Uptake2 transporters: Novel sex-dependent molecular targets to treat stimulant use disorder

治疗物质使用障碍（SUD）的药物存在于许多滥用药物中，但不包括精神兴奋剂， 例如安非他明（AMPH）和甲基安非他明。他们也缺乏越来越多地使用合成 模仿已知精神兴奋剂作用的药物。已知的和新的精神活性物质 继续构成重大和日益严重的公共卫生威胁。为了开发有效的治疗方法， 需要更好地了解这些兴奋剂是如何产生与滥用有关的影响的。许多兴奋剂相互作用 与多巴胺（DA）转运蛋白（DAT），这被认为是介导其滥用相关的影响。然而，在这方面， 针对DAT的策略在治疗精神兴奋剂成瘾方面几乎没有任何益处， 这些兴奋剂可能在其他地方具有调节多巴胺能神经传递的重要作用。 与此一致，快速增长的文献支持有机阳离子转运蛋白3（OCT 3）的突出作用。 调节多巴胺能神经传递。我们的初步数据支持这一观点，表明decyphin-22 (D22)OCT 3的抑制剂，在雄性OCT 3野生型（WT）小鼠体内抑制DA释放，这种作用在 组成型OCT 3敲除（KO）小鼠。此外，体外研究表明，AMPH诱导的底物外排 D22可以以独立于可卡因敏感转运蛋白的方式抑制。这些发现是 通过雄性小鼠对AMPH的条件性位置偏好（CPP）的丧失，以及雌性OCT 3 KO的减弱， 其中D22在两种WT性别中预防CPP。雄性小鼠的初步数据显示， 在雄性OCT 3 KO小鼠中，AMPH的自我给药相对于它们的 WT对应物。根据这些发现，我们建立并验证了OCT 3 floxed小鼠，以检查 OCT 3在不存在任何可能发生在种系科斯中的发育补偿时的作用。利用 这些小鼠的能力，新的数据表明，他莫昔芬诱导或病毒KO的OCT 3有力地减弱AMPH- 诱发DA释放和CPP。拟议的研究将询问OCT 3在AMPH影响能力中的作用 行为和DA外排通过测试新的和潜在的变革性假设，AMPH驱动DA释放 不仅由DAT介导，而且也由经由OCT 3的反向转运介导。因此，DAT-OCT 3冗余可 解释为什么DAT为基础的治疗已在治疗AMPH使用障碍不成功。我们将使用一个 结合药理学，新的和建立的转基因小鼠模型（DA神经元特异性与神经胶质细胞， 特异性）和脑区域特异性病毒敲低，以确定OCT 3在AMPH诱导的行为抑制中的作用。 作用（运动、直立、刻板、致敏、CPP、自我给药）（目的1）和AMPH诱导的DA 体内外排和清除（目的2）。在目标3中，我们将确定他莫昔芬如何诱导OCT 3的KO或病毒KO 影响DA信号传导的关键要素以及OCT 3如何适应慢性AMPH自我给药。这些研究 将填补关于OCT 3在AMPH滥用相关影响中的重要性的基本知识空白， 这将有助于确定治疗AMPH成瘾药物的新分子靶点。

Medications to treat substance use disorders (SUD) exist for many drugs of abuse, but not for psychostimulants, such as amphetamine (AMPH), and methamphetamine. They are also lacking for increasingly used synthetic drugs designed to mimic the actions of known psychostimulants. Both known and new psychoactive substances continue to pose a major and increasing public health threat. To develop effective treatments, the mechanisms by which these stimulants produce their abuse-related effects need to be better understood. Many stimulants interact with the dopamine (DA) transporter (DAT), which is thought to mediate their abuse-related effects. However, strategies targeting DAT have yielded little to no benefit in the treatment of psychostimulant addiction, raising the possibility that these stimulants have significant actions elsewhere to modulate dopaminergic neurotransmission. Consistent with this, a rapidly growing literature supports a prominent role for organic cation transporter 3 (OCT3) in regulating dopaminergic neurotransmission. Our preliminary data support this idea, showing that decynium-22 (D22), an inhibitor of OCT3, inhibits DA release in vivo in male OCT3 wild-type (WT) mice, an effect lost in constitutive OCT3 knockout (KO) mice. Furthermore, in vitro studies showed that AMPH-induced substrate efflux could be inhibited by D22 in a manner independent of cocaine-sensitive transporters. These finding were paralleled by a loss of conditioned place preference (CPP) for AMPH in male mice, and an attenuation in female OCT3 KO, with D22 preventing CPP in both WT sexes. Preliminary data in male mice show that the dose-response curve for self-administration of AMPH is dramatically downward and rightward shifted in male OCT3 KO mice relative to their WT counterpart. In light of these findings, we have established and validated OCT3 floxed mice so as to examine the role of OCT3 in the absence of any developmental compensation that may occur in germline KOs. Harnessing the power of these mice, new data show that tamoxifen inducible or viral KO of OCT3 robustly attenuates AMPH- evoked DA release and CPP. Proposed studies will interrogate OCT3’s role in the ability of AMPH to impact behavior and DA efflux by testing the novel and potentially transformative hypothesis that AMPH driven DA release is not mediated solely by DAT but by reverse transport via OCT3 as well. Thus, DAT-OCT3 redundancy may explain why DAT-based therapeutics have been unsuccessful in treating AMPH use disorders. We will use a combination of pharmacology, novel and established transgenic mouse models (DA neuron-specific vs glia- specific), and brain-region specific viral knockdown, to determine the role of OCT3 in AMPH-induced behavioral effects (locomotion, rearing, stereotypy, sensitization, CPP, self-administration) (Aim 1), and AMPH-induced DA efflux and clearance in vivo (Aim 2). In Aim 3 we will determine how tamoxifen inducible KO or viral KO of OCT3 impacts key elements in DA signaling and how OCT3 adapts to chronic AMPH self-administration. These studies will fill fundamental knowledge gaps about the importance of OCT3 in abuse-related effects of AMPH, and its congeners, which will help identify novel molecular targets for medications to treat AMPH addiction.