Physiology and Pharmacology of Brain Reward Circuits

大麻的主要精神活性成分被称为 δ9-四氢大麻酚 (THC)。此外，大脑中合成的内源性物质可以激活大麻素受体，这些物质被称为内源性大麻素（eCB）。所有作用于该物质受体的天然和合成药物统称为大麻素 (CB)。通过吸食或摄入大麻获得的大麻素药物之所以被使用，是因为它们通过与大脑的“奖励电路”相互作用来增强或奖励人类。这些研究的目的之一是了解大麻素改变脑细胞功能的潜在机制，以及最终产生这些药物维持其使用的愉悦效果的机制。该实验室的主要重点是研究滥用药物改变神经元电活动的机制以及这些神经元通过突触连接相互通信的方式。因此，我们的目标之一是确定特定的离子通道，其活性会被大麻、尼古丁、海洛因和可卡因等滥用药物改变。为了实现这些目标，我们利用从参与处理有关令人愉快和不愉快的环境刺激的信息的离散大脑区域敏锐获得的大鼠脑切片。 我们利用全细胞电生理记录和细胞解剖技术来重建我们记录的神经元。在这些正在进行的研究中，我们正在研究这些药物影响腹侧被盖区（VTA）神经元及其连接的机制。该大脑区域及其连接与所有滥用药物的强化和奖励作用以及调节食物、水等自然环境刺激的奖励作用密切相关。VTA 还参与处理有关生理应激反应、情绪和情感以及精神警觉性的信息。 由于 VTA 在这些过程中发挥着核心作用，因此它是导致成瘾、精神应激障碍、临床抑郁症和精神焦虑症等疾病的大脑区域。 Our recent work in the VTA is designed understand the mechanisms through which eCBs modulate VTA circuitry during psychostimulant exposure. Many studies show that antagonism of cannabinoid CB1Rs can block self-administration of several abused drugs. 此外，全身施用尼古丁、可卡因或乙醇后在伏核(NAc)中观察到的DA浓度增加通过全身施用CB1R拮抗剂/部分激动剂SR141716A2而大大降低。 该药物还会减少 NAc 中 DA 的释放，以响应奖励相关线索的出现，这表明 eCB 对于中脑边缘系统中因药物滥用相关的无条件和条件刺激而增加 DA 很重要。 还有证据表明，将 CB1R 拮抗剂直接输注到 VTA 中可以降低外周尼古丁的奖赏效应，并减少大鼠全身给予可卡因后 VTA 中 DA 的释放。 Data from our laboratory and others also show that eCBs are released from dopamine neurons in the VTA. 总的来说，这些数据表明，这些 eCB 在 VTA 内对 CB1R 起作用，在摄入滥用药物或暴露于药物配对刺激期间增加 NAc 中 DA 的释放。 尽管 CB1R 位于 VTA 内的 GABA 能和谷氨酸能轴突末端，但我们推测 THC 抑制 GABA 释放和去抑制 DA 神经元是其奖励效应的原因。 我们进一步假设滥用药物增加 DA 神经元活性会导致 eCB、2-花生四烯酰甘油 (2-AG) 释放，然后在 eCB 驱动的正反馈回路中通过抑制 GABA 释放来增强 DA 神经元兴奋。 出于这些原因，我们正在进行实验以确定滥用药物是否会触发 VTA 中的 eCB 释放，以及这种作用的机制可能是什么。 我们在 VTA 中发现的 eCB 功能最敏感的生理测量是突触 GABAB 受体介导的 IPSC，由电刺激激活。 这些反应受到突触前 CB1R 的强烈抑制，并受到 eCB、2-AG 的强弱抑制，因为在存在 CB1R 拮抗剂的情况下，或者当全细胞移液管中包含二酰基甘油脂肪酶-α (DGL-α) 抑制剂 THL 抑制 2-AG 合成时，它们的振幅会增加。 此外，当 DA 神经元短暂去极化以激发动作电位时，可以观察到阶段性、活动依赖性 2-AG 释放，这也是由 2-AG 释放增加介导的。 我们最近获得的数据表明，VTA DA 神经元中的 GABA-B IPSC 受到可卡因 (10 µM) 的抑制，并且可被 CB1R 拮抗剂 AM251 (2 µM) 减弱。可卡因的作用部分是由记录的神经元和周围细胞释放的 2-AG 介导的，因为与细胞内相比，THL 在细胞外应用时会更强烈地阻止这种作用。 These data suggest that cocaine stimulates the release of 2-AG in the VTA to reduce GABA release onto DA neurons. 我们假设可卡因抑制作用的减少会增加中脑 DA 神经元的兴奋性，从而增加 NAc 等投射区域中 DA 的释放，如体内研究所述。 由于可卡因的这种作用可能对其奖励和成瘾特性很重要，因此我们正在进行额外的实验来确定可卡因在 VTA 中释放 2-AG 的机制。 我们最近扩展了这些研究，表明仅在 DA 神经元中特异性敲除 DGL-α 可在体外阻止可卡因诱导的 VTA 中的 2-AG 功能。 此外，我们现在正在探索 2-AG 在 VTA 中释放的机制，并有强有力的证据表明，这是通过 sigma1 受体和特定细胞内信号通路的协调作用分泌的外泌体发生的。 总之，这些研究将确定可卡因诱导的 2-AG 释放在大脑奖励区域中的新作用，并将确定这些脂质分子释放的机制。

The main psychoactive component of marijuana is known as delta9-tetrahydrocannabinol (THC). In addition, endogenous substances are synthesized in the brain that can activate cannabinoid receptors, and these substances are referred to as endocannabinoids (eCB). All drugs, both natural and synthetic, that act at receptors for this substance are known collectively as cannabinoids (CBs). Cannabinoid drugs obtained by the smoking or ingestion of marijuana are used because they are reinforcing or rewarding to humans through interaction with the brain's "reward circuitry". One of the objectives of these studies is to gain knowledge about the underlying mechanisms through which cannabinoids alter brain cell function, and ultimately the mechanisms that produce the pleasurable effects of these drugs that sustain their use. The primary focus of this laboratory is to examine the mechanisms through which abused drugs alter the electrical activity of neurons and the ways in which these neurons communicate with each other via synaptic connections. Therefore, one of our goals is to identify specific ion channels whose activity is modified by abused drugs such as marijuana, nicotine, heroin, and cocaine. To achieve these goals we utilize rat brain slices acutely obtained from discrete brain areas involved in processing information regarding pleasurable and unpleasant environmental stimuli. We utilize whole-cell electrophysiological recordings, and cellular anatomical techniques to reconstruct the neurons from which we record. In these ongoing studies we are examining the mechanisms through which these drugs affect neurons and their connections in the ventral tegmental area (VTA). This brain area and its connections are strongly implicated in the reinforcing and rewarding actions of all abused drugs, as well as in mediating the rewarding effects of natural environmental stimuli, such as food, water, etc. The VTA is also involved in processing information regarding the physiological stress responses, mood and affect, and mental alertness. Because of its central role in these processes, the VTA is a brain area that contributes to disorders such as addiction, psychiatric stress disorders, clinical depression, and psychiatric anxiety disorders. Our recent work in the VTA is designed understand the mechanisms through which eCBs modulate VTA circuitry during psychostimulant exposure. Many studies show that antagonism of cannabinoid CB1Rs can block self-administration of several abused drugs. Furthermore, the increase in DA concentration that is observed in the nucleus accumbens (NAc) following systemic nicotine, cocaine or ethanol administration is greatly reduced by systemic administration of the CB1R antagonist/partial agonist SR141716A2. This drug also decreases DA release in the NAc in response to the presentation of reward-associated cues, suggesting that eCBs are important for DA increases in the mesolimbic system by unconditioned and conditioned stimuli associated with drug abuse. There is also evidence that infusion of CB1R antagonists directly into the VTA can reduce the rewarding effect of peripheral nicotine, and reduce the release of DA in the VTA following systemic cocaine administration in rats. Data from our laboratory and others also show that eCBs are released from dopamine neurons in the VTA. Collectively, these data suggest that these eCBs act within the VTA on CB1Rs to increase the release of DA in the NAc during the intake of abused drugs, or during exposure to drug-paired stimuli. Although CB1Rs are located on both GABAergic and glutamatergic axon terminals within the VTA, we hypothesize that it is the inhibition of GABA release and disinhibition of DA neurons by THC that is responsible for its rewarding effects. We further hypothesize that the increase in DA neuron activity by abused drugs causes release of the eCB, 2-aracchidonoylglycerol (2-AG), which then augments DA neuron excitation through inhibition of GABA release, in an eCB-driven positive feedback loop. For these reasons we are conducting experiments to determine whether abused drugs trigger eCB release in the VTA, and what mechanisms might underlie this action. The most sensitive physiological measure of eCB function that we have found in the VTA are synaptic GABAB-receptor-mediated IPSCs, activated by electrical stimulation. These responses are strongly inhibited by presynaptic CB1Rs, and are tonically inhibited by the eCB, 2-AG, since their amplitudes are increased in the presence of CB1R antagonists, or when 2-AG synthesis is inhibited by inclusion of a diacylglycerol lipase-alpha (DGL-alpha) inhibitor, THL, in the whole-cell pipettes. Additionally, phasic, activity-dependent 2-AG release can be observed when DA neurons are transiently depolarized to fire action potentials, and this is also mediated by an increase in 2-AG release. Our recently acquired data show that GABA-B IPSCs in VTA DA neurons are inhibited by cocaine (10 microM), and this is reduced by the CB1R antagonist AM251 (2 microM). The effect of cocaine is partly mediated by 2-AG released from the recorded neuron, and from surrounding cells, since THL blocks the effect more strongly when applied extracellularly, versus intracellularly. These data suggest that cocaine stimulates the release of 2-AG in the VTA to reduce GABA release onto DA neurons. We hypothesize that the reduction of inhibition by cocaine increases the excitability of midbrain DA neurons to augment release of DA in projection areas like the NAc, as described in in vivo studies. Because this cocaine action may be important for its rewarding and addicting properties, we are performing additional experiments to determine the mechanism(s) by which cocaine releases 2-AG in the VTA. We have recently expanded these studies by showing that specifically knocking DGL-alpha only in DA neurons prevents cocaine-induced 2-AG function in the VTA in vitro. Moreover, we are now exploring the mechanisms through which 2-AG is released in the VTA and have strong evidence that this occurs via exosomes that are secreted via the coordinated actions of the sigma1 receptor and specific intracellular signaling pathways. Together, these studies will identify a novel role for cocaine-induced 2-AG release in this brain reward area, and will identify the mechanisms through which these lipid molecules are released.