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. 2023 May 22;14:2939. doi: 10.1038/s41467-023-38583-6

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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 Left: strategy for labeling and photoactivating RTN^Phox2b/Atoh1 neurons unilaterally. Right: transfected somas. Scale bar, 250 µm. Representative of n = 6 mice. b Distribution of transfected neurons across pF subregions defined in Fig. 4f. Bars are means ± SD across n mice and circles are mean values of individual mice. c Transverse section at the preBötC level showing RTN^Phox2b/Atoh1 projections. Scale bar, 500 µm (left), 100 µm (right). d Distribution of RTN^Phox2b/Atoh1 projections in the preBötC. Bars are means ± SD across n mice and gray circles are mean values of individual mice. **p = 0.0039, Wilcoxon matched-pairs test. e Top: whole-body plethysmography recording (WBP, inspirations are upwards) around a single photoactivation of RTN^Phox2b/Atoh1 neurons which shortens the respiratory cycle. Φ: control cycle, φ_s: phase of photoactivation, θ: perturbed cycle. f Inspiratory phase-shift as a function of the phase of photoactivation showing a shortening. Blue circles are N random trials from n mice, white circles are averages ± SD across all trials within 0.1 ms bins. See Fig. 2 and methods for details. g WBP recordings during RTN^Phox2b/Atoh1 photoactivation at increasing frequencies. h Respiratory parameters before (CTL), during and after (OFF) photoactivations. Gray circles are means of individual animals and colored circles are means ± SD across n mice. In all graphs, §, p < 0.05; ** or §§, p < 0.01, §§§, p < 0.001; §§§§, p < 0.0001 using Wilcoxon matched-pairs test (amplitude) and paired t tests (frequency, Ti, Te). Exact p values between conditions (*) are, from left to right: amplitude: p = 0,25; p = 0,9375; p = 0,3125; p = 0,125; frequency: p = 0,125; p = 0,1215; p = 0,3527; p = 0,0066; Ti: p = 0,684; p = 0,0025; p = 0,2645; p = 0,0523; Te: p = 0,071; p = 0,0069; p = 0,1537; p = 0,0014. Additional statistics compared to CTL condition (§): amplitude: CTL vs 5 Hz, p = 0,125; CTL vs 10 Hz, p = 0,2188; CTL vs 20 Hz, p = 0,4688; CTL vs 40 Hz, p = 0,125; CTL vs OFF, p = 0,8125; frequency: CTL vs 5 Hz, p = 0,0204; CTL vs 10 Hz, p < 0,0001; CTL vs 20 Hz, p = 0,0001; CTL vs 40 Hz, p = 0,0017; CTL vs OFF, p = 0,3061; Ti: CTL vs 5 Hz, p = 0,0651; CTL vs 10 Hz, p = 0,042; CTL vs 20 Hz, p = 0,0031; CTL vs 40 Hz, p = 0,0903; CTL vs OFF, p = 0,7401; Te: CTL vs 5 Hz, p = 0,0082; CTL vs 10 Hz, p < 0,0001; CTL vs 20 Hz, p < 0,0001; CTL vs 40 Hz, p = 0,0003; CTL vs OFF, p = 0,6141. i Left: strategy for labeling RTN^Phox2b/Atoh1 synaptic contacts unilaterally. Middle: transverse section showing transfected somas in the pF. Scale bar, 250 µm. Right: magnification. Scale bar, 50 µm. j Transverse section at the preBötC level showing RTN^Phox2b/Atoh1 synaptic contacts. Scale bar, 1 mm (left), 100 µm (right). i, j are representative of n = 4 animals. For all graphs, source data are provided as a Source data file. See also Fig. S12.