脑中的存在一直存在争议。到目前为止，大多数证据表明，只有CB1大麻素受体存在于大脑和中枢神经系统，而大麻素CB2受体仅限于身体的外周-主要是在免疫系统中。然而，这一观点受到了最近CB2受体存在于中枢神经系统和CB2受体调节突触活动的说法的挑战。因此，我们使用高选择性的CB2激动剂和拮抗剂，结合使用CB1和CB2受体基因缺失的小鼠，研究CB2参与可卡因的行为和神经化学作用。我们发现CB2受体选择性激动剂JWH133减少了野生型和CB1基因缺失小鼠的静脉可卡因自我给药，但在CB2基因缺失小鼠中没有。这种作用被CB2受体选择性拮抗剂AM630消除。为了证实我们的发现，我们还使用了cb2选择性激动剂GW405833，并在野生型小鼠中发现了类似的静脉注射可卡因自我给药抑制作用。在递进比强化条件下，我们发现JWH133抑制了自我服用可卡因的激励动机，这可以通过递进比断点的显著降低来证明。当JWH133通过鼻内（通过筛板直接进入大脑）或直接将JWH133脑内显微注射到伏隔核时，也发现了类似的效果。同样，这种效果在野生型小鼠中出现，而在CB2受体基因缺失的小鼠中没有出现。通过静脉注射自我给药和条件位置偏好/厌恶实验，发现JWH133本身没有强化或厌恶作用。此外，JWH133在野生型和CB1基因缺失小鼠中抑制可卡因增强的运动，但在CB2基因缺失小鼠中没有。在野生型和CB1基因缺失的小鼠中，JWH133本身具有全身给药和脑内微注射到伏隔核的抑制运动作用，但在CB2基因缺失的小鼠中没有。CB2选择性拮抗剂AM630对野生型和CB1基因缺失小鼠的运动有刺激作用，无论是全身给药还是脑内微注射到伏隔核，但对CB2基因缺失小鼠没有刺激作用。通过实时活体脑微透析检测，JWH133本身抑制伏隔核胞外多巴胺。通过实时体内脑微透析测量，JWH133还抑制基础和可卡因增强的细胞外伏隔核多巴胺。这种作用被cb2选择性拮抗剂AM630阻断。AM630在脑内注入伏隔核，增加伏隔核基底细胞外多巴胺。我们得出结论，CB2大麻素受体存在于大脑中，CB2受体功能调节中伏隔多巴胺系统，CB2受体功能调节多巴胺介导的行为。此外，我们使用脑电刺激奖励临床前动物模型来研究几种大麻素的奖励和/或厌恶作用以及这些作用背后的受体机制。我们发现混合CB1/CB2大麻素激动剂δ -9-四氢大麻酚（THC）和WIN55212-2对脑奖励产生双相作用-低剂量增强脑奖励机制，高剂量抑制脑奖励机制。另一方面，选择性CB1大麻素受体激动剂ACEA只产生脑奖励增强，而选择性CB2受体激动剂只产生脑奖励抑制。此外，选择性大麻素CB1受体拮抗剂AM251选择性阻断低剂量THC或WIN55212-2产生的脑奖励增强，而选择性大麻素CB2受体拮抗剂AM630选择性阻断高剂量THC或WIN55212-2产生的脑奖励抑制。TRPV1拮抗剂capsazepine（一些研究人员假设通过非cb1，非cb2大麻素受体起作用）不能改变THC-或win55212 -2诱导的脑奖励变化。此外，CB1受体选择性拮抗剂AM251，而不是CB2受体选择性拮抗剂AM630，阻断了acea增强的脑奖励。CB2受体选择性拮抗剂AM630能阻断jwh133诱导的脑奖励抑制，而CB1受体选择性拮抗剂AM251则不能。鼻内JWH133抑制脑奖励，这一作用被鼻内联合给药CB2受体选择性拮抗剂AM630阻断。我们得出结论，大麻素激动剂对大脑奖励产生双相作用，低剂量增强和高剂量抑制大脑奖励机制。我们进一步得出结论，大麻素诱导的奖励增强是通过激活脑CB1受体介导的，而大麻素诱导的脑奖励机制抑制是通过激活脑CB2受体介导的。这些研究结果表明，脑CB1和CB2受体连接的神经系统可能以相互拮抗的方式在功能上相互拮抗。这种机制知识可以帮助寻找治疗药物成瘾和依赖的新的有效药物治疗化合物。我们还将这项工作扩展到实验室老鼠身上，看看脑CB2受体在不同哺乳动物物种中的作用是否不同。我们也开始了JWH133对多巴胺能神经元细胞放电影响的电生理研究——在单个解离的腹侧被盖区多巴胺神经元中，在中脑切片制备的腹侧被盖区多巴胺神经元中，以及在麻醉活体动物的腹侧被盖区多巴胺神经元中。