---
_id: '11393'
abstract:
- lang: eng
text: "AMPA receptors (AMPARs) mediate fast excitatory neurotransmission and their
role is\r\nimplicated in complex processes such as learning and memory and various
neurological\r\ndiseases. These receptors are composed of different subunits and
the subunit composition can\r\naffect channel properties, receptor trafficking
and interaction with other associated proteins.\r\nUsing the high sensitivity
SDS-digested freeze-fracture replica labeling (SDS-FRL) for\r\nelectron microscopy
I investigated the number, density, and localization of AMPAR subunits,\r\nGluA1,
GluA2, GluA3, and GluA1-3 (panAMPA) in pyramidal cells in the CA1 area of mouse\r\nhippocampus.
I have found that the immunogold labeling for all of these subunits in the\r\npostsynaptic
sites was highest in stratum radiatum and lowest in stratum lacunosummoleculare.
The labeling density for the all subunits in the extrasynaptic sites showed a
gradual\r\nincrease from the pyramidal cell soma towards the distal part of stratum
radiatum. The densities\r\nof extrasynaptic GluA1, GluA2 and panAMPA labeling
reached 10-15% of synaptic densities,\r\nwhile the ratio of extrasynaptic labeling
for GluA3 was significantly lower compared than those\r\nfor other subunits. The
labeling patterns for GluA1, GluA2 and GluA1-3 are similar and their\r\ndensities
were higher in the periphery than center of synapses. In contrast, the GluA3-\r\ncontaining
receptors were more centrally localized compared to the GluA1- and GluA2-\r\ncontaining
receptors.\r\nThe hippocampus plays a central role in learning and memory. Contextual
learning has been\r\nshown to require the delivery of AMPA receptors to CA1 synapses
in the dorsal hippocampus.\r\nHowever, proximodistal heterogeneity of this plasticity
and particular contribution of different\r\nAMPA receptor subunits are not fully
understood. By combining inhibitory avoidance task, a\r\nhippocampus-dependent
contextual fear-learning paradigm, with SDS-FRL, I have revealed an\r\nincrease
in synaptic density specific to GluA1-containing AMPA receptors in the CA1 area.\r\nThe
intrasynaptic distribution of GluA1 also changed from the periphery to center-preferred\r\npattern.
Furthermore, this synaptic plasticity was evident selectively in stratum radiatum
but\r\nnot stratum oriens, and in the CA1 subregion proximal but not distal to
CA2. These findings\r\nfurther contribute to our understanding of how specific
hippocampal subregions and AMPA\r\nreceptor subunits are involved in physiological
learning.\r\nAlthough the immunolabeling results above shed light on subunit-specific
plasticity in\r\nAMPAR distribution, no tools to visualize and study the subunit
composition at the single\r\nchannel level in situ have been available. Electron
microscopy with conventional immunogold\r\nlabeling approaches has limitations
in the single channel analysis because of the large size of\r\nantibodies and
steric hindrance hampering multiple subunit labeling of single channels. I\r\nmanaged
to develop a new chemical labeling system using a short peptide tag and small\r\nsynthetic
probes, which form specific covalent bond with a cysteine residue in the tag fused
to\r\nproteins of interest (reactive tag system). I additionally made substantial
progress into adapting\r\nthis system for AMPA receptor subunits."
acknowledged_ssus:
- _id: EM-Fac
alternative_title:
- ISTA Thesis
article_processing_charge: No
author:
- first_name: Marijo
full_name: Jevtic, Marijo
id: 4BE3BC94-F248-11E8-B48F-1D18A9856A87
last_name: Jevtic
citation:
ama: Jevtic M. Contextual fear learning induced changes in AMPA receptor subtypes
along the proximodistal axis in dorsal hippocampus. 2022. doi:10.15479/at:ista:11393
apa: Jevtic, M. (2022). Contextual fear learning induced changes in AMPA receptor
subtypes along the proximodistal axis in dorsal hippocampus. Institute of
Science and Technology Austria. https://doi.org/10.15479/at:ista:11393
chicago: Jevtic, Marijo. “Contextual Fear Learning Induced Changes in AMPA Receptor
Subtypes along the Proximodistal Axis in Dorsal Hippocampus.” Institute of Science
and Technology Austria, 2022. https://doi.org/10.15479/at:ista:11393.
ieee: M. Jevtic, “Contextual fear learning induced changes in AMPA receptor subtypes
along the proximodistal axis in dorsal hippocampus,” Institute of Science and
Technology Austria, 2022.
ista: Jevtic M. 2022. Contextual fear learning induced changes in AMPA receptor
subtypes along the proximodistal axis in dorsal hippocampus. Institute of Science
and Technology Austria.
mla: Jevtic, Marijo. Contextual Fear Learning Induced Changes in AMPA Receptor
Subtypes along the Proximodistal Axis in Dorsal Hippocampus. Institute of
Science and Technology Austria, 2022, doi:10.15479/at:ista:11393.
short: M. Jevtic, Contextual Fear Learning Induced Changes in AMPA Receptor Subtypes
along the Proximodistal Axis in Dorsal Hippocampus, Institute of Science and Technology
Austria, 2022.
date_created: 2022-05-17T08:57:41Z
date_published: 2022-05-16T00:00:00Z
date_updated: 2023-09-07T14:53:44Z
day: '16'
ddc:
- '570'
degree_awarded: PhD
department:
- _id: GradSch
- _id: RySh
doi: 10.15479/at:ista:11393
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- iso: eng
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page: '108'
publication_identifier:
issn:
- 2663-337X
publication_status: published
publisher: Institute of Science and Technology Austria
related_material:
record:
- id: '7391'
relation: part_of_dissertation
status: public
status: public
supervisor:
- first_name: Ryuichi
full_name: Shigemoto, Ryuichi
id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
last_name: Shigemoto
orcid: 0000-0001-8761-9444
title: Contextual fear learning induced changes in AMPA receptor subtypes along the
proximodistal axis in dorsal hippocampus
type: dissertation
user_id: 8b945eb4-e2f2-11eb-945a-df72226e66a9
year: '2022'
...
---
_id: '7391'
abstract:
- lang: eng
text: Electron microscopy (EM) is a technology that enables visualization of single
proteins at a nanometer resolution. However, current protein analysis by EM mainly
relies on immunolabeling with gold-particle-conjugated antibodies, which is compromised
by large size of antibody, precluding precise detection of protein location in
biological samples. Here, we develop a specific chemical labeling method for EM
detection of proteins at single-molecular level. Rational design of α-helical
peptide tag and probe structure provided a complementary reaction pair that enabled
specific cysteine conjugation of the tag. The developed chemical labeling with
gold-nanoparticle-conjugated probe showed significantly higher labeling efficiency
and detectability of high-density clusters of tag-fused G protein-coupled receptors
in freeze-fracture replicas compared with immunogold labeling. Furthermore, in
ultrathin sections, the spatial resolution of the chemical labeling was significantly
higher than that of antibody-mediated labeling. These results demonstrate substantial
advantages of the chemical labeling approach for single protein visualization
by EM.
article_processing_charge: No
article_type: original
author:
- first_name: Shigekazu
full_name: Tabata, Shigekazu
id: 4427179E-F248-11E8-B48F-1D18A9856A87
last_name: Tabata
- first_name: Marijo
full_name: Jevtic, Marijo
id: 4BE3BC94-F248-11E8-B48F-1D18A9856A87
last_name: Jevtic
- first_name: Nobutaka
full_name: Kurashige, Nobutaka
last_name: Kurashige
- first_name: Hirokazu
full_name: Fuchida, Hirokazu
last_name: Fuchida
- first_name: Munetsugu
full_name: Kido, Munetsugu
last_name: Kido
- first_name: Kazushi
full_name: Tani, Kazushi
last_name: Tani
- first_name: Naoki
full_name: Zenmyo, Naoki
last_name: Zenmyo
- first_name: Shohei
full_name: Uchinomiya, Shohei
last_name: Uchinomiya
- first_name: Harumi
full_name: Harada, Harumi
id: 2E55CDF2-F248-11E8-B48F-1D18A9856A87
last_name: Harada
orcid: 0000-0001-7429-7896
- first_name: Makoto
full_name: Itakura, Makoto
last_name: Itakura
- first_name: Itaru
full_name: Hamachi, Itaru
last_name: Hamachi
- first_name: Ryuichi
full_name: Shigemoto, Ryuichi
id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
last_name: Shigemoto
orcid: 0000-0001-8761-9444
- first_name: Akio
full_name: Ojida, Akio
last_name: Ojida
citation:
ama: Tabata S, Jevtic M, Kurashige N, et al. Electron microscopic detection of single
membrane proteins by a specific chemical labeling. iScience. 2019;22(12):256-268.
doi:10.1016/j.isci.2019.11.025
apa: Tabata, S., Jevtic, M., Kurashige, N., Fuchida, H., Kido, M., Tani, K., … Ojida,
A. (2019). Electron microscopic detection of single membrane proteins by a specific
chemical labeling. IScience. Elsevier. https://doi.org/10.1016/j.isci.2019.11.025
chicago: Tabata, Shigekazu, Marijo Jevtic, Nobutaka Kurashige, Hirokazu Fuchida,
Munetsugu Kido, Kazushi Tani, Naoki Zenmyo, et al. “Electron Microscopic Detection
of Single Membrane Proteins by a Specific Chemical Labeling.” IScience.
Elsevier, 2019. https://doi.org/10.1016/j.isci.2019.11.025.
ieee: S. Tabata et al., “Electron microscopic detection of single membrane
proteins by a specific chemical labeling,” iScience, vol. 22, no. 12. Elsevier,
pp. 256–268, 2019.
ista: Tabata S, Jevtic M, Kurashige N, Fuchida H, Kido M, Tani K, Zenmyo N, Uchinomiya
S, Harada H, Itakura M, Hamachi I, Shigemoto R, Ojida A. 2019. Electron microscopic
detection of single membrane proteins by a specific chemical labeling. iScience.
22(12), 256–268.
mla: Tabata, Shigekazu, et al. “Electron Microscopic Detection of Single Membrane
Proteins by a Specific Chemical Labeling.” IScience, vol. 22, no. 12, Elsevier,
2019, pp. 256–68, doi:10.1016/j.isci.2019.11.025.
short: S. Tabata, M. Jevtic, N. Kurashige, H. Fuchida, M. Kido, K. Tani, N. Zenmyo,
S. Uchinomiya, H. Harada, M. Itakura, I. Hamachi, R. Shigemoto, A. Ojida, IScience
22 (2019) 256–268.
date_created: 2020-01-29T15:56:56Z
date_published: 2019-12-20T00:00:00Z
date_updated: 2024-03-28T23:30:12Z
day: '20'
ddc:
- '570'
department:
- _id: RySh
doi: 10.1016/j.isci.2019.11.025
ec_funded: 1
external_id:
isi:
- :000504652000020
pmid:
- '31786521'
file:
- access_level: open_access
checksum: f3e90056a49f09b205b1c4f8c739ffd1
content_type: application/pdf
creator: dernst
date_created: 2020-02-04T10:48:36Z
date_updated: 2020-07-14T12:47:57Z
file_id: '7448'
file_name: 2019_iScience_Tabata.pdf
file_size: 7197776
relation: main_file
file_date_updated: 2020-07-14T12:47:57Z
has_accepted_license: '1'
intvolume: ' 22'
issue: '12'
language:
- iso: eng
license: https://creativecommons.org/licenses/by/4.0/
month: '12'
oa: 1
oa_version: Published Version
page: 256-268
pmid: 1
project:
- _id: 25CA28EA-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '694539'
name: 'In situ analysis of single channel subunit composition in neurons: physiological
implication in synaptic plasticity and behaviour'
- _id: 25CBA828-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '720270'
name: Human Brain Project Specific Grant Agreement 1 (HBP SGA 1)
publication: iScience
publication_identifier:
issn:
- 2589-0042
publication_status: published
publisher: Elsevier
quality_controlled: '1'
related_material:
record:
- id: '11393'
relation: dissertation_contains
status: public
scopus_import: '1'
status: public
title: Electron microscopic detection of single membrane proteins by a specific chemical
labeling
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1
volume: 22
year: '2019'
...