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. 2020 Jul 7;11:3394. doi: 10.1038/s41467-020-17110-x

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© The Author(s) 2020

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. 1 — a Registration of VIP^Cre and VPAC2^Cre cells to corresponding phase maps of SCN spatiotemporal bioluminescent wave. For both cell types, left panel shows projections of average bioluminescence emissions recorded by CCD; middle panel shows the corresponding phase map categorically coloured as 1-h phase bins overlaid with manually annotated sequential phase clusters; and right panel shows distribution of Cre-expressing cells revealed by Cre-conditional EYFP intensity. The perimeter of the SCN is delineated by the white outline, and the location of the third ventricle is denoted by asterisks. Note registration of VIP^Cre and VPAC2^Cre cells with earlier and later phase clusters, respectively. These images are representative of a set of three experiments for VIP^EYFP and VPAC2^EYFP each, which are presented in Supplementary Fig. 3. Scale bars = 50 µm. b Representative micrographs of Per2::Luciferase in register with GCaMP6f conditionally targetted to VIP (upper) or VPAC2 (lower) cells. Scale bars = 250 µm. c Representative detrended Per2::Luciferase (purple) and conditionally targetted GCaMP6f traces (green) from recordings from VIP^Cre (upper) and VPAC2^Cre (lower) slices. d Paired period measures of Per2::Luciferase (purple) and conditional GCaMP6f (green) rhythms. e Phase-aligned normalised single cycles of Per2::Luciferase (two overlaid purple) versus corresponding VIP^GCaMP6f (blue) and VPAC2^GCaMP6f (pink) cycles. Lines and shading are mean ± SEM. f Circadian-normalised phase measures for pan-neuronal GCaMP6f (grey), VIP^GCaMP6f (blue) and VPAC2^GCaMP6f (pink), (**p = 0.004, ***p = 0.0005). g Circadian-normalised phase-maps of neuronal GCaMP6f (left), VIP^GCaMP6f (middle) and VPAC2^GCaMP6f (right). SCN outlines from corresponding Per2::Luciferase phase-maps, asterisks mark third ventricle. In all histograms, individual points represent individual SCN slices and bars are mean ± SEM. Statistics: d paired two-tailed t-tests; f ordinary one-way ANOVA with Holm-Sidak’s correction for multiple comparisons. For GCaMP6f imaging (d–f): n = 6, 9 and 10 for neuronal, VIP^GCaMP6f and VPAC2^GCaMP6f. Only significant comparisons (p < 0.05) are shown. Source data are provided as a source data file.