Serotonin Transporter Kinetics In Vivo by Microdialysis/Capillary UPLC

描述(申请人提供)：我们建议开发一种分析方法，使用高温高压下的毛细管高效液相色谱与在线微透析采样后的光致发光检测(UPLC-PFET)相结合，以实现对清醒行为动物大脑中5-羟色胺神经递质的快速测量。这种方法将在检测速度和限度方面比目前的方法(例如，高效液相色谱-EC，CE-LIF)提供实质性的改进。它将广泛适用于其他单胺类神经递质和代谢物和多肽。大脑5-羟色胺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、奖赏相关行为、运动活动和食欲行为。突触前质膜5-羟色胺转运体(SERT)从细胞外间隙摄取5-羟色胺，在调节5-羟色胺能神经传递中发挥重要作用。它是最广泛使用的抗抑郁和抗焦虑药物--5-羟色胺再摄取抑制剂(SRIs)--的主要靶点。在人类中，由一种常见的基因变异导致的SERT表达减少与焦虑相关的人格特征的增加有关。以及对压力相关抑郁的易感性。为了研究SERT降低的影响，我们制造了表达SERT降低的小鼠。在小鼠中，SERT降低会导致类似焦虑的行为增加，与在人类中观察到的类似。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。我们将使用这里开发的优化的UPLC-PFET方法来测量这些动力学。 与公共健康相关：为了了解大脑的内部通信，我们需要绝对有选择性且快速的分析方法。没有一种通用的方法兼具这两种属性。我们将开发一个，并将其应用到关于血清素的问题上--血清素是导致焦虑、抑郁和相关障碍的关键因素。

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.