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- EMDB-53931: Memory engram synapse 3D molecular architecture visualized by cry... -

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Basic information

Entry
Database: EMDB / ID: EMD-53931
TitleMemory engram synapse 3D molecular architecture visualized by cryoCLEM-guided cryoET
Map dataRepresentative tomographic volume of engram labelled synapse
Sample
  • Tissue: Macromolecular resolution of synapses between engram cells within tissue
KeywordsSynapse / Engram / In Situ / In Tissue / CEMOVIS / cryo-section / CELL ADHESION
Biological speciesMus musculus (house mouse)
Methodelectron tomography
AuthorsFrank RAW / Lovatt CA
Funding support United Kingdom, European Union, Ireland, 7 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/M011151/1 United Kingdom
European Research Council (ERC)715968European Union
Science Foundation Ireland15/YI/3187 Ireland
Irish Research CouncilGOID/2019-812 Ireland
UK Research and Innovation (UKRI)MR/V022644/1 United Kingdom
Wellcome Trust108466/Z/15/Z & 221524/Z/20/Z United Kingdom
Wellcome Trust208395/Z/17/Z United Kingdom
CitationJournal: bioRxiv / Year: 2025
Title: Memory engram synapse 3D molecular architecture visualized by cryoCLEM-guided cryoET.
Authors: Charlie Lovatt / Thomas J O'Sullivan / Clara Ortega-de San Luis / Tomás J Ryan / René A W Frank /
Abstract: Memory is incorporated into the brain as physicochemical changes to engram cells. These are neuronal populations that form complex neuroanatomical circuits, are modified by experiences to store ...Memory is incorporated into the brain as physicochemical changes to engram cells. These are neuronal populations that form complex neuroanatomical circuits, are modified by experiences to store information, and allow for memory recall. At the molecular level, learning modifies synaptic communication to rewire engram circuits, a mechanism known as synaptic plasticity. However, despite its functional role on memory formation, the 3D molecular architecture of synapses within engram circuits is unknown. Here, we demonstrate the use of engram labelling technology and cryogenic correlated light and electron microscopy (cryoCLEM)-guided cryogenic electron tomography (cryoET) to visualize the in-tissue 3D molecular architecture of engram synapses of a contextual fear memory within the CA1 region of the mouse hippocampus. Engram cells exhibited structural diversity of macromolecular constituents and organelles in both pre- and postsynaptic compartments and within the synaptic cleft, including in clusters of membrane proteins, synaptic vesicle occupancy, and F-actin copy number. This 'engram to tomogram' approach, harnessing functional neuroscience and structural biology, provides a methodological framework for testing fundamental molecular plasticity mechanisms within engram circuits during memory encoding, storage and recall.
History
DepositionJun 2, 2025-
Header (metadata) releaseNov 26, 2025-
Map releaseNov 26, 2025-
UpdateNov 26, 2025-
Current statusNov 26, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_53931.png

Downloads & links

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Map

FileDownload / File: emd_53931.map.gz / Format: CCP4 / Size: 1.6 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRepresentative tomographic volume of engram labelled synapse
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
12 Å/pix.
x 400 pix.
= 4800. Å		12 Å/pix.
x 1024 pix.
= 12288. Å		12 Å/pix.
x 1024 pix.
= 12288. Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 12 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-82.238174000000001 - 12.668004
Average (Standard dev.)-6.706597 (±3.284889)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions10241024400
Spacing10241024400
CellA: 12288.0 Å / B: 12288.0 Å / C: 4800.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Macromolecular resolution of synapses between engram cells within...

EntireName: Macromolecular resolution of synapses between engram cells within tissue
Components
  • Tissue: Macromolecular resolution of synapses between engram cells within tissue

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Supramolecule #1: Macromolecular resolution of synapses between engram cells within...

SupramoleculeName: Macromolecular resolution of synapses between engram cells within tissue
type: tissue / ID: 1 / Parent: 0 / Details: cryoCLEM-targeted engram-labelled synapses
Source (natural)Organism: Mus musculus (house mouse) / Strain: C57B16/J / Organ: Brain / Tissue: Brain

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Experimental details

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Structure determination

Processingelectron tomography
Aggregation statetissue

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Sample preparation

BufferpH: 7.3
Component:
ConcentrationFormulaName
23.25 mMNMDGn-methyl-d-glucamine
0.625 mMKClPotassium chloride
0.3 mMNaH2PO4Socium Phosphate
7.5 mMNaHCO3Sodium carbonate
5.0 mMHEPESHEPES
6.25 mMC6H12O6Glucose
1.25 mMC6H7O6NaAscorbic acid
0.5 mMCH4N2SThiourea
0.75 mMC3H3NaO3Sodium Pyruvate
2.5 mMMgSO4.7H2OMagnesium Sulphate
0.125 mMCaCl2.2H2OCalcium chloride

Details: pH 7.2-7.4, 304-310 mOsm
GridModel: Quantifoil R3.5/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
DetailsHigh pressure frozen brain tissue with 20% dextran cryoprotectant
High pressure freezingInstrument: OTHER
Details: 3 mm diameter, 100 um thick carriers. 20% dextran cryoprotectant.. The value given for _em_high_pressure_freezing.instrument is Leica EM ICE. This is not in a list of allowed values {'LEICA ...Details: 3 mm diameter, 100 um thick carriers. 20% dextran cryoprotectant.. The value given for _em_high_pressure_freezing.instrument is Leica EM ICE. This is not in a list of allowed values {'LEICA EM PACT2', 'BAL-TEC HPM 010', 'LEICA EM HPM100', 'OTHER', 'EMS-002 RAPID IMMERSION FREEZER', 'LEICA EM PACT'} so OTHER is written into the XML file.
Cryo protectant20% dextran
SectioningUltramicrotomy - Instrument: Leica EM UC7 / Ultramicrotomy - Temperature: 113 K / Ultramicrotomy - Final thickness: 100
Ultramicrotomy - Details: High pressure frozen tissue carriers were loaded into Leica UC7 cryo-ultramicrotome and tissue ribbons were collected at 100-190 nm thickness.

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 3 pixel / Digitization - Dimensions - Height: 3 pixel / Number grids imaged: 4 / Number real images: 2867 / Average exposure time: 2.0 sec. / Average electron dose: 109.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated defocus max: 8.0 µm / Calibrated defocus min: 5.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 8.0 µm / Nominal defocus min: 5.0 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsTilt series were reconstructed in imod and tomograms were denoised in isonet
Final reconstructionAlgorithm: SIMULTANEOUS ITERATIVE (SIRT) / Software: (Name: IMOD (ver. 4.9), UCSF ChimeraX (ver. 1.8)) / Number images used: 2396
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION

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