---
_id: '15084'
abstract:
- lang: eng
text: "GABAB receptor (GBR) activation inhibits neurotransmitter release in axon
terminals in the brain, except in medial habenula (MHb) terminals, which show
robust potentiation. However, mechanisms underlying this enigmatic potentiation
remain elusive. Here, we report that GBR activation on MHb terminals induces an
activity-dependent transition from a facilitating, tonic to a depressing, phasic
neurotransmitter release mode. This transition is accompanied by a 4.1-fold increase
in readily releasable vesicle pool (RRP) size and a 3.5-fold increase of docked
synaptic vesicles (SVs) at the presynaptic active zone (AZ). Strikingly, the depressing
phasic release exhibits looser coupling distance than the tonic release. Furthermore,
the tonic and phasic release are selectively affected by deletion of synaptoporin
(SPO) and Ca\r\n 2+\r\n -dependent
activator protein for secretion 2 (CAPS2), respectively. SPO modulates augmentation,
the short-term plasticity associated with tonic release, and CAPS2 retains the
increased RRP for initial responses in phasic response trains. The cytosolic protein
CAPS2 showed a SV-associated distribution similar to the vesicular transmembrane
protein SPO, and they were colocalized in the same terminals. We developed the
“Flash and Freeze-fracture” method, and revealed the release of SPO-associated
vesicles in both tonic and phasic modes and activity-dependent recruitment of
CAPS2 to the AZ during phasic release, which lasted several minutes. Overall,
these results indicate that GBR activation translocates CAPS2 to the AZ along
with the fusion of CAPS2-associated SVs, contributing to persistency of the RRP
increase. Thus, we identified structural and molecular mechanisms underlying tonic
and phasic neurotransmitter release and their transition by GBR activation in
MHb terminals."
acknowledged_ssus:
- _id: M-Shop
- _id: PreCl
- _id: EM-Fac
acknowledgement: We thank Erwin Neher and Ipe Ninan for critical comments on the manuscript.
This project has received funding from the European Research Council (ERC) and European
Commission, under the European Union’s Horizon 2020 research and innovation program
(ERC grant agreement no. 694539 to R.S. and the Marie Skłodowska-Curie grant agreement
no. 665385 to C.Ö.). This study was supported by the Cooperative Study Program of
Center for Animal Resources and Collaborative Study of NINS. We thank Kohgaku Eguchi
for statistical analysis, Yu Kasugai for additional EM imaging, Robert Beattie for
the design of the slice recovery chamber for Flash and Freeze experiments, Todor
Asenov from the ISTA machine shop for custom part preparations for high-pressure
freezing, the ISTA preclinical facility for animal caretaking, and the ISTA EM facilities
for technical support.
article_number: e2301449121
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: Peter
full_name: Koppensteiner, Peter
id: 3B8B25A8-F248-11E8-B48F-1D18A9856A87
last_name: Koppensteiner
orcid: 0000-0002-3509-1948
- first_name: Pradeep
full_name: Bhandari, Pradeep
id: 45EDD1BC-F248-11E8-B48F-1D18A9856A87
last_name: Bhandari
orcid: 0000-0003-0863-4481
- first_name: Hüseyin C
full_name: Önal, Hüseyin C
id: 4659D740-F248-11E8-B48F-1D18A9856A87
last_name: Önal
orcid: 0000-0002-2771-2011
- first_name: Carolina
full_name: Borges Merjane, Carolina
id: 4305C450-F248-11E8-B48F-1D18A9856A87
last_name: Borges Merjane
orcid: 0000-0003-0005-401X
- first_name: Elodie
full_name: Le Monnier, Elodie
id: 3B59276A-F248-11E8-B48F-1D18A9856A87
last_name: Le Monnier
- first_name: Utsa
full_name: Roy, Utsa
id: 4d26cf11-5355-11ee-ae5a-eb05e255b9b2
last_name: Roy
- first_name: Yukihiro
full_name: Nakamura, Yukihiro
last_name: Nakamura
- first_name: Tetsushi
full_name: Sadakata, Tetsushi
last_name: Sadakata
- first_name: Makoto
full_name: Sanbo, Makoto
last_name: Sanbo
- first_name: Masumi
full_name: Hirabayashi, Masumi
last_name: Hirabayashi
- first_name: JeongSeop
full_name: Rhee, JeongSeop
last_name: Rhee
- first_name: Nils
full_name: Brose, Nils
last_name: Brose
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
- first_name: Ryuichi
full_name: Shigemoto, Ryuichi
id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
last_name: Shigemoto
orcid: 0000-0001-8761-9444
citation:
ama: Koppensteiner P, Bhandari P, Önal C, et al. GABAB receptors induce phasic release
from medial habenula terminals through activity-dependent recruitment of release-ready
vesicles. Proceedings of the National Academy of Sciences. 2024;121(8).
doi:10.1073/pnas.2301449121
apa: Koppensteiner, P., Bhandari, P., Önal, C., Borges Merjane, C., Le Monnier,
E., Roy, U., … Shigemoto, R. (2024). GABAB receptors induce phasic release from
medial habenula terminals through activity-dependent recruitment of release-ready
vesicles. Proceedings of the National Academy of Sciences. Proceedings
of the National Academy of Sciences. https://doi.org/10.1073/pnas.2301449121
chicago: Koppensteiner, Peter, Pradeep Bhandari, Cihan Önal, Carolina Borges Merjane,
Elodie Le Monnier, Utsa Roy, Yukihiro Nakamura, et al. “GABAB Receptors Induce
Phasic Release from Medial Habenula Terminals through Activity-Dependent Recruitment
of Release-Ready Vesicles.” Proceedings of the National Academy of Sciences.
Proceedings of the National Academy of Sciences, 2024. https://doi.org/10.1073/pnas.2301449121.
ieee: P. Koppensteiner et al., “GABAB receptors induce phasic release from
medial habenula terminals through activity-dependent recruitment of release-ready
vesicles,” Proceedings of the National Academy of Sciences, vol. 121, no.
8. Proceedings of the National Academy of Sciences, 2024.
ista: Koppensteiner P, Bhandari P, Önal C, Borges Merjane C, Le Monnier E, Roy U,
Nakamura Y, Sadakata T, Sanbo M, Hirabayashi M, Rhee J, Brose N, Jonas PM, Shigemoto
R. 2024. GABAB receptors induce phasic release from medial habenula terminals
through activity-dependent recruitment of release-ready vesicles. Proceedings
of the National Academy of Sciences. 121(8), e2301449121.
mla: Koppensteiner, Peter, et al. “GABAB Receptors Induce Phasic Release from Medial
Habenula Terminals through Activity-Dependent Recruitment of Release-Ready Vesicles.”
Proceedings of the National Academy of Sciences, vol. 121, no. 8, e2301449121,
Proceedings of the National Academy of Sciences, 2024, doi:10.1073/pnas.2301449121.
short: P. Koppensteiner, P. Bhandari, C. Önal, C. Borges Merjane, E. Le Monnier,
U. Roy, Y. Nakamura, T. Sadakata, M. Sanbo, M. Hirabayashi, J. Rhee, N. Brose,
P.M. Jonas, R. Shigemoto, Proceedings of the National Academy of Sciences 121
(2024).
date_created: 2024-03-05T09:23:55Z
date_published: 2024-02-20T00:00:00Z
date_updated: 2024-03-12T13:44:18Z
day: '20'
ddc:
- '570'
department:
- _id: RySh
- _id: PeJo
doi: 10.1073/pnas.2301449121
ec_funded: 1
external_id:
pmid:
- '38346189'
file:
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file_size: 13648221
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intvolume: ' 121'
issue: '8'
language:
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month: '02'
oa: 1
oa_version: Published Version
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: 2564DBCA-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '665385'
name: International IST Doctoral Program
publication: Proceedings of the National Academy of Sciences
publication_identifier:
eissn:
- 1091-6490
issn:
- 0027-8424
publication_status: published
publisher: Proceedings of the National Academy of Sciences
quality_controlled: '1'
related_material:
link:
- description: News on ISTA Website
relation: press_release
url: https://ista.ac.at/en/news/neuronal-insights-flash-and-freeze-fracture/
record:
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relation: research_data
status: public
status: public
title: GABAB receptors induce phasic release from medial habenula terminals through
activity-dependent recruitment of release-ready vesicles
tmp:
image: /images/cc_by_nc_nd.png
legal_code_url: https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode
name: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
(CC BY-NC-ND 4.0)
short: CC BY-NC-ND (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 121
year: '2024'
...
---
_id: '14843'
abstract:
- lang: eng
text: The coupling between Ca2+ channels and release sensors is a key factor defining
the signaling properties of a synapse. However, the coupling nanotopography at
many synapses remains unknown, and it is unclear how it changes during development.
To address these questions, we examined coupling at the cerebellar inhibitory
basket cell (BC)-Purkinje cell (PC) synapse. Biophysical analysis of transmission
by paired recording and intracellular pipette perfusion revealed that the effects
of exogenous Ca2+ chelators decreased during development, despite constant reliance
of release on P/Q-type Ca2+ channels. Structural analysis by freeze-fracture replica
labeling (FRL) and transmission electron microscopy (EM) indicated that presynaptic
P/Q-type Ca2+ channels formed nanoclusters throughout development, whereas docked
vesicles were only clustered at later developmental stages. Modeling suggested
a developmental transformation from a more random to a more clustered coupling
nanotopography. Thus, presynaptic signaling developmentally approaches a point-to-point
configuration, optimizing speed, reliability, and energy efficiency of synaptic
transmission.
acknowledged_ssus:
- _id: EM-Fac
- _id: PreCl
- _id: M-Shop
acknowledgement: We thank Drs. David DiGregorio and Erwin Neher for critically reading
an earlier version of the manuscript, Ralf Schneggenburger for helpful discussions,
Benjamin Suter and Katharina Lichter for support with image analysis, Chris Wojtan
for advice on numerical solution of partial differential equations, Maria Reva for
help with Ripley analysis, Alois Schlögl for programming, and Akari Hagiwara and
Toshihisa Ohtsuka for anti-ELKS antibody. We are grateful to Florian Marr, Christina
Altmutter, and Vanessa Zheden for excellent technical assistance and to Eleftheria
Kralli-Beller for manuscript editing. This research was supported by the Scientific
Services Units (SSUs) of ISTA (Electron Microscopy Facility, Preclinical Facility,
and Machine Shop). The project received funding from the European Research Council
(ERC) under the European Union’s Horizon 2020 research and innovation program (grant
agreement no. 692692), the Fonds zur Förderung der Wissenschaftlichen Forschung
(Z 312-B27, Wittgenstein award; P 36232-B), all to P.J., and a DOC fellowship of
the Austrian Academy of Sciences to J.-J.C.
article_processing_charge: No
article_type: original
author:
- first_name: JingJing
full_name: Chen, JingJing
id: 2C4E65C8-F248-11E8-B48F-1D18A9856A87
last_name: Chen
- first_name: Walter
full_name: Kaufmann, Walter
id: 3F99E422-F248-11E8-B48F-1D18A9856A87
last_name: Kaufmann
orcid: 0000-0001-9735-5315
- first_name: Chong
full_name: Chen, Chong
id: 3DFD581A-F248-11E8-B48F-1D18A9856A87
last_name: Chen
- first_name: Itaru
full_name: Arai, Itaru
id: 32A73F6C-F248-11E8-B48F-1D18A9856A87
last_name: Arai
- first_name: Olena
full_name: Kim, Olena
id: 3F8ABDDA-F248-11E8-B48F-1D18A9856A87
last_name: Kim
- first_name: Ryuichi
full_name: Shigemoto, Ryuichi
id: 499F3ABC-F248-11E8-B48F-1D18A9856A87
last_name: Shigemoto
orcid: 0000-0001-8761-9444
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
citation:
ama: Chen J, Kaufmann W, Chen C, et al. Developmental transformation of Ca2+ channel-vesicle
nanotopography at a central GABAergic synapse. Neuron. doi:10.1016/j.neuron.2023.12.002
apa: Chen, J., Kaufmann, W., Chen, C., Arai, itaru, Kim, O., Shigemoto, R., &
Jonas, P. M. (n.d.). Developmental transformation of Ca2+ channel-vesicle nanotopography
at a central GABAergic synapse. Neuron. Elsevier. https://doi.org/10.1016/j.neuron.2023.12.002
chicago: Chen, JingJing, Walter Kaufmann, Chong Chen, itaru Arai, Olena Kim, Ryuichi
Shigemoto, and Peter M Jonas. “Developmental Transformation of Ca2+ Channel-Vesicle
Nanotopography at a Central GABAergic Synapse.” Neuron. Elsevier, n.d.
https://doi.org/10.1016/j.neuron.2023.12.002.
ieee: J. Chen et al., “Developmental transformation of Ca2+ channel-vesicle
nanotopography at a central GABAergic synapse,” Neuron. Elsevier.
ista: Chen J, Kaufmann W, Chen C, Arai itaru, Kim O, Shigemoto R, Jonas PM. Developmental
transformation of Ca2+ channel-vesicle nanotopography at a central GABAergic synapse.
Neuron.
mla: Chen, JingJing, et al. “Developmental Transformation of Ca2+ Channel-Vesicle
Nanotopography at a Central GABAergic Synapse.” Neuron, Elsevier, doi:10.1016/j.neuron.2023.12.002.
short: J. Chen, W. Kaufmann, C. Chen, itaru Arai, O. Kim, R. Shigemoto, P.M. Jonas,
Neuron (n.d.).
date_created: 2024-01-21T23:00:56Z
date_published: 2024-01-11T00:00:00Z
date_updated: 2024-03-14T13:14:18Z
day: '11'
department:
- _id: PeJo
- _id: EM-Fac
- _id: RySh
doi: 10.1016/j.neuron.2023.12.002
ec_funded: 1
external_id:
pmid:
- '38215739'
language:
- iso: eng
month: '01'
oa_version: None
pmid: 1
project:
- _id: 25B7EB9E-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '692692'
name: Biophysics and circuit function of a giant cortical glumatergic synapse
- _id: 25C5A090-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: Z00312
name: The Wittgenstein Prize
- _id: bd88be38-d553-11ed-ba76-81d5a70a6ef5
grant_number: P36232
name: Mechanisms of GABA release in hippocampal circuits
- _id: 26B66A3E-B435-11E9-9278-68D0E5697425
grant_number: '25383'
name: Development of nanodomain coupling between Ca2+ channels and release sensors
at a central inhibitory synapse
publication: Neuron
publication_identifier:
eissn:
- 1097-4199
issn:
- 0896-6273
publication_status: inpress
publisher: Elsevier
quality_controlled: '1'
related_material:
link:
- description: News on ISTA Website
relation: press_release
url: https://ista.ac.at/en/news/synapses-brought-to-the-point/
record:
- id: '15101'
relation: dissertation_contains
status: public
scopus_import: '1'
status: public
title: Developmental transformation of Ca2+ channel-vesicle nanotopography at a central
GABAergic synapse
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
year: '2024'
...
---
_id: '15101'
acknowledged_ssus:
- _id: EM-Fac
alternative_title:
- ISTA Thesis
article_processing_charge: No
author:
- first_name: JingJing
full_name: Chen, JingJing
id: 2C4E65C8-F248-11E8-B48F-1D18A9856A87
last_name: Chen
citation:
ama: Chen J. Developmental transformation of nanodomain coupling between Ca2+ channels
and release sensors at a central GABAergic synapse. 2024. doi:10.15479/at:ista:15101
apa: Chen, J. (2024). Developmental transformation of nanodomain coupling between
Ca2+ channels and release sensors at a central GABAergic synapse. Institute
of Science and Technology Austria. https://doi.org/10.15479/at:ista:15101
chicago: Chen, JingJing. “Developmental Transformation of Nanodomain Coupling between
Ca2+ Channels and Release Sensors at a Central GABAergic Synapse.” Institute of
Science and Technology Austria, 2024. https://doi.org/10.15479/at:ista:15101.
ieee: J. Chen, “Developmental transformation of nanodomain coupling between Ca2+
channels and release sensors at a central GABAergic synapse,” Institute of Science
and Technology Austria, 2024.
ista: Chen J. 2024. Developmental transformation of nanodomain coupling between
Ca2+ channels and release sensors at a central GABAergic synapse. Institute of
Science and Technology Austria.
mla: Chen, JingJing. Developmental Transformation of Nanodomain Coupling between
Ca2+ Channels and Release Sensors at a Central GABAergic Synapse. Institute
of Science and Technology Austria, 2024, doi:10.15479/at:ista:15101.
short: J. Chen, Developmental Transformation of Nanodomain Coupling between Ca2+
Channels and Release Sensors at a Central GABAergic Synapse, Institute of Science
and Technology Austria, 2024.
date_created: 2024-03-11T10:09:54Z
date_published: 2024-03-11T00:00:00Z
date_updated: 2024-03-14T13:14:19Z
day: '11'
ddc:
- '570'
degree_awarded: PhD
department:
- _id: GradSch
- _id: PeJo
doi: 10.15479/at:ista:15101
ec_funded: 1
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date_updated: 2024-03-12T07:12:17Z
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file_name: Thesis_Jingjing CHEN.docx
file_size: 11271363
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file_date_updated: 2024-03-12T07:12:17Z
has_accepted_license: '1'
language:
- iso: eng
month: '03'
oa_version: Published Version
page: '84'
project:
- _id: 25B7EB9E-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '692692'
name: Biophysics and circuit function of a giant cortical glumatergic synapse
- _id: 25C5A090-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: Z00312
name: The Wittgenstein Prize
- _id: bd88be38-d553-11ed-ba76-81d5a70a6ef5
grant_number: P36232
name: Mechanisms of GABA release in hippocampal circuits
- _id: 26B66A3E-B435-11E9-9278-68D0E5697425
grant_number: '25383'
name: Development of nanodomain coupling between Ca2+ channels and release sensors
at a central inhibitory synapse
publication_identifier:
issn:
- 2663 - 337X
publication_status: published
publisher: Institute of Science and Technology Austria
related_material:
record:
- id: '14843'
relation: part_of_dissertation
status: public
status: public
supervisor:
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
title: Developmental transformation of nanodomain coupling between Ca2+ channels and
release sensors at a central GABAergic synapse
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: dissertation
user_id: 8b945eb4-e2f2-11eb-945a-df72226e66a9
year: '2024'
...
---
_id: '15117'
abstract:
- lang: eng
text: 'The hippocampal mossy fiber synapse, formed between axons of dentate gyrus
granule cells and dendrites of CA3 pyramidal neurons, is a key synapse in the
trisynaptic circuitry of the hippocampus. Because of its comparatively large size,
this synapse is accessible to direct presynaptic recording, allowing a rigorous
investigation of the biophysical mechanisms of synaptic transmission and plasticity.
Furthermore, because of its placement in the very center of the hippocampal memory
circuit, this synapse seems to be critically involved in several higher network
functions, such as learning, memory, pattern separation, and pattern completion.
Recent work based on new technologies in both nanoanatomy and nanophysiology,
including presynaptic patch-clamp recording, paired recording, super-resolution
light microscopy, and freeze-fracture and “flash-and-freeze” electron microscopy,
has provided new insights into the structure, biophysics, and network function
of this intriguing synapse. This brings us one step closer to answering a fundamental
question in neuroscience: how basic synaptic properties shape higher network computations.'
acknowledgement: "We thank previous students, postdocs, and collaborators, particularly
J. Geiger, and (in alphabetical order) H. Alle, J. Bischofberger, C. Borges-Merjane,
D. Engel, M. Frotscher, S. Hallermann, M. Heckmann, S. Jamrichova, O. Kim, L. Li,
K. Lichter, P. Lin, J. Lübke, Y. Okamoto, C. Pawlu, C. Schmidt-Hieber, N. Spruston,
and N. Vyleta for their outstanding experimental contributions. We also thank P.
Castillo, J. Geiger, T. Sakaba, S. Siegert, T. Vogels, and J. Watson for critically
reading the manuscript, E. Kralli-Beller for text editing, and J. Malikovic and
L. Slomianka for useful discussions. We apologize that, due to space constraints,
not all relevant papers could be cited.\r\nThis project was supported by the European
Research Council (ERC) under the European Union’s Horizon 2020 research and innovation
program (grant agreement 692692, AdG “GIANTSYN”) and the Fonds zur Förderung der
Wissenschaftlichen Forschung (Z 312-B27, Wittgenstein Award; P 36232-B, stand-alone
grant), both to P.J."
article_processing_charge: No
article_type: review
author:
- first_name: David H
full_name: Vandael, David H
id: 3AE48E0A-F248-11E8-B48F-1D18A9856A87
last_name: Vandael
orcid: 0000-0001-7577-1676
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
citation:
ama: Vandael DH, Jonas PM. Structure, biophysics, and circuit function of a “giant”
cortical presynaptic terminal. Science. 2024;383(6687):eadg6757. doi:10.1126/science.adg6757
apa: Vandael, D. H., & Jonas, P. M. (2024). Structure, biophysics, and circuit
function of a “giant” cortical presynaptic terminal. Science. AAAS. https://doi.org/10.1126/science.adg6757
chicago: Vandael, David H, and Peter M Jonas. “Structure, Biophysics, and Circuit
Function of a ‘Giant’ Cortical Presynaptic Terminal.” Science. AAAS, 2024.
https://doi.org/10.1126/science.adg6757.
ieee: D. H. Vandael and P. M. Jonas, “Structure, biophysics, and circuit function
of a ‘giant’ cortical presynaptic terminal,” Science, vol. 383, no. 6687.
AAAS, p. eadg6757, 2024.
ista: Vandael DH, Jonas PM. 2024. Structure, biophysics, and circuit function of
a ‘giant’ cortical presynaptic terminal. Science. 383(6687), eadg6757.
mla: Vandael, David H., and Peter M. Jonas. “Structure, Biophysics, and Circuit
Function of a ‘Giant’ Cortical Presynaptic Terminal.” Science, vol. 383,
no. 6687, AAAS, 2024, p. eadg6757, doi:10.1126/science.adg6757.
short: D.H. Vandael, P.M. Jonas, Science 383 (2024) eadg6757.
date_created: 2024-03-17T23:00:57Z
date_published: 2024-03-08T00:00:00Z
date_updated: 2024-03-20T07:42:52Z
day: '08'
department:
- _id: PeJo
doi: 10.1126/science.adg6757
ec_funded: 1
external_id:
pmid:
- '38452088'
intvolume: ' 383'
issue: '6687'
language:
- iso: eng
month: '03'
oa_version: None
page: eadg6757
pmid: 1
project:
- _id: 25B7EB9E-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '692692'
name: Biophysics and circuit function of a giant cortical glumatergic synapse
- _id: 25C5A090-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: Z00312
name: The Wittgenstein Prize
- _id: bd88be38-d553-11ed-ba76-81d5a70a6ef5
grant_number: P36232
name: Mechanisms of GABA release in hippocampal circuits
publication: Science
publication_identifier:
eissn:
- 1095-9203
publication_status: published
publisher: AAAS
quality_controlled: '1'
scopus_import: '1'
status: public
title: Structure, biophysics, and circuit function of a "giant" cortical presynaptic
terminal
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 383
year: '2024'
...
---
_id: '12720'
abstract:
- lang: eng
text: Here we describe the in vivo DNA assembly approach, where molecular cloning
procedures are performed using an E. coli recA-independent recombination pathway,
which assembles linear fragments of DNA with short homologous termini. This pathway
is present in all standard laboratory E. coli strains and, by bypassing the need
for in vitro DNA assembly, allows simplified molecular cloning to be performed
without the plasmid instability issues associated with specialized recombination-cloning
bacterial strains. The methodology requires specific primer design and can perform
all standard plasmid modifications (insertions, deletions, mutagenesis, and sub-cloning)
in a rapid, simple, and cost-efficient manner, as it does not require commercial
kits or specialized bacterial strains. Additionally, this approach can be used
to perform complex procedures such as multiple modifications to a plasmid, as
up to 6 linear fragments can be assembled in vivo by this recombination pathway.
Procedures generally require less than 3 h, involving PCR amplification, DpnI
digestion of template DNA, and transformation, upon which circular plasmids are
assembled. In this chapter we describe the requirements, procedure, and potential
pitfalls when using this technique, as well as protocol variations to overcome
the most common issues.
alternative_title:
- Methods in Molecular Biology
article_processing_charge: No
author:
- first_name: Sandra
full_name: Arroyo-Urea, Sandra
last_name: Arroyo-Urea
- first_name: Jake
full_name: Watson, Jake
id: 63836096-4690-11EA-BD4E-32803DDC885E
last_name: Watson
orcid: 0000-0002-8698-3823
- first_name: Javier
full_name: García-Nafría, Javier
last_name: García-Nafría
citation:
ama: 'Arroyo-Urea S, Watson J, García-Nafría J. Molecular Cloning Using In Vivo
DNA Assembly. In: Scarlett G, ed. DNA Manipulation and Analysis. Vol 2633.
MIMB. New York, NY, United States: Springer Nature; 2023:33-44. doi:10.1007/978-1-0716-3004-4_3'
apa: 'Arroyo-Urea, S., Watson, J., & García-Nafría, J. (2023). Molecular Cloning
Using In Vivo DNA Assembly. In G. Scarlett (Ed.), DNA Manipulation and Analysis
(Vol. 2633, pp. 33–44). New York, NY, United States: Springer Nature. https://doi.org/10.1007/978-1-0716-3004-4_3'
chicago: 'Arroyo-Urea, Sandra, Jake Watson, and Javier García-Nafría. “Molecular
Cloning Using In Vivo DNA Assembly.” In DNA Manipulation and Analysis,
edited by Garry Scarlett, 2633:33–44. MIMB. New York, NY, United States: Springer
Nature, 2023. https://doi.org/10.1007/978-1-0716-3004-4_3.'
ieee: 'S. Arroyo-Urea, J. Watson, and J. García-Nafría, “Molecular Cloning Using
In Vivo DNA Assembly,” in DNA Manipulation and Analysis, vol. 2633, G.
Scarlett, Ed. New York, NY, United States: Springer Nature, 2023, pp. 33–44.'
ista: 'Arroyo-Urea S, Watson J, García-Nafría J. 2023.Molecular Cloning Using In
Vivo DNA Assembly. In: DNA Manipulation and Analysis. Methods in Molecular Biology,
vol. 2633, 33–44.'
mla: Arroyo-Urea, Sandra, et al. “Molecular Cloning Using In Vivo DNA Assembly.”
DNA Manipulation and Analysis, edited by Garry Scarlett, vol. 2633, Springer
Nature, 2023, pp. 33–44, doi:10.1007/978-1-0716-3004-4_3.
short: S. Arroyo-Urea, J. Watson, J. García-Nafría, in:, G. Scarlett (Ed.), DNA
Manipulation and Analysis, Springer Nature, New York, NY, United States, 2023,
pp. 33–44.
date_created: 2023-03-12T23:01:02Z
date_published: 2023-03-01T00:00:00Z
date_updated: 2023-03-16T08:34:24Z
day: '01'
department:
- _id: PeJo
doi: 10.1007/978-1-0716-3004-4_3
editor:
- first_name: Garry
full_name: Scarlett, Garry
last_name: Scarlett
external_id:
pmid:
- '36853454'
intvolume: ' 2633'
language:
- iso: eng
month: '03'
oa_version: None
page: 33-44
place: New York, NY, United States
pmid: 1
publication: DNA Manipulation and Analysis
publication_identifier:
eisbn:
- 978-1-0716-3004-4
eissn:
- 1940-6029
isbn:
- 978-1-0716-3003-7
issn:
- 1064-3745
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
series_title: MIMB
status: public
title: Molecular Cloning Using In Vivo DNA Assembly
type: book_chapter
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 2633
year: '2023'
...
---
_id: '12567'
abstract:
- lang: eng
text: Single-molecule localization microscopy (SMLM) greatly advances structural
studies of diverse biological tissues. For example, presynaptic active zone (AZ)
nanotopology is resolved in increasing detail. Immunofluorescence imaging of AZ
proteins usually relies on epitope preservation using aldehyde-based immunocompetent
fixation. Cryofixation techniques, such as high-pressure freezing (HPF) and freeze
substitution (FS), are widely used for ultrastructural studies of presynaptic
architecture in electron microscopy (EM). HPF/FS demonstrated nearer-to-native
preservation of AZ ultrastructure, e.g., by facilitating single filamentous structures.
Here, we present a protocol combining the advantages of HPF/FS and direct stochastic
optical reconstruction microscopy (dSTORM) to quantify nanotopology of the AZ
scaffold protein Bruchpilot (Brp) at neuromuscular junctions (NMJs) of Drosophila
melanogaster. Using this standardized model, we tested for preservation of Brp
clusters in different FS protocols compared to classical aldehyde fixation. In
HPF/FS samples, presynaptic boutons were structurally well preserved with ~22%
smaller Brp clusters that allowed quantification of subcluster topology. In summary,
we established a standardized near-to-native preparation and immunohistochemistry
protocol for SMLM analyses of AZ protein clusters in a defined model synapse.
Our protocol could be adapted to study protein arrangements at single-molecule
resolution in other intact tissue preparations.
acknowledgement: This work has been supported by funding of the German Research Foundation
(Deutsche Forschungsgemeinschaft [DFG], CRC 166, Project B06 to M.H. and A.-L.S.,
FOR 3004 SYNABS P1 to M.H.) and by the Interdisciplinary Clinical Research Center
(IZKF) Würzburg (Z-3/69 to M.M.P., N-229 to M.H. and A.-L.S.). A.M. is funded by
the University of Leipzig Clinician Scientist Program.
article_number: '2128'
article_processing_charge: No
article_type: original
author:
- first_name: Achmed
full_name: Mrestani, Achmed
last_name: Mrestani
- first_name: Katharina
full_name: Lichter, Katharina
id: 39302e62-fcfc-11ec-8196-8b01447dbd3d
last_name: Lichter
- first_name: Anna Leena
full_name: Sirén, Anna Leena
last_name: Sirén
- first_name: Manfred
full_name: Heckmann, Manfred
last_name: Heckmann
- first_name: Mila M.
full_name: Paul, Mila M.
last_name: Paul
- first_name: Martin
full_name: Pauli, Martin
last_name: Pauli
citation:
ama: Mrestani A, Lichter K, Sirén AL, Heckmann M, Paul MM, Pauli M. Single-molecule
localization microscopy of presynaptic active zones in Drosophila melanogaster
after rapid cryofixation. International Journal of Molecular Sciences.
2023;24(3). doi:10.3390/ijms24032128
apa: Mrestani, A., Lichter, K., Sirén, A. L., Heckmann, M., Paul, M. M., & Pauli,
M. (2023). Single-molecule localization microscopy of presynaptic active zones
in Drosophila melanogaster after rapid cryofixation. International Journal
of Molecular Sciences. MDPI. https://doi.org/10.3390/ijms24032128
chicago: Mrestani, Achmed, Katharina Lichter, Anna Leena Sirén, Manfred Heckmann,
Mila M. Paul, and Martin Pauli. “Single-Molecule Localization Microscopy of Presynaptic
Active Zones in Drosophila Melanogaster after Rapid Cryofixation.” International
Journal of Molecular Sciences. MDPI, 2023. https://doi.org/10.3390/ijms24032128.
ieee: A. Mrestani, K. Lichter, A. L. Sirén, M. Heckmann, M. M. Paul, and M. Pauli,
“Single-molecule localization microscopy of presynaptic active zones in Drosophila
melanogaster after rapid cryofixation,” International Journal of Molecular
Sciences, vol. 24, no. 3. MDPI, 2023.
ista: Mrestani A, Lichter K, Sirén AL, Heckmann M, Paul MM, Pauli M. 2023. Single-molecule
localization microscopy of presynaptic active zones in Drosophila melanogaster
after rapid cryofixation. International Journal of Molecular Sciences. 24(3),
2128.
mla: Mrestani, Achmed, et al. “Single-Molecule Localization Microscopy of Presynaptic
Active Zones in Drosophila Melanogaster after Rapid Cryofixation.” International
Journal of Molecular Sciences, vol. 24, no. 3, 2128, MDPI, 2023, doi:10.3390/ijms24032128.
short: A. Mrestani, K. Lichter, A.L. Sirén, M. Heckmann, M.M. Paul, M. Pauli, International
Journal of Molecular Sciences 24 (2023).
date_created: 2023-02-19T23:00:56Z
date_published: 2023-01-21T00:00:00Z
date_updated: 2023-08-01T13:16:36Z
day: '21'
ddc:
- '570'
department:
- _id: PeJo
doi: 10.3390/ijms24032128
external_id:
isi:
- '000930324700001'
file:
- access_level: open_access
checksum: 69a35dcd3e0249f902ab881b06ee2e58
content_type: application/pdf
creator: dernst
date_created: 2023-02-20T07:09:27Z
date_updated: 2023-02-20T07:09:27Z
file_id: '12569'
file_name: 2023_IJMS_Mrestani.pdf
file_size: 2823025
relation: main_file
success: 1
file_date_updated: 2023-02-20T07:09:27Z
has_accepted_license: '1'
intvolume: ' 24'
isi: 1
issue: '3'
language:
- iso: eng
month: '01'
oa: 1
oa_version: Published Version
publication: International Journal of Molecular Sciences
publication_identifier:
eissn:
- 1422-0067
publication_status: published
publisher: MDPI
quality_controlled: '1'
scopus_import: '1'
status: public
title: Single-molecule localization microscopy of presynaptic active zones in Drosophila
melanogaster after rapid cryofixation
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: 4359f0d1-fa6c-11eb-b949-802e58b17ae8
volume: 24
year: '2023'
...
---
_id: '12759'
abstract:
- lang: eng
text: Stereological methods for estimating the 3D particle size and density from
2D projections are essential to many research fields. These methods are, however,
prone to errors arising from undetected particle profiles due to sectioning and
limited resolution, known as ‘lost caps’. A potential solution developed by Keiding,
Jensen, and Ranek in 1972, which we refer to as the Keiding model, accounts for
lost caps by quantifying the smallest detectable profile in terms of its limiting
‘cap angle’ (ϕ), a size-independent measure of a particle’s distance from the
section surface. However, this simple solution has not been widely adopted nor
tested. Rather, model-independent design-based stereological methods, which do
not explicitly account for lost caps, have come to the fore. Here, we provide
the first experimental validation of the Keiding model by comparing the size and
density of particles estimated from 2D projections with direct measurement from
3D EM reconstructions of the same tissue. We applied the Keiding model to estimate
the size and density of somata, nuclei and vesicles in the cerebellum of mice
and rats, where high packing density can be problematic for design-based methods.
Our analysis reveals a Gaussian distribution for ϕ rather than a single value.
Nevertheless, curve fits of the Keiding model to the 2D diameter distribution
accurately estimate the mean ϕ and 3D diameter distribution. While systematic
testing using simulations revealed an upper limit to determining ϕ, our analysis
shows that estimated ϕ can be used to determine the 3D particle density from the
2D density under a wide range of conditions, and this method is potentially more
accurate than minimum-size-based lost-cap corrections and disector methods. Our
results show the Keiding model provides an efficient means of accurately estimating
the size and density of particles from 2D projections even under conditions of
a high density.
acknowledged_ssus:
- _id: EM-Fac
acknowledgement: "We thank the IST Austria Electron Microscopy Facility for technical
support, and Diccon Coyle, Andrea Lőrincz and Zoltan Nusser for their helpful comments
and discussions.\r\nFunding for JSR and RAS was from the Wellcome Trust (203048;
224499; https://\r\nwellcome.org/). RAS is in receipt of a Wellcome Trust Principal
Research Fellowship (224499).\r\nFunding for CBM and PJ was from Fond zur Förderung
der Wissenschaftlichen Forschung (V\r\n739-B27 Elise-Richter Programme to CBM, Z
312-B27 Wittgenstein Award to PJ; \r\nhttps://www.fwf.ac.at). PJ received funding
from the European Research Council (ERC; https://erc.europa.eu) under the European
Union’s Horizon 2020 research and innovation programme (grant agreement no. 692692).
NH was supported by a European\r\nResearch Council Advanced Grant (ERC-AG787157)."
article_number: e0277148
article_processing_charge: No
article_type: original
author:
- first_name: Jason Seth
full_name: Rothman, Jason Seth
last_name: Rothman
- first_name: Carolina
full_name: Borges Merjane, Carolina
id: 4305C450-F248-11E8-B48F-1D18A9856A87
last_name: Borges Merjane
orcid: 0000-0003-0005-401X
- first_name: Noemi
full_name: Holderith, Noemi
last_name: Holderith
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
- first_name: R.
full_name: Angus Silver, R.
last_name: Angus Silver
citation:
ama: Rothman JS, Borges Merjane C, Holderith N, Jonas PM, Angus Silver R. Validation
of a stereological method for estimating particle size and density from 2D projections
with high accuracy. PLoS ONE. 2023;18(3 March). doi:10.1371/journal.pone.0277148
apa: Rothman, J. S., Borges Merjane, C., Holderith, N., Jonas, P. M., & Angus
Silver, R. (2023). Validation of a stereological method for estimating particle
size and density from 2D projections with high accuracy. PLoS ONE. Public
Library of Science. https://doi.org/10.1371/journal.pone.0277148
chicago: Rothman, Jason Seth, Carolina Borges Merjane, Noemi Holderith, Peter M
Jonas, and R. Angus Silver. “Validation of a Stereological Method for Estimating
Particle Size and Density from 2D Projections with High Accuracy.” PLoS ONE.
Public Library of Science, 2023. https://doi.org/10.1371/journal.pone.0277148.
ieee: J. S. Rothman, C. Borges Merjane, N. Holderith, P. M. Jonas, and R. Angus
Silver, “Validation of a stereological method for estimating particle size and
density from 2D projections with high accuracy,” PLoS ONE, vol. 18, no.
3 March. Public Library of Science, 2023.
ista: Rothman JS, Borges Merjane C, Holderith N, Jonas PM, Angus Silver R. 2023.
Validation of a stereological method for estimating particle size and density
from 2D projections with high accuracy. PLoS ONE. 18(3 March), e0277148.
mla: Rothman, Jason Seth, et al. “Validation of a Stereological Method for Estimating
Particle Size and Density from 2D Projections with High Accuracy.” PLoS ONE,
vol. 18, no. 3 March, e0277148, Public Library of Science, 2023, doi:10.1371/journal.pone.0277148.
short: J.S. Rothman, C. Borges Merjane, N. Holderith, P.M. Jonas, R. Angus Silver,
PLoS ONE 18 (2023).
date_created: 2023-03-26T22:01:07Z
date_published: 2023-03-17T00:00:00Z
date_updated: 2023-08-01T13:46:39Z
day: '17'
ddc:
- '570'
department:
- _id: PeJo
doi: 10.1371/journal.pone.0277148
ec_funded: 1
external_id:
isi:
- '001024737400001'
file:
- access_level: open_access
checksum: 2380331ec27cc87808826fc64419ac1c
content_type: application/pdf
creator: dernst
date_created: 2023-03-27T06:51:09Z
date_updated: 2023-03-27T06:51:09Z
file_id: '12770'
file_name: 2023_PLoSOne_Rothman.pdf
file_size: 7290413
relation: main_file
success: 1
file_date_updated: 2023-03-27T06:51:09Z
has_accepted_license: '1'
intvolume: ' 18'
isi: 1
issue: 3 March
language:
- iso: eng
month: '03'
oa: 1
oa_version: Published Version
project:
- _id: 25B7EB9E-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '692692'
name: Biophysics and circuit function of a giant cortical glumatergic synapse
- _id: 25C5A090-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: Z00312
name: The Wittgenstein Prize
- _id: 2696E7FE-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: V00739
name: Structural plasticity at mossy fiber-CA3 synapses
publication: PLoS ONE
publication_identifier:
eissn:
- 1932-6203
publication_status: published
publisher: Public Library of Science
quality_controlled: '1'
scopus_import: '1'
status: public
title: Validation of a stereological method for estimating particle size and density
from 2D projections with high accuracy
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: 4359f0d1-fa6c-11eb-b949-802e58b17ae8
volume: 18
year: '2023'
...
---
_id: '12515'
abstract:
- lang: eng
text: "Introduction: The olfactory system in most mammals is divided into several
subsystems based on the anatomical locations of the neuroreceptor cells involved
and the receptor families that are expressed. In addition to the main olfactory
system and the vomeronasal system, a range of olfactory subsystems converge onto
the transition zone located between the main olfactory bulb (MOB) and the accessory
olfactory bulb (AOB), which has been termed the olfactory limbus (OL). The OL
contains specialized glomeruli that receive noncanonical sensory afferences and
which interact with the MOB and AOB. Little is known regarding the olfactory subsystems
of mammals other than laboratory rodents.\r\nMethods: We have focused on characterizing
the OL in the red fox by performing general and specific histological stainings
on serial sections, using both single and double immunohistochemical and lectin-histochemical
labeling techniques.\r\nResults: As a result, we have been able to determine that
the OL of the red fox (Vulpes vulpes) displays an uncommonly high degree of development
and complexity.\r\nDiscussion: This makes this species a novel mammalian model,
the study of which could improve our understanding of the noncanonical pathways
involved in the processing of chemosensory cues."
acknowledgement: This work was partially supported by a grant from “Consello Social
Universidade de Santiago de Compostela” 2022-PU004.We would like to show special
gratitude to Prof. Ludwig Wagner (Medical University, Vienna) for kindly providing
us with the secretagogin antibody. We thank the Wildlife Recovery Centres of Galicia,
Dirección Xeral de Patrimonio Natural (Xunta de Galicia, Spain), and Federación
Galega de Caza for providing the red foxes used in this study.
article_number: '1097467'
article_processing_charge: No
article_type: original
author:
- first_name: Irene
full_name: Ortiz-Leal, Irene
last_name: Ortiz-Leal
- first_name: Mateo V.
full_name: Torres, Mateo V.
last_name: Torres
- first_name: Victor M
full_name: Vargas Barroso, Victor M
id: 2F55A9DE-F248-11E8-B48F-1D18A9856A87
last_name: Vargas Barroso
- first_name: Luis Eusebio
full_name: Fidalgo, Luis Eusebio
last_name: Fidalgo
- first_name: Ana María
full_name: López-Beceiro, Ana María
last_name: López-Beceiro
- first_name: Jorge A.
full_name: Larriva-Sahd, Jorge A.
last_name: Larriva-Sahd
- first_name: Pablo
full_name: Sánchez-Quinteiro, Pablo
last_name: Sánchez-Quinteiro
citation:
ama: Ortiz-Leal I, Torres MV, Vargas Barroso VM, et al. The olfactory limbus of
the red fox (Vulpes vulpes). New insights regarding a noncanonical olfactory bulb
pathway. Frontiers in Neuroanatomy. 2023;16. doi:10.3389/fnana.2022.1097467
apa: Ortiz-Leal, I., Torres, M. V., Vargas Barroso, V. M., Fidalgo, L. E., López-Beceiro,
A. M., Larriva-Sahd, J. A., & Sánchez-Quinteiro, P. (2023). The olfactory
limbus of the red fox (Vulpes vulpes). New insights regarding a noncanonical olfactory
bulb pathway. Frontiers in Neuroanatomy. Frontiers. https://doi.org/10.3389/fnana.2022.1097467
chicago: Ortiz-Leal, Irene, Mateo V. Torres, Victor M Vargas Barroso, Luis Eusebio
Fidalgo, Ana María López-Beceiro, Jorge A. Larriva-Sahd, and Pablo Sánchez-Quinteiro.
“The Olfactory Limbus of the Red Fox (Vulpes Vulpes). New Insights Regarding a
Noncanonical Olfactory Bulb Pathway.” Frontiers in Neuroanatomy. Frontiers,
2023. https://doi.org/10.3389/fnana.2022.1097467.
ieee: I. Ortiz-Leal et al., “The olfactory limbus of the red fox (Vulpes
vulpes). New insights regarding a noncanonical olfactory bulb pathway,” Frontiers
in Neuroanatomy, vol. 16. Frontiers, 2023.
ista: Ortiz-Leal I, Torres MV, Vargas Barroso VM, Fidalgo LE, López-Beceiro AM,
Larriva-Sahd JA, Sánchez-Quinteiro P. 2023. The olfactory limbus of the red fox
(Vulpes vulpes). New insights regarding a noncanonical olfactory bulb pathway.
Frontiers in Neuroanatomy. 16, 1097467.
mla: Ortiz-Leal, Irene, et al. “The Olfactory Limbus of the Red Fox (Vulpes Vulpes).
New Insights Regarding a Noncanonical Olfactory Bulb Pathway.” Frontiers in
Neuroanatomy, vol. 16, 1097467, Frontiers, 2023, doi:10.3389/fnana.2022.1097467.
short: I. Ortiz-Leal, M.V. Torres, V.M. Vargas Barroso, L.E. Fidalgo, A.M. López-Beceiro,
J.A. Larriva-Sahd, P. Sánchez-Quinteiro, Frontiers in Neuroanatomy 16 (2023).
date_created: 2023-02-05T23:01:00Z
date_published: 2023-01-10T00:00:00Z
date_updated: 2023-08-16T11:37:52Z
day: '10'
ddc:
- '570'
department:
- _id: PeJo
doi: 10.3389/fnana.2022.1097467
external_id:
isi:
- '000919786900001'
pmid:
- '36704406'
file:
- access_level: open_access
checksum: 49cd40f3bda6f267079427042e7d15e3
content_type: application/pdf
creator: dernst
date_created: 2023-02-06T07:56:14Z
date_updated: 2023-02-06T07:56:14Z
file_id: '12518'
file_name: 2022_FrontiersNeuroanatomy_OrtizLeal.pdf
file_size: 21943473
relation: main_file
success: 1
file_date_updated: 2023-02-06T07:56:14Z
has_accepted_license: '1'
intvolume: ' 16'
isi: 1
language:
- iso: eng
month: '01'
oa: 1
oa_version: Published Version
pmid: 1
publication: Frontiers in Neuroanatomy
publication_identifier:
eissn:
- 1662-5129
publication_status: published
publisher: Frontiers
quality_controlled: '1'
scopus_import: '1'
status: public
title: The olfactory limbus of the red fox (Vulpes vulpes). New insights regarding
a noncanonical olfactory bulb pathway
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: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 16
year: '2023'
...
---
_id: '12786'
abstract:
- lang: eng
text: AMPA glutamate receptors (AMPARs) mediate excitatory neurotransmission throughout
the brain. Their signalling is uniquely diversified by brain region-specific auxiliary
subunits, providing an opportunity for the development of selective therapeutics.
AMPARs associated with TARP γ8 are enriched in the hippocampus, and are targets
of emerging anti-epileptic drugs. To understand their therapeutic activity, we
determined cryo-EM structures of the GluA1/2-γ8 receptor associated with three
potent, chemically diverse ligands. We find that despite sharing a lipid-exposed
and water-accessible binding pocket, drug action is differentially affected by
binding-site mutants. Together with patch-clamp recordings and MD simulations
we also demonstrate that ligand-triggered reorganisation of the AMPAR-TARP interface
contributes to modulation. Unexpectedly, one ligand (JNJ-61432059) acts bifunctionally,
negatively affecting GluA1 but exerting positive modulatory action on GluA2-containing
AMPARs, in a TARP stoichiometry-dependent manner. These results further illuminate
the action of TARPs, demonstrate the sensitive balance between positive and negative
modulatory action, and provide a mechanistic platform for development of both
positive and negative selective AMPAR modulators.
acknowledgement: We thank James Krieger for generating the ‘proDy’ interaction maps
in Fig. 5B and S7C, and Jan-Niklas Dohrke for critically reading the manuscript.
We thank members of the Greger lab for insightful comments during this study. We
acknowledge Trevor Rutherford for confirming ligand integrity by NMR. We are also
grateful to LMB scientific computing and the EM facility for their support. This
research was funded in part by the Wellcome Trust (223194/Z/21/Z) to I.H.G. For
the purpose of Open Access, the MRC Laboratory of Molecular Biology has applied
a CC BY public copyright licence to any Author Accepted Manuscript (AAM) version
arising from this submission. Further funding came from the Medical Research Council
(MRU105174197) to I.H.G, and NIH grant (R56/R01MH123474) to T.N.
article_number: '1659'
article_processing_charge: No
article_type: original
author:
- first_name: Danyang
full_name: Zhang, Danyang
last_name: Zhang
- first_name: Remigijus
full_name: Lape, Remigijus
last_name: Lape
- first_name: Saher A.
full_name: Shaikh, Saher A.
last_name: Shaikh
- first_name: Bianka K.
full_name: Kohegyi, Bianka K.
last_name: Kohegyi
- first_name: Jake
full_name: Watson, Jake
id: 63836096-4690-11EA-BD4E-32803DDC885E
last_name: Watson
orcid: 0000-0002-8698-3823
- first_name: Ondrej
full_name: Cais, Ondrej
last_name: Cais
- first_name: Terunaga
full_name: Nakagawa, Terunaga
last_name: Nakagawa
- first_name: Ingo H.
full_name: Greger, Ingo H.
last_name: Greger
citation:
ama: Zhang D, Lape R, Shaikh SA, et al. Modulatory mechanisms of TARP γ8-selective
AMPA receptor therapeutics. Nature Communications. 2023;14. doi:10.1038/s41467-023-37259-5
apa: Zhang, D., Lape, R., Shaikh, S. A., Kohegyi, B. K., Watson, J., Cais, O., …
Greger, I. H. (2023). Modulatory mechanisms of TARP γ8-selective AMPA receptor
therapeutics. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-023-37259-5
chicago: Zhang, Danyang, Remigijus Lape, Saher A. Shaikh, Bianka K. Kohegyi, Jake
Watson, Ondrej Cais, Terunaga Nakagawa, and Ingo H. Greger. “Modulatory Mechanisms
of TARP Γ8-Selective AMPA Receptor Therapeutics.” Nature Communications.
Springer Nature, 2023. https://doi.org/10.1038/s41467-023-37259-5.
ieee: D. Zhang et al., “Modulatory mechanisms of TARP γ8-selective AMPA receptor
therapeutics,” Nature Communications, vol. 14. Springer Nature, 2023.
ista: Zhang D, Lape R, Shaikh SA, Kohegyi BK, Watson J, Cais O, Nakagawa T, Greger
IH. 2023. Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics.
Nature Communications. 14, 1659.
mla: Zhang, Danyang, et al. “Modulatory Mechanisms of TARP Γ8-Selective AMPA Receptor
Therapeutics.” Nature Communications, vol. 14, 1659, Springer Nature, 2023,
doi:10.1038/s41467-023-37259-5.
short: D. Zhang, R. Lape, S.A. Shaikh, B.K. Kohegyi, J. Watson, O. Cais, T. Nakagawa,
I.H. Greger, Nature Communications 14 (2023).
date_created: 2023-04-02T22:01:09Z
date_published: 2023-03-25T00:00:00Z
date_updated: 2023-12-13T11:15:58Z
day: '25'
ddc:
- '570'
department:
- _id: PeJo
doi: 10.1038/s41467-023-37259-5
external_id:
isi:
- '001066658700003'
file:
- access_level: open_access
checksum: 0a97b31191432dae5853bbb5ccb7698d
content_type: application/pdf
creator: dernst
date_created: 2023-04-03T06:38:56Z
date_updated: 2023-04-03T06:38:56Z
file_id: '12797'
file_name: 2023_NatureComm_Zhang.pdf
file_size: 2613996
relation: main_file
success: 1
file_date_updated: 2023-04-03T06:38:56Z
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intvolume: ' 14'
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language:
- iso: eng
month: '03'
oa: 1
oa_version: Published Version
publication: Nature Communications
publication_identifier:
eissn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics
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: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 14
year: '2023'
...
---
_id: '13267'
abstract:
- lang: eng
text: Three-dimensional (3D) reconstruction of living brain tissue down to an individual
synapse level would create opportunities for decoding the dynamics and structure–function
relationships of the brain’s complex and dense information processing network;
however, this has been hindered by insufficient 3D resolution, inadequate signal-to-noise
ratio and prohibitive light burden in optical imaging, whereas electron microscopy
is inherently static. Here we solved these challenges by developing an integrated
optical/machine-learning technology, LIONESS (live information-optimized nanoscopy
enabling saturated segmentation). This leverages optical modifications to stimulated
emission depletion microscopy in comprehensively, extracellularly labeled tissue
and previous information on sample structure via machine learning to simultaneously
achieve isotropic super-resolution, high signal-to-noise ratio and compatibility
with living tissue. This allows dense deep-learning-based instance segmentation
and 3D reconstruction at a synapse level, incorporating molecular, activity and
morphodynamic information. LIONESS opens up avenues for studying the dynamic functional
(nano-)architecture of living brain tissue.
acknowledged_ssus:
- _id: ScienComp
- _id: Bio
- _id: PreCl
- _id: E-Lib
- _id: LifeSc
- _id: M-Shop
acknowledgement: "We thank J. Vorlaufer, N. Agudelo and A. Wartak for microscope maintenance
and troubleshooting, C. Kreuzinger and A. Freeman for technical assistance, M. Šuplata
for hardware control support and M. Cunha dos Santos for initial exploration of
software. We\r\nthank P. Henderson for advice on deep-learning training and M. Sixt,
S. Boyd and T. Weiss for discussions and critical reading of the manuscript. L.
Lavis (Janelia Research Campus) generously provided the JF585-HaloTag ligand. We
acknowledge expert support by IST\r\nAustria’s scientific computing, imaging and
optics, preclinical, library and laboratory support facilities and by the Miba machine
shop. We gratefully acknowledge funding by the following sources: Austrian Science
Fund (F.W.F.) grant no. I3600-B27 (J.G.D.), grant no. DK W1232\r\n(J.G.D. and J.M.M.)
and grant no. Z 312-B27, Wittgenstein award (P.J.); the Gesellschaft für Forschungsförderung
NÖ grant no. LSC18-022 (J.G.D.); an ISTA Interdisciplinary project grant (J.G.D.
and B.B.); the European Union’s Horizon 2020 research and innovation programme,\r\nMarie-Skłodowska
Curie grant 665385 (J.M.M. and J.L.); the European Union’s Horizon 2020 research
and innovation programme, European Research Council grant no. 715767, MATERIALIZABLE
(B.B.); grant no. 715508, REVERSEAUTISM (G.N.); grant no. 695568, SYNNOVATE (S.G.N.G.);
and grant no. 692692, GIANTSYN (P.J.); the Simons\r\nFoundation Autism Research
Initiative grant no. 529085 (S.G.N.G.); the Wellcome Trust Technology Development
grant no. 202932 (S.G.N.G.); the Marie Skłodowska-Curie Actions Individual Fellowship
no. 101026635 under the EU Horizon 2020 program (J.F.W.);\r\nthe Human Frontier
Science Program postdoctoral fellowship LT000557/2018 (W.J.); and the National Science
Foundation grant no. IIS-1835231 (H.P.) and NCS-FO-2124179 (H.P.)."
article_processing_charge: Yes
article_type: original
author:
- first_name: Philipp
full_name: Velicky, Philipp
id: 39BDC62C-F248-11E8-B48F-1D18A9856A87
last_name: Velicky
orcid: 0000-0002-2340-7431
- first_name: Eder
full_name: Miguel Villalba, Eder
id: 3FB91342-F248-11E8-B48F-1D18A9856A87
last_name: Miguel Villalba
orcid: 0000-0001-5665-0430
- first_name: Julia M
full_name: Michalska, Julia M
id: 443DB6DE-F248-11E8-B48F-1D18A9856A87
last_name: Michalska
orcid: 0000-0003-3862-1235
- first_name: Julia
full_name: Lyudchik, Julia
id: 46E28B80-F248-11E8-B48F-1D18A9856A87
last_name: Lyudchik
- first_name: Donglai
full_name: Wei, Donglai
last_name: Wei
- first_name: Zudi
full_name: Lin, Zudi
last_name: Lin
- first_name: Jake
full_name: Watson, Jake
id: 63836096-4690-11EA-BD4E-32803DDC885E
last_name: Watson
orcid: 0000-0002-8698-3823
- first_name: Jakob
full_name: Troidl, Jakob
last_name: Troidl
- first_name: Johanna
full_name: Beyer, Johanna
last_name: Beyer
- first_name: Yoav
full_name: Ben Simon, Yoav
id: 43DF3136-F248-11E8-B48F-1D18A9856A87
last_name: Ben Simon
- first_name: Christoph M
full_name: Sommer, Christoph M
id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87
last_name: Sommer
orcid: 0000-0003-1216-9105
- first_name: Wiebke
full_name: Jahr, Wiebke
id: 425C1CE8-F248-11E8-B48F-1D18A9856A87
last_name: Jahr
- first_name: Alban
full_name: Cenameri, Alban
id: 9ac8f577-2357-11eb-997a-e566c5550886
last_name: Cenameri
- first_name: Johannes
full_name: Broichhagen, Johannes
last_name: Broichhagen
- first_name: Seth G.N.
full_name: Grant, Seth G.N.
last_name: Grant
- first_name: Peter M
full_name: Jonas, Peter M
id: 353C1B58-F248-11E8-B48F-1D18A9856A87
last_name: Jonas
orcid: 0000-0001-5001-4804
- first_name: Gaia
full_name: Novarino, Gaia
id: 3E57A680-F248-11E8-B48F-1D18A9856A87
last_name: Novarino
orcid: 0000-0002-7673-7178
- first_name: Hanspeter
full_name: Pfister, Hanspeter
last_name: Pfister
- first_name: Bernd
full_name: Bickel, Bernd
id: 49876194-F248-11E8-B48F-1D18A9856A87
last_name: Bickel
orcid: 0000-0001-6511-9385
- first_name: Johann G
full_name: Danzl, Johann G
id: 42EFD3B6-F248-11E8-B48F-1D18A9856A87
last_name: Danzl
orcid: 0000-0001-8559-3973
citation:
ama: Velicky P, Miguel Villalba E, Michalska JM, et al. Dense 4D nanoscale reconstruction
of living brain tissue. Nature Methods. 2023;20:1256-1265. doi:10.1038/s41592-023-01936-6
apa: Velicky, P., Miguel Villalba, E., Michalska, J. M., Lyudchik, J., Wei, D.,
Lin, Z., … Danzl, J. G. (2023). Dense 4D nanoscale reconstruction of living brain
tissue. Nature Methods. Springer Nature. https://doi.org/10.1038/s41592-023-01936-6
chicago: Velicky, Philipp, Eder Miguel Villalba, Julia M Michalska, Julia Lyudchik,
Donglai Wei, Zudi Lin, Jake Watson, et al. “Dense 4D Nanoscale Reconstruction
of Living Brain Tissue.” Nature Methods. Springer Nature, 2023. https://doi.org/10.1038/s41592-023-01936-6.
ieee: P. Velicky et al., “Dense 4D nanoscale reconstruction of living brain
tissue,” Nature Methods, vol. 20. Springer Nature, pp. 1256–1265, 2023.
ista: Velicky P, Miguel Villalba E, Michalska JM, Lyudchik J, Wei D, Lin Z, Watson
J, Troidl J, Beyer J, Ben Simon Y, Sommer CM, Jahr W, Cenameri A, Broichhagen
J, Grant SGN, Jonas PM, Novarino G, Pfister H, Bickel B, Danzl JG. 2023. Dense
4D nanoscale reconstruction of living brain tissue. Nature Methods. 20, 1256–1265.
mla: Velicky, Philipp, et al. “Dense 4D Nanoscale Reconstruction of Living Brain
Tissue.” Nature Methods, vol. 20, Springer Nature, 2023, pp. 1256–65, doi:10.1038/s41592-023-01936-6.
short: P. Velicky, E. Miguel Villalba, J.M. Michalska, J. Lyudchik, D. Wei, Z. Lin,
J. Watson, J. Troidl, J. Beyer, Y. Ben Simon, C.M. Sommer, W. Jahr, A. Cenameri,
J. Broichhagen, S.G.N. Grant, P.M. Jonas, G. Novarino, H. Pfister, B. Bickel,
J.G. Danzl, Nature Methods 20 (2023) 1256–1265.
date_created: 2023-07-23T22:01:13Z
date_published: 2023-08-01T00:00:00Z
date_updated: 2024-01-10T08:37:48Z
day: '01'
department:
- _id: PeJo
- _id: GaNo
- _id: BeBi
- _id: JoDa
- _id: Bio
doi: 10.1038/s41592-023-01936-6
ec_funded: 1
external_id:
isi:
- '001025621500001'
pmid:
- '37429995'
intvolume: ' 20'
isi: 1
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41592-023-01936-6
month: '08'
oa: 1
oa_version: Published Version
page: 1256-1265
pmid: 1
project:
- _id: 265CB4D0-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: I03600
name: Optical control of synaptic function via adhesion molecules
- _id: 2548AE96-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: W1232-B24
name: Molecular Drug Targets
- _id: 25C5A090-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: Z00312
name: The Wittgenstein Prize
- _id: 23889792-32DE-11EA-91FC-C7463DDC885E
name: High content imaging to decode human immune cell interactions in health and
allergic disease
- _id: 2564DBCA-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '665385'
name: International IST Doctoral Program
- _id: 24F9549A-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '715767'
name: 'MATERIALIZABLE: Intelligent fabrication-oriented Computational Design and
Modeling'
- _id: 25444568-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '715508'
name: Probing the Reversibility of Autism Spectrum Disorders by Employing in vivo
and in vitro Models
- _id: 25B7EB9E-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '692692'
name: Biophysics and circuit function of a giant cortical glumatergic synapse
- _id: fc2be41b-9c52-11eb-aca3-faa90aa144e9
call_identifier: H2020
grant_number: '101026635'
name: Synaptic computations of the hippocampal CA3 circuitry
- _id: 2668BFA0-B435-11E9-9278-68D0E5697425
grant_number: LT00057
name: High-speed 3D-nanoscopy to study the role of adhesion during 3D cell migration
publication: Nature Methods
publication_identifier:
eissn:
- 1548-7105
issn:
- 1548-7091
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
related_material:
link:
- relation: software
url: https://github.com/danzllab/LIONESS
record:
- id: '12817'
relation: research_data
status: public
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relation: shorter_version
status: public
scopus_import: '1'
status: public
title: Dense 4D nanoscale reconstruction of living brain tissue
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 20
year: '2023'
...