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. 2023 Mar 23;26(4):682–695. doi: 10.1038/s41593-023-01281-z

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — a, Top left: two-photon image of axons from an SS31232-Gal4 animal expressing OpGCaMP6f (cyan) and tdTomato (red). ROIs are numbered. Scale bar, 5 μm. Bottom: behavioral epochs are color-coded. Representative ΔF/F time series from two ROIs (green) overlaid with a prediction (black) obtained by convolving PE epochs with a Ca²⁺ indicator response function. Explained variance is indicated (R²). b, ΔF/F, normalized with respect to the neuron’s 90th percentile, as a function of PE number within a PE train for ROIs 0 (solid boxes, filled circles) or 1 (dashed boxes, open circles). Data include 25 PE trains from eight animals and are presented as IQR (box), median (center), 1.5× IQR (whisker) and outliers (circles). c, Explained variance (R²) between ΔF/F time series and a prediction obtained by convolving PE epochs with a Ca²⁺ indicator response function and a time window. Time windows that maximize the correlation for ROIs 0 (solid line) and 1 (dashed line) are indicated (red circles). d, Behavioral epochs are color-coded. Representative ΔF/F time series from two ROIs (green) are overlaid with a prediction (black) obtained by convolving PE epochs with a Ca²⁺ response function as well as the time windows indicated in c (red circles). Explained variance is indicated (R²). e, Standard deviation projection image of a SS31232-Gal4 nervous system expressing smFP and stained for GFP (green) and Nc82 (blue). Cell bodies are indicated (white asterisks). Scale bar, 40 μm. f, Projection as in e but for one MCFO-expressing, traced neuron (black asterisks). The brain’s GNG (yellow) and the VNC’s intermediate tectulum (green) and prothoracic leg neuromere (yellow) are color-coded. Scale bar, 40 μm. g,h, Higher magnification projections of brains (top) and VNCs (bottom) for SS31232-Gal4 animals expressing the stochastic label MCFO (g) or the synaptic marker, syt:GFP (green) and tdTomato (red) (h). Insets magnify dashed boxes. Indicated are cell bodies (asterisks) and bouton-like structures (white arrowheads). Scale bars for brain images are 10 μm. Scale bars for VNC images and insets are 20 μm and 2 μm, respectively.

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