scholarly journals 3D ultrastructural study of synapses in the human entorhinal cortex

2020 ◽  
Author(s):  
M Domínguez-Álvaro ◽  
M Montero-Crespo ◽  
L Blazquez-Llorca ◽  
J DeFelipe ◽  
L Alonso-Nanclares
Keyword(s):  
Electron Microscopy ◽  
Human Brain ◽  
Entorhinal Cortex ◽  
Brain Region ◽  
Spatial Navigation ◽  
Ion Beam ◽  
Synaptic Organization ◽  
3D Electron Microscopy ◽  
Quantitative Ultrastructure ◽  
3D Electron

AbstractThe entorhinal cortex (EC) is a brain region that has been shown to be essential for memory functions and spatial navigation. However, detailed 3D synaptic morphology analysis and identification of postsynaptic targets at the ultrastructural level have not been performed before in the human EC. In the present study, we used Focused Ion Beam/Scanning Electron Microscopy (FIB/SEM) to perform a three-dimensional analysis of the synapses in the neuropil of medial EC in layers II and III from human brain autopsies. Specifically, we studied synaptic structural parameters of 3561 synapses, which were fully reconstructed in 3D. We analyzed the synaptic density, 3D spatial distribution, and type (excitatory and inhibitory), as well as the shape and size of each synaptic junction. Moreover, the postsynaptic targets of synapses could be clearly determined. The present work constitutes a detailed description of the synaptic organization of the human EC, which is a necessary step to better understand the functional organization of this region in both health and disease.Significance StatementThe present study represents the first attempt to unveil the detailed synaptic organization of the neuropil of the human entorhinal cortex — a brain region that is essential for memory function and spatial navigation. Using 3D electron microscopy, we have characterized the synaptic morphology and identified the postsynaptic targets of thousands of synapses. The results provide a new, large, quantitative ultrastructure dataset of the synaptic organization of the human entorhinal cortex. These data provide critical information to better understand synaptic functionality in the human brain.HighlightEstimation of the number of synapses, as well as determination of their type, shapes, sizes and postsynaptic targets, provides critical data to better understand synaptic functionality. This study provides a new, large, quantitative ultrastructure dataset of the synaptic organization of the human entorhinal cortex using 3D electron microscopy.

scholarly journals Association of intracellular and synaptic organization in cochlear inner hair cells revealed by 3D electron microscopy

2015 ◽  
Vol 128 (14) ◽  
pp. 2529-2540 ◽  
Author(s):  
Anwen Bullen ◽  
Timothy West ◽  
Carolyn Moores ◽  
Jonathan Ashmore ◽  
Roland A. Fleck ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Hair Cells ◽  
Synaptic Organization ◽  
Inner Hair Cells ◽  
3D Electron Microscopy ◽  
3D Electron

scholarly journals Fluorescence-based 3D targeting of FIB-SEM acquisition of small volumes in large samples

2021 ◽  
Author(s):  
Paolo Ronchi ◽  
Pedro Machado ◽  
Edoardo D’Imprima ◽  
Giulia Mizzon ◽  
Benedikt T. Best ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Focused Ion Beam ◽  
Single Cells ◽  
Ion Beam ◽  
Three Dimensional ◽  
3D Culture ◽  
Fluorescence Signal ◽  
Three Dimensional Objects ◽  
3D Electron Microscopy ◽  
3D Electron

AbstractCells are three dimensional objects. Therefore, 3D electron microscopy is often crucial for correct interpretation of ultrastructural data. Today samples are frequently imaged in 3D at ultrastructural resolution using volume Scanning Electron Microscopy (SEM) methods such as Focused Ion Beam (FIB) SEM and Serial Block face SEM. While these imaging modalities allow for automated data acquisition, precise targeting of (small) volumes of interest within a large sample remains challenging. Here, we provide an easy and reliable approach to target FIB-SEM acquisition of fluorescently labelled cells or subcellular structures with micrometer precision. The strategy relies on fluorescence preservation during sample preparation and targeting based on confocal acquisition of the fluorescence signal in the resin block. Targeted trimming of the block exposes the cell of interest and laser branding of the surface after trimming creates landmarks to precisely position the FIB-SEM acquisition. Using this method, we acquired volumes of specific single cells within large tissues such as a 3D culture of mouse primary mammary gland organoids, tracheal terminal cells in Drosophila melanogaster larvae and ovarian follicular cells in adult Drosophila, discovering ultrastructural details that could not be appreciated before.SummaryRonchi et al. present a workflow to facilitate the precise targeting of three-dimensional (3D) Electron Microscopy acquisitions, guided by fluorescence. This method allows ultrastructural visualization of single cells within a millimeter-range large specimen, based on molecular identity characterized by fluorescence.

scholarly journals Protocol for preparation of heterogeneous biological samples for 3D electron microscopy: a case study for insects

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alexey A. Polilov ◽  
Anastasia A. Makarova ◽  
Song Pang ◽  
C. Shan Xu ◽  
Harald Hess
Keyword(s):  
Electron Microscopy ◽  
Biological Samples ◽  
Focused Ion Beam ◽  
Ion Beam ◽  
Three Dimensional ◽  
Entire Volume ◽  
Simple Protocol ◽  
3D Electron Microscopy ◽  
3D Em

AbstractModern morphological and structural studies are coming to a new level by incorporating the latest methods of three-dimensional electron microscopy (3D-EM). One of the key problems for the wide usage of these methods is posed by difficulties with sample preparation, since the methods work poorly with heterogeneous (consisting of tissues different in structure and in chemical composition) samples and require expensive equipment and usually much time. We have developed a simple protocol allows preparing heterogeneous biological samples suitable for 3D-EM in a laboratory that has a standard supply of equipment and reagents for electron microscopy. This protocol, combined with focused ion-beam scanning electron microscopy, makes it possible to study 3D ultrastructure of complex biological samples, e.g., whole insect heads, over their entire volume at the cellular and subcellular levels. The protocol provides new opportunities for many areas of study, including connectomics.

scholarly journals Patterns of organelle ontogeny through a cell cycle revealed by whole-cell reconstructions using 3D electron microscopy

Development ◽  
2017 ◽  
Vol 144 (4) ◽  
pp. e1.2-e1.2
Author(s):  
Louise Hughes ◽  
Samantha Borrett ◽  
Katie Towers ◽  
Tobias Starborg ◽  
Sue Vaughan
Keyword(s):  
Electron Microscopy ◽  
Cell Cycle ◽  
Whole Cell ◽  
3D Electron Microscopy ◽  
A Cell ◽  
3D Electron

scholarly journals Quantitative method for estimating stain density in electron microscopy of conventionally prepared biological specimens

2019 ◽  
Author(s):  
Andrea Fera ◽  
Qianping He ◽  
Guofeng Zhang ◽  
Richard D. Leapman
Keyword(s):  
Electron Microscopy ◽  
Focused Ion Beam ◽  
Electron Tomography ◽  
Blood Platelets ◽  
Ion Beam ◽  
Simple Expression ◽  
Heavy Atoms ◽  
3D Electron Microscopy ◽  
Block Face ◽  
Microscopy Techniques

SummaryStain density is an important parameter for optimizing the quality of ultrastructural data obtained from several types of 3D electron microscopy techniques, including serial block-face electron microscopy (SBEM), and focused ion beam scanning electron microscopy (FIB-SEM). Here, we show how some straightforward measurements in the TEM can be used to determine the stain density based on a simple expression that we derive. Numbers of stain atoms per unit volume are determined from the measured ratio of the bright-field intensities from regions of the specimen that contain both pure embedding material and the embedded biological structures of interest. The determination only requires knowledge of the section thickness, which can either be estimated from the microtome setting, or from low-dose electron tomography, and the elastic scattering cross section for the heavy atoms used to stain the specimen. The method is tested on specimens of embedded blood platelets, brain tissue, and liver tissue.

Sign in / Sign up

Export Citation Format

Share Document