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.

本报告期内，该研究项目取得了非常重大的进展。 迄今为止，大脑中大麻素 CB2 受体的存在一直存在争议。迄今为止，大多数证据表明，仅在大脑和中枢神经系统中发现了 CB1 大麻素受体，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，这种观点受到最近关于 CB2 受体存在于中枢神经系统中的说法以及最近关于 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 会减弱野生型和 CB1 基因缺失小鼠的静脉内可卡因自我给药，但不会减弱 CB2 基因缺失小鼠的静脉注射可卡因。这种效应被 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，并发现野生型小鼠静脉注射可卡因自我给药具有类似的抑制作用。在渐进比率强化条件下，我们发现 JWH133 抑制自我施用可卡因的激励动机，渐进比率断点的大幅降低证明了这一点。当 JWH133 鼻内给药（通过筛板直接进入大脑）或通过直接脑内显微注射 JWH133 进入伏隔核时，发现了类似的效果。同样，这种效应在野生型小鼠中出现，但在 CB2 受体基因缺失的小鼠中却没有出现。通过静脉自我给药和条件性位置偏好/厌恶实验评估，发现 JWH133 本身没有增强或厌恶作用。此外，JWH133 可以抑制野生型和 CB1 基因缺失小鼠中可卡因增强的运动，但不能抑制 CB2 基因缺失小鼠中的运动。 JWH133 本身对野生型和 CB1 基因缺失小鼠的全身给药和伏核脑内显微注射均具有运动抑制作用，但在 CB2 基因缺失小鼠中则没有。 CB2选择性拮抗剂AM630对野生型和CB1基因缺失小鼠的全身给药和伏核脑内显微注射均具有运动刺激作用，但在CB2基因缺失小鼠中则不然。通过实时体内脑微透析测量，JWH133 本身抑制细胞外伏隔核多巴胺。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核多巴胺。这种效应被 CB2 选择性拮抗剂 AM630 阻断。 AM630 本身——通过脑内显微注射到伏隔核中——增强了基底细胞外伏隔核多巴胺。我们得出结论，CB2 大麻素受体存在于大脑中，CB2 受体功能性调节中伏隔多巴胺系统，并且 CB2 受体功能性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/或厌恶作用以及实验室大鼠中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 delta-9-四氢大麻酚 (THC) 和 WIN55212-2 对大脑奖赏产生双相作用 - 低剂量增强大脑奖赏机制，高剂量抑制它们。 另一方面，选择性CB1大麻素受体激动剂ACEA仅产生大脑奖赏增强，而选择性CB2受体激动剂仅产生大脑奖赏抑制。 此外，选择性大麻素CB1受体拮抗剂AM251选择性阻断由低剂量THC或WIN55212-2产生的增强的脑奖赏，而选择性大麻素CB2受体拮抗剂AM630选择性阻断由高剂量THC或WIN55212-2产生的脑奖赏抑制。 TRPV1 拮抗剂辣椒西平（一些研究人员假设通过非 CB1、非 CB2 大麻素受体发挥作用）无法改变 THC 或 WIN55212-2 诱导的大脑奖赏变化。 此外，CB1 受体选择性拮抗剂 AM251（而非 CB2 受体选择性拮抗剂 AM630）可阻断 ACEA 增强的大脑奖赏。 CB2 受体选择性拮抗剂 AM630（而非 CB1 受体选择性拮抗剂 AM251）可阻断 JWH133 诱导的脑奖赏抑制。 鼻内 JWH133 抑制脑奖赏，这种作用可通过鼻内同时给予 CB2 受体选择性拮抗剂 AM630 来阻断。 我们得出的结论是，大麻素激动剂对大脑奖励机制产生双相效应，低剂量增强大脑奖励机制，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大脑 CB1 受体的激活介导的，而大麻素诱导的大脑奖励机制的抑制是通过大脑中 CB2 受体的激活介导的。 这些研究结果表明，大脑 CB1 和 CB2 受体相关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新的有效的药物治疗化合物来治疗药物成瘾和依赖性。 我们还将这项工作扩展到实验室大鼠，以观察大脑 CB2 受体在不同哺乳动物物种中的作用是否不同。 我们还开始了 JWH133 对多巴胺能神经元放电影响的电生理学研究 - 在单个解离的腹侧被盖区多巴胺神经元、中脑切片制剂中的腹侧被盖区多巴胺神经元以及活体麻醉动物的腹侧被盖区多巴胺神经元中。