Does neurokinin-1 modulate amphetamine reward via catecholamine transport?

 描述(申请人提供)：目前，尚无有效的药物治疗兴奋剂滥用、上瘾或复发。因此，有关神经底物被精神刺激药物修饰的新知识将极大地帮助确定新的靶点，以开发干预这些过程的治疗策略。去甲肾上腺素传递的改变和突触前去甲肾上腺素转运体(NET)的表达长期以来一直与抑郁症和药物成瘾有关。事实上，安非他明(Amph)针对的是net，并下调了其功能。我们已经证明Amph通过转运蛋白T258/S259转运基序下调Net功能和表面表达。我们还表明，神经激肽-1受体(NK1R)的激活通过同一基序的PKC磷酸化来负向调节Net。这些发现具有生理意义，因为(1)内源性NK1R激动剂P物质在AMPH治疗后释放。SP增强AMPH刺激效应，而NK1R拮抗剂阻断这种效应；(Ii)NK1R基因敲除小鼠表现出ADHD样表型；以及(Iii)AMPH是治疗人类ADHD的治疗剂。因此，Net、NK1R和Amph之间显然存在生理上的相关关系。在这方面，我们的发现NK1R和AMPH都介导了Net下调，并且都与T258/S259基序相连，这表明这一途径可能在调节Amph诱导的行为中发挥重要作用。我们的初步体内研究表明，Amph诱导的Net下调和运动激活对NK1R拮抗剂和针对T258/S259基序的操作敏感。我们提出了两个具体的目标来验证我们的假设，即NK1R介导的T258/S259特异性网络下调有助于AMPH行为效应。目的1验证NK1R介导AMPH诱导的小鼠净调节和奖励样行为的假说。我们将通过单独或联合脑核特异性微量注射激动剂和拮抗剂来确定腹侧纹状体NK1R在调节AMPH诱导的突触网络调节和条件性位置偏爱中的作用。目的2将验证这样的假设，即净T258/S259基序是AMPH介导的奖励样行为和净下调所必需的。我们将通过腹侧纹状体微量注射细胞通透性TAT-Net多肽来研究T258/S269基序的作用，这些TAT-Net多肽干扰依赖Amph的Net运输。由于多巴胺转运蛋白(DAT)也被NK1R和AMPH下调，而且Net和DAT在T258/S259区域都有高度保守的序列同源性，DAT-KO小鼠也将被用于这两个目的，以分离Net调节和DAT的作用。因此，了解NK1R如何调节胺转运体可能有助于我们确定AMPH扰乱儿茶酚胺转运体功能从而扰乱动物行为的特定机制(可能成为治疗靶点)。

 DESCRIPTION (provided by applicant): Currently, there are no proven medications to treat stimulant abuse, addiction or relapse. Therefore, new knowledge on the neuronal substrates that are modified by psychostimulant drugs will greatly aid in identifying novel targets for developing therapeutic strategies to intervene in these processes. Altered noradrenergic transmission and presynaptic norepinephrine transporter (NET) expression have long been associated with depression and drug-addiction. Indeed, amphetamine (AMPH) targets NET and downregulates its function. We have shown that AMPH downregulates NET function and surface expression via transporter T258/S259 trafficking motif. We have also shown that activation of the neurokinin-1 receptor (NK1R) negatively regulates NET via PKC phosphorylation of the same motif. These findings are of physiological significance because (i) Substance P, the endogenous NK1R agonist, is released following AMPH treatment. SP enhances AMPH stimulant effects, whereas NK1R antagonists block this effect; (ii) NK1R knockout mice exhibit ADHD-like phenotype; and (iii) AMPH is a therapeutic agent for treating ADHD in the human. Thus, there is, apparently a physiologically relevant relationship between NET, NK1R and AMPH. In this regard, our discovery that both NK1R and AMPH mediate NET downregulation and that both are linked to the T258/S259 motif suggests that this pathway may play a significant role in regulating AMPH-elicited behaviors. In support, our preliminary in vivo studies suggest that AMPH-induced NET downregulation and locomotor activation are sensitive to NK1R antagonists and to manipulations targeting the T258/S259 motif. We propose two specific aims to test our hypothesis that the NK1R-mediated T258/S259-specific NET downregulation contributes to AMPH behavioral effects. Aim 1 will test the hypothesis that NK1R mediates AMPH-induced NET regulation and reward-like behavior in mice. We will identify the role of ventral striatal NK1R in modulating AMPH-induced synaptic NET regulation and conditioned place preference by brain nuclei-specific microinjections of agonists and antagonists alone or in combination. Aim 2 will test the hypothesis that NET T258/S259 motif is required for AMPH-mediated reward-like behavior and NET downregulation in mice. We will study the role of the T258/S269 motif by ventral striatal microinjections of cell permeable TAT-NET peptides that interfere with the AMPH-dependent trafficking of NET. Since dopamine transporter (DAT) is also downregulated by both NK1R and AMPH, and both NET and DAT share highly conserved sequence homology at the T258/S259 region, DAT-KO mice will also be utilized in both aims to dissociate the role of NET regulation from that of DAT. Thus, understanding how NK1R regulates amine transporters may help us identify the specific mechanisms (which could be therapeutic targets) whereby AMPH disrupts catecholamine transporter function and hence animal behavior.