5-HT Reuptake in SERT Knockout Mice by Voltammetry

该建议涉及应用“快速”电化学方法测定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。 这些小鼠的产生是为了更清楚地了解5-羟色胺转运体在规范行为、焦虑和情绪障碍中的作用。 5-羟色胺转运蛋白基因敲除小鼠对3，4-亚甲二氧基甲基苯丙胺（MDMA）（一种流行的滥用药物）的反应显示自发性焦虑样行为升高和运动激活降低。据推测，这些表型的改变是直接归因于长期的血清素转运蛋白功能的下降，最终导致神经适应性的变化，在多巴胺能系统及其突触后的目标。迄今为止研究的许多神经化学和行为参数显示，携带一个功能性5-羟色胺转运体基因拷贝的小鼠发生了中等水平的变化。 然而，转运功能最初评估的[3 H]血清素摄取出现在杂合子敲除小鼠不变。 因此，我们假设，通过经典的放射化学方法的5-羟色胺摄取的表征不提供必要的时间分辨率来检测摄取过程的动力学的重要变化。 所述研究将：（1）使用高速计时电流法在源自5-羟色胺转运蛋白敲除小鼠的突触体中评价5-羟色胺摄取的动力学;和（2）使用快速扫描循环伏安法表征5-羟色胺转运蛋白敲除小鼠的切片制备物中特定脑区域中5-羟色胺再摄取和释放的动力学。 我们的目标是回答这个问题，是否转运蛋白表达的中间变化导致转运蛋白功能的显着修改。 该建议代表了伏安技术在转基因小鼠模型中表征多巴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.