5-HT Reuptake in SERT Knockout Mice by Voltammetry

这项建议涉及应用“快速”电化学方法来测定5-羟色胺转运体基因敲除小鼠脑组织中5-羟色胺的实时再摄取和释放。制造这些小鼠是为了更清楚地了解5-羟色胺转运体在正常行为以及焦虑和情绪障碍中的作用。5-羟色胺转运体基因敲除小鼠的自发焦虑样行为增加，运动激活减少，对滥用药物3，4-亚甲基二氧基甲基苯丙胺(MDMA)的反应。据推测，这些表型的改变直接归因于5-羟色胺转运体功能的长期下降，最终导致5-羟色胺能系统及其突触后靶点的神经适应性变化。到目前为止所研究的许多神经化学和行为参数显示，携带一个功能副本的5-羟色胺转运体基因的小鼠发生了中等水平的变化。然而，在杂合子基因敲除小鼠中，最初通过[~3H]5-羟色胺摄取来评估转运蛋白功能似乎没有改变。因此，我们假设，经典放射化学方法对5-羟色胺摄取的表征不能提供必要的时间分辨率来检测摄取过程动力学中的重要变化。所描述的研究将：(1)使用高速计时电流法评估5-羟色胺转运体敲除小鼠的突触体内摄取5-羟色胺的动力学；以及(2)使用快速扫描循环伏安法表征5-羟色胺转运体敲除小鼠脑片制备中的特定脑区5-羟色胺再摄取和释放的动力学。我们的目标是回答转运蛋白表达的中间变化是否会导致转运蛋白功能的显著改变的问题。这项建议代表了伏安技术在表征转基因小鼠模型中5-羟色胺能神经传递变化的新应用。

This proposal is concerned with the application of "fast" electrochemical methods for the determination of real-time serotonin reuptake and release in brain tissue derived from serotonin transporter knockout mice. These mice were produced to gain a clearer understanding of the role of the serotonin transporter in normative behavior, and in anxiety and mood disorders. Serotonin transporter knockout mice display elevations in spontaneous anxiety-like behavior and decreases in locomotor activation in response to 3,4- methylenedioxymethamphetamine (MDMA), a popular drug of abuse. It is hypothesized that these alterations in phenotype are directly attributable to long-term decreases in serotonin transporter function, which ultimately result in neuroadaptive changes in the serotonergic system and its postsynaptic targets. Many of the neurochemical and behavioral parameters studied to date show intermediate levels of change in mice bearing one functional copy of the serotonin transporter gene. However, transporter function initially assessed by [3H]serotonin uptake appears unaltered in heterozygote knockout mice. Therefore, we hypothesize that the characterization of serotonin uptake by classical radiochemical methods does not provide the temporal resolution necessary to detect important variations in the kinetics of the uptake process. The research described will: (1) Evaluate the kinetics of serotonin uptake using high-speed chronoamperometry in synaptosomes derived from serotonin transporter knockout mice; and (2) Characterize the dynamics of serotonin reuptake and release in specific brain regions in slice preparations from serotonin transporter knockout mice using fast scan cyclic voltammetry. We aim to answer the question of whether intermediate changes in transporter expression lead to significant modifications in transporter function. This proposal represents the novel application of voltammetric techniques to the characterization of changes in serotonergic neurotransmission in transgenic mouse models.