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. 2019 Feb 8;9:1693. doi: 10.1038/s41598-018-38412-7

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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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3D Serial Block-Face scanning electron microscopy of dendritic spines and core compartment of postsynaptic densities (PSD-core) in the organotypic hippocampal slices (OHCs). (a) Experimental design: OHC were treated with cLTP (n = 4) or DMSO (control, n = 4) for 30 min and then fixed. (a) OHC transfected with AAV1/2_CaMKII-mCherry to show its morphology. Scale bar, 200 μm. For 3D EM, only non-transfected OHC were used. (c) Low-magnification (5k) electron microphotograph of CA1 region of the organotypic slices showing conserved morphology. Dashed lines denote borders between strata. Scale bar, 10 μm. (d) 3D reconstruction of a full SBEM scan. Scale bar, 1 μm. (e,f) High magnification (25k) images of neuropil; M – mitochondrion, Sp – dendritic spine, T - axon terminal. Scale bars, 1 μm. (g,h) Consecutive images from SBEM scans with examples of dendritic spines from (g) control and (h) cLTP sample. Example dendritic spines are marked in blue and the PSDs are marked in red. Scale bars, 1 μm. (i) Example of a PSD segmentation (from a scan). Arrows indicate two parts of a perforated PSD. PSD-core volume was measured by outlining the electron-dense PSD core. PSD area was annotated with open traces along PSD opposing active zone (see methods for details).