Mechanism of drug addiction

描述(申请人提供)：目前，还没有针对可卡因成瘾的药物治疗，部分原因是可卡因成瘾的机制尚不清楚。可卡因在大脑中有3个高亲和力靶点，即多巴胺转运体(DAT)、5-羟色胺转运体(SERT)和去甲肾上腺素转运体(NET)。可卡因结合并阻断了这些转运蛋白的摄取功能，导致突触中的递质水平延长和升高，这被认为是可卡因刺激和奖励作用的基础。令人惊讶的是，单独干扰DAT、Net或SERT的基因敲除小鼠仍然表现出可卡因奖赏，这表明可卡因奖赏不需要单一转运体，阻止可卡因抑制单一转运体的药物可能并不有效。然而，基因敲除小鼠的代偿性变化可能改变了奖励途径。为了避免代偿性变化，我们产生了一个携带DAT突变体的敲入小鼠系，该突变体保留了摄取功能，但对可卡因抑制不敏感(DAT-Ki小鼠)。在这些小鼠中，正常剂量的可卡因不再阻断DAT，刺激运动，或产生奖励。我们的结果表明，在DAT功能正常的小鼠中，需要阻断DAT才能获得可卡因奖励。最重要的是，它表明，拮抗可卡因对DAT的抑制的药物应该有效地阻断可卡因奖励。DAT-KI小鼠为研究复杂可卡因效应的机制提供了独特的新工具。我们建议继续我们目前的研究。我们将在其他几个行为测试中检测DAT-Ki小鼠的可卡因反应，以剖析DAT在可卡因复杂效应中的作用。此外，我们将使用AAV载体将野生型DAT重新引入DAT-Ki小鼠的选定脑区，以仅在这些选定区域恢复可卡因诱导的DA升高，并研究哪些可卡因反应被恢复。这一独特的方法使我们能够将特定大脑区域的可卡因行为与特定的可卡因效应联系起来。我们的初步数据显示，我们现在能够在有限的大脑区域和具有不同AAV注射的小鼠组中注射AAV，无论是在条件位置偏爱试验、仅运动刺激试验、两个试验中还是在不试验中恢复可卡因反应，证明了我们方法的可行性。拟议项目的成功将大大加强我们对可卡因如何产生复杂影响的了解，这对我们努力开发有效的可卡因成瘾治疗方法至关重要。公共卫生相关性：在拟议的研究中，我们将使用分子、遗传、生化和行为分析工具来了解药物成瘾的分子机制。拟议项目的成功将大大加强我们对可卡因如何产生复杂影响的了解，这对我们努力开发有效的可卡因成瘾治疗方法至关重要。

DESCRIPTION (provided by applicant): Currently, there are no pharmacological treatments for cocaine addiction partly because the mechanism of cocaine addiction is not clear. Cocaine has 3 high affinity targets in the brain, dopamine transporter (DAT), serotonin transporter (SERT), and norepinephrine transporter (NET). Cocaine binds and blocks the uptake functions of these transporters, resulting in prolonged and elevated transmitter levels in the synapses, which are believed to underlie the stimulating and rewarding effects of cocaine. Surprisingly, knockout mice with DAT, NET, or SERT individually disrupted still exhibit cocaine reward, suggesting that no single transporter is explicitly required for cocaine reward and drugs preventing cocaine inhibition of a single transporter may not be effective. However, compensatory changes in the knockout mice may have altered the reward pathway. To avoid the compensatory changes, we have generated a knock-in mouse line carrying a DAT mutant that retains the uptake function but is insensitive to cocaine inhibition (DAT-KI mice). In these mice, normal doses of cocaine no longer block DAT, stimulate locomotion, or produce reward. Our results indicate that DAT blockade is required for cocaine reward in mice with a functional DAT. Most significantly, it suggests drugs that antagonize cocaine inhibition of DAT should be effective in blocking cocaine reward. DAT-KI mice provide a unique novel tool to investigate the mechanism of the complex cocaine effects. We propose to continue our current studies. We will examine cocaine responses by DAT-KI mice in several other behavioral tests to dissect out the contribution of DAT in the complex effects of cocaine. In addition, we will use AAV vector to re-introduce wild type DAT back into selected brain regions of DAT-KI mice to restore cocaine-induced DA elevation in those selected regions only and study what cocaine responses are restored. This unique approach allows us to correlate cocaine actions in specific brain regions to specific cocaine effects. Our preliminary data show that we are now able to inject AAV in confined brain regions and groups of mice with varying AAV injections restored cocaine responses in either conditioned place preference test only, locomotor stimulation only, both tests, or none of the tests, demonstrating the feasibility of our approach. The success of the proposed project will significantly enhance our understanding on how cocaine produces its complex effects, which are crucial for our efforts in the development of effective treatment for cocaine addiction. PUBLIC HEALTH RELEVANCE: In the proposed study, we will use molecular, genetic, biochemical and behavioral analysis tools to understand the molecular mechanisms of drug addiction. The success of the proposed project will significantly enhance our understanding on how cocaine produces its complex effects, which are crucial for our efforts in the development of effective treatment for cocaine addiction.