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. 2019 Jul 19;10:3223. doi: 10.1038/s41467-019-11207-8

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© The Author(s) 2019

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. 3 — Orai2-independent replenishment and overcharging of Ry-sensitive Ca²⁺ stores. a Left: Caffeine was locally applied to the somata of whole-cell patch-clamped CA1 pyramidal neurons (PNs) filled with OGB-1 through the patch pipette. Red traces: Fluorescence recordings in an Orai2−/− mouse in response to applications of caffeine before (1), and 30s after (2) a depolarization to 0 mV for 1 s. b Summary of the experiments shown in a (left bars) and c (right bars). Bar graph showing mean amplitudes of relative fluorescence changes in response to caffeine (a, n = 13 cells, p = 0.0014) or 3,5-dihydroxyphenylglycine (DHPG; c, n = 17 cells, p = 0.65) application before (1) and after the depolarization (2), normalized to the control amplitude (1). c Analogous experiment in another CA1 PN from the same mouse with DHPG applied after the depolarization. d Mean area under the curve for 15 s following the caffeine (2) and DHPG (3) applications following the depolarization (marked by dashed boxes in a and b, caffeine: p = 0.01, DHPG: p = 0.831). e Left: Caffeine was locally applied to somata of CA1 PNs filled with Cal520-AM by multi-cell bolus loading (MCBL). Traces: Fluorescence recordings from the soma of a single CA1 PN in a wild-type mouse during consecutive caffeine applications (arrowheads) in the presence of D600 (blue bar) and after washout of D600. f Summary of the experiments shown in e. Mean amplitudes of relative fluorescence changes in response to caffeine applications at the time points indicated in e (n = 41 cells, analysis of variance (ANOVA): p = 0.001 (1 vs 2) and p = 4.98 × 10⁻⁶ (2 vs 3). g Analogous experiment to e with DHPG instead of caffeine applications. h Analogous summary to f for the experiments shown in g (n = 41 cells, ANOVA: p = 0.694 (1 vs 2) and p = 0.058 (2 vs 3)). ** - very significant (p <= 0.01), *** - extremely significant (p <= 0.0001)