Biological And Biochemical Characterization Of Sigma Receptors

自从我在NIDA IRP的实验室于1982年发现了sigma-1受体以来，许多临床前研究表明sigma-1受体和相关配体与中风、健忘症、抑郁症、癌症、阿尔茨海默病、疼痛和可卡因成瘾有关。 在2019-2020财年，我们对可卡因的新作用有了根本性的发现。可卡因是一种成瘾性药物，作用于大脑奖励区域。最近的证据表明，可卡因刺激中脑内源性大麻素2-花生四烯酸甘油（2-AG）的合成，通过去抑制增加多巴胺神经元的活性。虽然可卡因刺激的2-AG合成的机制是已知的，我们对2-AG释放的理解是有限的。在NG 108细胞和小鼠中脑组织中，我们发现2-AG位于非突触细胞外囊泡（EV）中，这些囊泡在可卡因存在下通过与伴侣蛋白sigma-1受体（Sig-1 R）相互作用分泌。当可卡因导致Sig-1 R从ADP-核糖基化因子（ARF 6）（一种调节EV运输的G蛋白）解离，导致肌球蛋白轻链激酶（MLCK）活化时，EV释放。阻断Sig-1 R功能，或抑制ARF 6或MLCK也阻止了可卡因诱导的EV释放和可卡因刺激的DA神经元抑制性突触的2-AG调制。我们的研究结果表明，Sig-1 R-ARF 6复合物控制EV释放，并证明可卡因介导的2-AG释放可以通过EV发生。 我们还发现了一个基本的机制，除了作用于质膜上传统上公认的受体或转运蛋白外，几种滥用药物，包括安非他明，甲基苯丙胺，尼古丁，阿片类药物，可卡因，氯胺酮和大麻素，可以通过作用于质膜上或细胞内的其他分子靶点来发挥其作用。这些目标通常是新生受体或蛋白质，可以引起下游信号传导或分子事件，导致有利于成瘾过程的生理结果改变。然而，这些非典型的滥用药物的目标，一般来说，没有得到广泛的认识，在药物滥用研究。这一观点将注意力转移到那些认识不足的目标上，以期促进对滥用药物的作用有更全面的了解。

In the years since my laboratory at the NIDA IRP identified the sigma-1 receptor in 1982, many preclinical studies have shown that sigma-1 receptors and associated ligands are involved in stroke, amnesia, depression, cancer, Alzheimers disease, pain, and cocaine addiction. In this fiscal year, 2019-2020, we made a fundamental discovery on the new action of cocaine. Cocaine is an addictive drug that acts in brain reward areas. Recent evidence suggests that cocaine stimulates synthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) in midbrain, increasing dopamine neuron activity via disinhibition. Although a mechanism for cocaine-stimulated 2-AG synthesis is known, our understanding of 2-AG release is limited. In NG108 cells and mouse midbrain tissue, we find that 2-AG is localized in non-synaptic extracellular vesicles (EVs) that are secreted in the presence of cocaine via interaction with the chaperone protein sigma-1 receptor (Sig-1R). The release of EVs occurs when cocaine causes dissociation of the Sig-1R from ADP-ribosylation factor (ARF6), a G-protein regulating EV trafficking, leading to activation of myosin light chain kinase (MLCK). Blockade of Sig-1R function, or inhibition of ARF6 or MLCK also prevented cocaine-induced EV release and cocaine-stimulated 2-AG-modulation of inhibitory synapses in DA neurons. Our results implicate the Sig-1R-ARF6 complex in control of EV release and demonstrate that cocaine-mediated 2-AG release can occur via EVs. We also made a fundamental mechanistic finding that in addition to acting on traditionally recognized receptors or transporters on the plasma membrane, several drugs of abuse, including amphetamine, methamphetamine, nicotine, opioid, cocaine, ketamine, and cannabinoid can work to exert their effects by acting on additional molecular targets either on the plasma membrane or inside a cell. These targets are usually nascent receptors or proteins that can cause downstream signaling or molecular events, leading to altered physiological outcomes favoring addictive processes. However, those non-canonical targets of drugs of abuse, in general, have not been widely recognized in drug abuse research. This perspective diverts attention to those underrecognized targets, in the hope of promoting a more complete understanding of the action of drugs of abuse.