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. 2018 Jun 18;8:9316. doi: 10.1038/s41598-018-27690-w

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© The Author(s) 2018

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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Neuronal silencing performance of eKR2. (a) Confocal image of a hippocampal neuron in culture (Z stack) with zoom-ins (0.5-µm equatorial slice) on dendrites and soma; scale bar corresponds to 10 µm. (b) Left: Representative voltage traces recorded in current-clamp configuration using a squared somatic current injection protocol to determine the rheobase (540 nm LED, 500 ms) Right: Dependence of average rheobase on illumination intensity (mean ± SD; n = 10). Dotted line represents the average rheobase without illumination. (c) Left: current-clamp traces at different light intensities using a somatic current injection ramp protocol with 0.44 pA/ms (0 to 400 pA in 900 ms). Right: Cell hyperpolarization at 10 mW/mm² green light stimulation (mean ± SD; n = 11). Delta membrane voltage was calculated as the minimum membrane voltage value under light stimulation minus the membrane voltage at 0 somatic current injection and no light stimulus. (d) Relative number of spikes during the ramp protocol at different light intensities normalized to no light stimulation (mean ± SD; n = 11). (e) Pulse trial protocol traces of an eKR2-expressing neuron with (light blue) and without (black) light stimulation; pulse width 10 ms, LJP corrected. (f) Firing probability during pulse trial protocol pre-, during, and post- light stimulation (mean ± SD; n = 11). Left: At AP threshold; middle: injection amplitudes at AP threshold plus 300 pA. Right: At AP threshold plus 500 pA (only eKR2). The Kruskal Wallis test was used followed by Dunn’s test.