Department of Cellular Biology and Physiology, Brigham Young University, Provo, Utah 84602, United States.
Department of Neurobiology & Anatomy, Drexel University, Philadelphia, Pennsylvania 28619, United States.
ACS Chem Neurosci. 2022 May 18;13(10):1534-1548. doi: 10.1021/acschemneuro.2c00033. Epub 2022 Apr 28.
Fast-scan cyclic voltammetry (FSCV) is an effective tool for measuring dopamine release and clearance throughout the brain, especially the striatum where dopamine terminals are abundant and signals are heavily regulated by release machinery and the dopamine transporter (DAT). Peak height measurement is perhaps the most common method for measuring dopamine release, but it is influenced by changes in clearance. Michaelis-Menten-based modeling has been a standard in measuring dopamine clearance, but it is problematic in that it requires experimenter fitted modeling subject to experimenter bias. This study presents the use of the first derivative (velocity) of evoked dopamine signals as an alternative approach for measuring and distinguishing dopamine release from clearance. Maximal upward velocity predicts reductions in dopamine peak height due to D and GABA receptor stimulation and by alterations in calcium concentrations. The Michaelis-Menten maximal velocity () measure, an approximation for DAT levels, predicts maximal downward velocity in slices and in vivo. Dopamine peak height and upward velocity were similar between wild-type and DAT knock-out (DATKO) mice. In contrast, downward velocity was lower and exponential decay (tau) was higher in DATKO mice, supporting the use of both measures for extreme changes in DAT activity. In slices, the competitive DAT inhibitors cocaine, PTT, and WF23 increased peak height and upward velocity differentially across increasing concentrations, with PTT and cocaine reducing these measures at high concentrations. Downward velocity and tau values decreased and increased respectively across concentrations, with greater potency and efficacy observed with WF23 and PTT. In vivo recordings demonstrated similar effects of WF23, PTT, and cocaine on measures of release and clearance. Tau was a more sensitive measure at low concentrations, supporting its use as a surrogate for the Michaelis-Menten measure of apparent affinity (). Together, these results inform on the use of these various measures for dopamine release and clearance.
快速扫描循环伏安法（FSCV）是测量整个大脑中多巴胺释放和清除的有效工具，特别是在纹状体中，多巴胺末梢丰富，信号受到释放机制和多巴胺转运体（DAT）的强烈调节。峰高测量可能是测量多巴胺释放最常用的方法，但它受到清除变化的影响。基于米氏方程的建模一直是测量多巴胺清除的标准方法，但它存在问题，因为它需要实验者拟合建模，容易受到实验者偏见的影响。本研究提出了使用诱发多巴胺信号的一阶导数（速度）作为替代方法来测量和区分多巴胺释放和清除。最大向上速度预测由于 D 和 GABA 受体刺激以及钙浓度变化导致的多巴胺峰高降低。米氏方程最大速度（）测量值是 DAT 水平的近似值，可预测切片和体内的最大向下速度。野生型和多巴胺转运体敲除（DATKO）小鼠之间的多巴胺峰高和向上速度相似。相比之下，DATKO 小鼠的向下速度较低，指数衰减（tau）较高，支持这两种方法用于 DAT 活性的极端变化。在切片中，竞争性 DAT 抑制剂可卡因、PTT 和 WF23 以不同的浓度增加峰高和向上速度，PTT 和可卡因在高浓度下降低了这些指标。向下速度和 tau 值分别随浓度降低和升高，WF23 和 PTT 表现出更大的效力和功效。体内记录显示 WF23、PTT 和可卡因对释放和清除的测量有类似的影响。tau 在低浓度下是一种更敏感的指标，支持将其用作米氏方程测量表观亲和力（）的替代指标。这些结果为使用这些各种方法测量多巴胺释放和清除提供了信息。
