Wallace Conner W, DiMarco Emily K, Slinkard Clare Y, Shaughnessy Rachael, Holleran Katherine M, Centanni Samuel W, Lapish Christopher C, Jones Sara R
Department of Translational Neuroscience, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, United States.
Eastern Virginia Medical School, Old Dominion University, Norfolk, Virginia 23501, United States.
ACS Chem Neurosci. 2025 Sep 3;16(17):3239-3256. doi: 10.1021/acschemneuro.5c00078. Epub 2025 Aug 13.
Fluorescent sensors have revolutionized the measurement of molecules in the brain, and the dLight dopamine sensor has been used extensively to examine reward- and cue-evoked dopamine release, but only recently has the field turned its attention to spontaneous release events. Analysis of spontaneous events typically requires evaluation of hundreds of events over minutes to hours, and the most common method of analysis, -scoring, was not designed for this purpose. Here, we compare the accuracy and reliability of three different analysis methods to identify pharmacologically induced changes in dopamine release and uptake in the nucleus accumbens core of freely moving C57BL/6J mice. The D1-like receptor antagonist SCH23390 was used to prevent dLight sensors from interacting with dopamine in the extracellular space, while cocaine was used to inhibit uptake and raclopride to increase the release of dopamine. We examined peak-to-peak frequency, peak amplitude, and width, the time spent above an established cutoff. The three methods were (1) the "-score method", which automatically smooths baseline drift and normalizes recordings using signal-to-noise ratios; (2) a "manual method", in which local baselines were adjusted manually and individual cutoffs were determined for each subject; and (3) the "prominence method" that combines -scoring with prominence assessment to tag individual peaks and then returns to the preprocessed data for kinetic analysis. First, SCH23390 reduced the amplitude of release, but this effect was diminished using the Z-score method, and there was a variable effect on frequency between methods. Cocaine increased signal width as expected using the manual and prominence methods but not the Z-score method. Finally, raclopride-induced increases in amplitude were correctly identified by all three methods, but this effect was again diminished by using the Z-score method. Thus, analysis of spontaneous dopamine signals requires assessment of the %Δ/ values, and the use of -scoring is not appropriate.
荧光传感器彻底改变了大脑中分子的测量方式，dLight多巴胺传感器已被广泛用于检测奖励和线索诱发的多巴胺释放，但该领域直到最近才将注意力转向自发释放事件。对自发事件的分析通常需要在数分钟到数小时内评估数百个事件，而最常用的分析方法——Z评分，并非为此目的而设计。在这里，我们比较了三种不同分析方法在识别自由活动的C57BL/6J小鼠伏隔核核心中多巴胺释放和摄取的药理学诱导变化方面的准确性和可靠性。D1样受体拮抗剂SCH23390用于防止dLight传感器与细胞外空间中的多巴胺相互作用，而可卡因用于抑制摄取，雷氯必利用于增加多巴胺的释放。我们检查了峰峰值频率、峰值幅度和宽度，以及高于既定阈值的时间。这三种方法分别是：（1）“Z评分法”，该方法使用信噪比自动平滑基线漂移并对记录进行归一化；（2）“手动法”，即手动调整局部基线并为每个受试者确定单独的阈值；（3）“突出度法”，该方法将Z评分与突出度评估相结合以标记单个峰值，然后返回预处理数据进行动力学分析。首先，SCH23390降低了释放幅度，但使用Z评分法时这种效果减弱，并且不同方法之间对频率的影响存在差异。可卡因使用手动法和突出度法时如预期增加了信号宽度，但Z评分法未显示此效果。最后，所有三种方法都正确识别出雷氯必利诱导的幅度增加，但使用Z评分法时这种效果再次减弱。因此，对自发多巴胺信号的分析需要评估%Δ/值，使用Z评分并不合适。
