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. 2020 Mar 9;10:90. doi: 10.1038/s41398-020-0767-0

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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. 2 — Live cell calcium recordings in DVC brain slices in astrocytes and neurons simultaneously. a Cells prelabeled with Cal520 (calcium sensitive dye; taken up by both neurons and astrocytes) encircled in green. b Cells prelabeled with SR101 (astrocyte-specific dye) are identified as astrocytes and encircled in red. c Yellow localization confirms that the calcium-sensitive dye is localized within astrocytes and distinguishable from neurons (green circle). d Total field of view (showing both neurons and astrocytes) at “rest” (i.e., baseline) and the same field of view shown in e at period of peak response leptin perfusion (see Supplementary Video 2 and 3). f Response to leptin perfusion of individual astrocyte marked by red arrow in g Response to leptin perfusion of individual neuron marked by green arrow in panel. Changes in fluorescence were measured in brain slices prepared from lean rats (n = 4) and rats exposed to a high-fat diet (n = 3) following bath leptin exposure (100ng/mL). h Magnitude of the leptin response was attenuated in neurons but not astrocytes of obese compared to lean animals. Obese animals had lower proportions of leptin-responsive (i) neurons and (j) astrocytes compared to lean animals. Changes in magnitude are represented as the mean ± standard error of the mean and were analyzed with a 2-way ANOVA and Bonferroni post hoc tests; *p < 0.05. Percentages of leptin-responsive cells were analyzed using Fisher’s exact test; *p < 0.05. See Supplementary Video 2 and Supplementary Video 3.