Serotonin Transporter Kinetics In Vivo by Microdialysis/Capillary UPLC

描述（由申请人提供）：我们建议开发一种分析方法，使用高温高压毛细管高效液相色谱法，结合在线微透析采样后的电子转移后光致发光检测（uplc - pet），以便在清醒行为动物的大脑中快速测量5 -羟色胺神经递质。与现有方法（如HPLC-EC， ce - liff）相比，该方法将在速度和检测限制方面提供实质性改进。它将广泛适用于其他单胺类神经递质及其代谢物和多肽。脑血清素神经递质系统调节许多重要的心理生理功能，包括情绪、焦虑状态、认知、奖励相关行为、运动活动和食欲行为。突触前质膜5 -羟色胺转运蛋白（SERT）从细胞外空间吸收5 -羟色胺，并在调节5 -羟色胺能神经传递中起核心作用。它是最广泛使用的抗抑郁和抗焦虑药物——血清素再摄取抑制剂（SRIs）的主要目标。在人类中，由一种常见的基因变异驱动的SERT表达减少与焦虑相关人格特征的增加有关。以及对压力相关抑郁症的易感性。为了研究SERT减少的影响，我们制造了表达SERT减少的小鼠。小鼠SERT降低导致焦虑样行为增加，这与在人类中观察到的相似。尽管5 -羟色胺摄取起着关键作用，但内源性5 -羟色胺摄取的动力学从未被测量过。我们将采用优化的uplc - pet方法来测量这些动力学。

DESCRIPTION (provided by applicant): We propose to develop an analytical method using capillary high-performance liquid chromatography at high temperature and pressure coupled to photoluminescence detection following electron transfer (UPLC-PFET) following online microdialysis sampling to enable fast serotonin neurotransmitter measurements in the brains of awake behaving animals. This method will provide substantial improvements in the speed and limits of detection over current methods (e.g., HPLC-EC, CE-LIF). It will be broadly applicable to other monoamine neurotransmitters and metabolites and peptides. The brain serotonin neurotransmitter system regulates many important psychophysiological functions including mood, anxiety states, cognition, reward-related behavior, motor activity and appetitive behavior. The presynaptic plasma membrane serotonin transporter (SERT) takes up serotonin from the extracellular space and plays a central role in regulating serotonergic neurotransmission. It is the primary target for the most widely prescribed class of antidepressant and anti-anxiety medications } the serotonin reuptake inhibitors (SRIs). In humans, reduced SERT expression driven by a commonly occurring gene variant has been associated with increases in anxiety-related personality traits. and susceptibility to stress-associated depression. To investigate effects of decreased SERT, we have produced mice that express reduced SERT. Decreased SERT in mice results in increased anxiety-like behavior similar to that observed in humans. Despite the key role that serotonin uptake plays, the kinetics of the uptake of endogenous serotonin has never been measured We will employ the optimized UPLC-PFET method developed here to measure these kinetics. PUBLIC HEALTH RELEVANCE: To understand the brain's internal communications, we need analytical methods that are absolutely selective and also fast. There are no general approaches that have both attributes. We will develop one, and apply it to a question about serotonin - the key player in anxiety, depression and related disorders.