Dopamine Transporter--structure/function Studies Of Tran

多巴胺转运体(DAT)一直是大脑的主要受体部位，与可卡因的奖赏和愉悦特性相关。MNB科学家发现，DAT和SERT的缺失是消除小鼠对可卡因条件位置偏好所必需的。目前产生最佳帕金森氏病模型的每一种多巴胺选择性毒素的作用都需要DAT。对DAT结构-功能关系及其与SERT的关系的分析在这一年继续进行，进一步表征了选定的氨基酸侧链在转运蛋白功能中的作用。这些研究集中在大鼠多巴胺转运体中感兴趣的单域和多域氨基酸的变化，以及在今年报告的其他研究中发现的人类DAT序列等位基因变体所产生的氨基酸变化的特征。今年报告的研究记录了丝氨酸和苏氨酸取代对激活或抑制PKC、MAP、MEK和IP3激酶通路的药物的DAT活性的惊人的巨大影响。今年完成的研究还发现，PKC依赖的PP1抑制剂Kepi(在本实验室中被鉴定为吗啡上调基因)的共表达对共表达的DAT的功能有很大影响。今年报告的研究还记录了DAT点突变对DAT表达细胞的多巴胺外流的令人惊讶的选择性影响。这些见解将继续有助于鉴定可能在体内作为可卡因拮抗剂的小分子化合物的结构-功能特征，与DAT调节相关的结构-功能关系，以及DAT药理学中的人类个体差异。

The dopamine transporter (DAT) has been a principal brain receptor site that has been correlated with the rewarding and euphoric properties of cocaine. MNB scientists have found that deletion of DAT and SERT are required to eliminate cocaine conditioned place preferences in mice. DAT is required for the actions of each of the current dopamine-selective toxins that produce the best models of Parkinson's disease. Analyses of DAT structure- function relationships, and their relationships with SERT, continued during this year with further characterization of the roles of selected amino acid sidechains in transporter functions. These studies have focused on single- and multiple-domaine amino acid changes of interest in the rat dopamine transporter and on characterization of the amino acid changes produced by human allelic variants of the DAT sequence identified in other studies reported during this year. Studies reported during this year document surprisingly large effects of serine and threonine substitutions on the DAT activity effects of drugs that activate or inhibit PKC, MAP, MEK kinase, and IP3 kinase pathways. Studies completed during this year have also identified a large effect of coexpression of the PKC-dependent PP1 inhibitor, KEPI (identified in this laboratory as a morphine-upregulated gene)on the function of coexpressed DAT. Studies reported during this year also document surprisingly-selective effects of DAT point mutations on efflux of dopamine from DAT-expreessing cells. These insights should continue to help in identification of structure- function features of small molecule compounds possibly active in vivo as cocaine antagonists, structure-function relationships relevant to DAT regulation, and human individual differences in DAT pharmacology.