--- _id: '8532' abstract: - lang: eng text: The molecular anatomy of synapses defines their characteristics in transmission and plasticity. Precise measurements of the number and distribution of synaptic proteins are important for our understanding of synapse heterogeneity within and between brain regions. Freeze–fracture replica immunogold electron microscopy enables us to analyze them quantitatively on a two-dimensional membrane surface. Here, we introduce Darea software, which utilizes deep learning for analysis of replica images and demonstrate its usefulness for quick measurements of the pre- and postsynaptic areas, density and distribution of gold particles at synapses in a reproducible manner. We used Darea for comparing glutamate receptor and calcium channel distributions between hippocampal CA3-CA1 spine synapses on apical and basal dendrites, which differ in signaling pathways involved in synaptic plasticity. We found that apical synapses express a higher density of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors and a stronger increase of AMPA receptors with synaptic size, while basal synapses show a larger increase in N-methyl-D-aspartate (NMDA) receptors with size. Interestingly, AMPA and NMDA receptors are segregated within postsynaptic sites and negatively correlated in density among both apical and basal synapses. In the presynaptic sites, Cav2.1 voltage-gated calcium channels show similar densities in apical and basal synapses with distributions consistent with an exclusion zone model of calcium channel-release site topography. acknowledgement: "This research was funded by Austrian Academy of Sciences, DOC fellowship to D.K., European Research\r\nCouncil Advanced Grant 694539 and European Union Human Brain Project (HBP) SGA2 785907 to R.S.\r\nWe acknowledge Elena Hollergschwandtner for technical support." article_number: '6737' article_processing_charge: No article_type: original author: - first_name: David full_name: Kleindienst, David id: 42E121A4-F248-11E8-B48F-1D18A9856A87 last_name: Kleindienst - first_name: Jacqueline-Claire full_name: Montanaro-Punzengruber, Jacqueline-Claire id: 3786AB44-F248-11E8-B48F-1D18A9856A87 last_name: Montanaro-Punzengruber - first_name: Pradeep full_name: Bhandari, Pradeep id: 45EDD1BC-F248-11E8-B48F-1D18A9856A87 last_name: Bhandari orcid: 0000-0003-0863-4481 - first_name: Matthew J full_name: Case, Matthew J id: 44B7CA5A-F248-11E8-B48F-1D18A9856A87 last_name: Case - first_name: Yugo full_name: Fukazawa, Yugo last_name: Fukazawa - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 citation: ama: Kleindienst D, Montanaro-Punzengruber J-C, Bhandari P, Case MJ, Fukazawa Y, Shigemoto R. Deep learning-assisted high-throughput analysis of freeze-fracture replica images applied to glutamate receptors and calcium channels at hippocampal synapses. International Journal of Molecular Sciences. 2020;21(18). doi:10.3390/ijms21186737 apa: Kleindienst, D., Montanaro-Punzengruber, J.-C., Bhandari, P., Case, M. J., Fukazawa, Y., & Shigemoto, R. (2020). Deep learning-assisted high-throughput analysis of freeze-fracture replica images applied to glutamate receptors and calcium channels at hippocampal synapses. International Journal of Molecular Sciences. MDPI. https://doi.org/10.3390/ijms21186737 chicago: Kleindienst, David, Jacqueline-Claire Montanaro-Punzengruber, Pradeep Bhandari, Matthew J Case, Yugo Fukazawa, and Ryuichi Shigemoto. “Deep Learning-Assisted High-Throughput Analysis of Freeze-Fracture Replica Images Applied to Glutamate Receptors and Calcium Channels at Hippocampal Synapses.” International Journal of Molecular Sciences. MDPI, 2020. https://doi.org/10.3390/ijms21186737. ieee: D. Kleindienst, J.-C. Montanaro-Punzengruber, P. Bhandari, M. J. Case, Y. Fukazawa, and R. Shigemoto, “Deep learning-assisted high-throughput analysis of freeze-fracture replica images applied to glutamate receptors and calcium channels at hippocampal synapses,” International Journal of Molecular Sciences, vol. 21, no. 18. MDPI, 2020. ista: Kleindienst D, Montanaro-Punzengruber J-C, Bhandari P, Case MJ, Fukazawa Y, Shigemoto R. 2020. Deep learning-assisted high-throughput analysis of freeze-fracture replica images applied to glutamate receptors and calcium channels at hippocampal synapses. International Journal of Molecular Sciences. 21(18), 6737. mla: Kleindienst, David, et al. “Deep Learning-Assisted High-Throughput Analysis of Freeze-Fracture Replica Images Applied to Glutamate Receptors and Calcium Channels at Hippocampal Synapses.” International Journal of Molecular Sciences, vol. 21, no. 18, 6737, MDPI, 2020, doi:10.3390/ijms21186737. short: D. Kleindienst, J.-C. Montanaro-Punzengruber, P. Bhandari, M.J. Case, Y. Fukazawa, R. Shigemoto, International Journal of Molecular Sciences 21 (2020). date_created: 2020-09-20T22:01:35Z date_published: 2020-09-14T00:00:00Z date_updated: 2024-03-27T23:30:30Z day: '14' ddc: - '570' department: - _id: RySh doi: 10.3390/ijms21186737 ec_funded: 1 external_id: isi: - '000579945300001' file: - access_level: open_access checksum: 2e4f62f3cfe945b7391fc3070e5a289f content_type: application/pdf creator: dernst date_created: 2020-09-21T14:08:58Z date_updated: 2020-09-21T14:08:58Z file_id: '8551' file_name: 2020_JournMolecSciences_Kleindienst.pdf file_size: 5748456 relation: main_file success: 1 file_date_updated: 2020-09-21T14:08:58Z has_accepted_license: '1' intvolume: ' 21' isi: 1 issue: '18' language: - iso: eng month: '09' oa: 1 oa_version: Published Version 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: 25D32BC0-B435-11E9-9278-68D0E5697425 name: Mechanism of formation and maintenance of input side-dependent asymmetry in the hippocampus - _id: 26436750-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '785907' name: Human Brain Project Specific Grant Agreement 2 (HBP SGA 2) publication: International Journal of Molecular Sciences publication_identifier: eissn: - '14220067' issn: - '16616596' publication_status: published publisher: MDPI quality_controlled: '1' related_material: record: - id: '9562' relation: dissertation_contains status: public scopus_import: '1' status: public title: Deep learning-assisted high-throughput analysis of freeze-fracture replica images applied to glutamate receptors and calcium channels at hippocampal synapses 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: 21 year: '2020' ... --- _id: '51' abstract: - lang: eng text: Asymmetries have long been known about in the central nervous system. From gross anatomical differences, such as the presence of the parapineal organ in only one hemisphere of the developing zebrafish, to more subtle differences in activity between both hemispheres, as seen in freely roaming animals or human participants under PET and fMRI imaging analysis. The presence of asymmetries has been demonstrated to have huge behavioural implications, with their disruption often leading to the generation of neurological disorders, memory problems, changes in personality, and in an organism's health and well-being. For my Ph.D. work I aimed to tackle two important avenues of research. The first being the process of input-side dependency in the hippocampus, with the goal of finding a key gene responsible for its development (Gene X). The second project was to do with experience-induced laterality formation in the hippocampus. Specifically, how laterality in the synapse density of the CA1 stratum radiatum (s.r.) could be induced purely through environmental enrichment. Through unilateral tracer injections into the CA3, I was able to selectively measure the properties of synapses within the CA1 and investigate how they differed based upon which hemisphere the presynaptic neurone originated. Having found the existence of a previously unreported reversed (left-isomerism) i.v. mutant, through morpholocal examination of labelled terminals in the CA1 s.r., I aimed to elucidate a key gene responsible for the process of left or right determination of inputs to the CA1 s.r.. This work relates to the previous finding of input-side dependent asymmetry in the wild-type rodent, where the origin of the projecting neurone to the CA1 will determine the morphology of a synapse, to a greater degree than the hemisphere in which the projection terminates. Using left- and right-isomerism i.v. mice, in combination with whole genome sequence analysis, I highlight Ena/VASP-like (Evl) as a potential target for Gene X. In relation to this topic, I also highlight my work in the recently published paper of how knockout of PirB can lead to a lack of input-side dependency in the murine hippocampus. For the second question, I show that the environmental enrichment paradigm will lead to an asymmetry in the synapse densities in the hippocampus of mice. I also highlight that the nature of the enrichment is of less consequence than the process of enrichment itself. I demonstrate that the CA3 region will dramatically alter its projection targets, in relation to environmental stimulation, with the asymmetry in synaptic density, caused by enrichment, relying heavily on commissural fibres. I also highlight the vital importance of input-side dependent asymmetry, as a necessary component of experience-dependent laterality formation in the CA1 s.r.. However, my results suggest that it isn't the only cause, as there appears to be a CA1 dependent mechanism also at play. Upon further investigation, I highlight the significant, and highly important, finding that the changes seen in the CA1 s.r. were predominantly caused through projections from the left-CA3, with the right-CA3 having less involvement in this mechanism. alternative_title: - ISTA Thesis article_processing_charge: No author: - first_name: Matthew J full_name: Case, Matthew J id: 44B7CA5A-F248-11E8-B48F-1D18A9856A87 last_name: Case citation: ama: 'Case MJ. From the left to the right: A tale of asymmetries, environments, and hippocampal development. 2018. doi:10.15479/AT:ISTA:th_1032' apa: 'Case, M. J. (2018). From the left to the right: A tale of asymmetries, environments, and hippocampal development. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:th_1032' chicago: 'Case, Matthew J. “From the Left to the Right: A Tale of Asymmetries, Environments, and Hippocampal Development.” Institute of Science and Technology Austria, 2018. https://doi.org/10.15479/AT:ISTA:th_1032.' ieee: 'M. J. Case, “From the left to the right: A tale of asymmetries, environments, and hippocampal development,” Institute of Science and Technology Austria, 2018.' ista: 'Case MJ. 2018. From the left to the right: A tale of asymmetries, environments, and hippocampal development. Institute of Science and Technology Austria.' mla: 'Case, Matthew J. From the Left to the Right: A Tale of Asymmetries, Environments, and Hippocampal Development. Institute of Science and Technology Austria, 2018, doi:10.15479/AT:ISTA:th_1032.' short: 'M.J. Case, From the Left to the Right: A Tale of Asymmetries, Environments, and Hippocampal Development, Institute of Science and Technology Austria, 2018.' date_created: 2018-12-11T11:44:22Z date_published: 2018-06-27T00:00:00Z date_updated: 2023-09-07T12:39:22Z day: '27' ddc: - '571' - '576' degree_awarded: PhD department: - _id: RySh doi: 10.15479/AT:ISTA:th_1032 file: - access_level: closed checksum: dcc7b55619d8509dd62b8e99d6cdee44 content_type: application/msword creator: dernst date_created: 2019-04-09T07:16:26Z date_updated: 2021-02-11T23:30:13Z embargo_to: open_access file_id: '6251' file_name: 2018_Thesis_Case_Source.doc file_size: 141270528 relation: source_file - access_level: open_access checksum: f69fdd5c8709c4e618aa8c1a1221153d content_type: application/pdf creator: dernst date_created: 2019-04-09T07:16:23Z date_updated: 2021-02-11T11:17:14Z embargo: 2019-07-05 file_id: '6252' file_name: 2018_Thesis_Case.pdf file_size: 15193621 relation: main_file file_date_updated: 2021-02-11T23:30:13Z has_accepted_license: '1' language: - iso: eng month: '06' oa: 1 oa_version: Published Version page: '186' publication_identifier: issn: - 2663-337X publication_status: published publisher: Institute of Science and Technology Austria publist_id: '8003' pubrep_id: '1032' related_material: record: - id: '682' 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: 'From the left to the right: A tale of asymmetries, environments, and hippocampal development' type: dissertation user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 year: '2018' ... --- _id: '682' abstract: - lang: eng text: Left-right asymmetry is a fundamental feature of higher-order brain structure; however, the molecular basis of brain asymmetry remains unclear. We recently identified structural and functional asymmetries in mouse hippocampal circuitry that result from the asymmetrical distribution of two distinct populations of pyramidal cell synapses that differ in the density of the NMDA receptor subunit GluRε2 (also known as NR2B, GRIN2B or GluN2B). By examining the synaptic distribution of ε2 subunits, we previously found that β2-microglobulin-deficient mice, which lack cell surface expression of the vast majority of major histocompatibility complex class I (MHCI) proteins, do not exhibit circuit asymmetry. In the present study, we conducted electrophysiological and anatomical analyses on the hippocampal circuitry of mice with a knockout of the paired immunoglobulin-like receptor B (PirB), an MHCI receptor. As in β2-microglobulin-deficient mice, the PirB-deficient hippocampus lacked circuit asymmetries. This finding that MHCI loss-of-function mice and PirB knockout mice have identical phenotypes suggests that MHCI signals that produce hippocampal asymmetries are transduced through PirB. Our results provide evidence for a critical role of the MHCI/PirB signaling system in the generation of asymmetries in hippocampal circuitry. article_number: e0179377 article_type: original author: - first_name: Hikari full_name: Ukai, Hikari last_name: Ukai - first_name: Aiko full_name: Kawahara, Aiko last_name: Kawahara - first_name: Keiko full_name: Hirayama, Keiko last_name: Hirayama - first_name: Matthew J full_name: Case, Matthew J id: 44B7CA5A-F248-11E8-B48F-1D18A9856A87 last_name: Case - first_name: Shotaro full_name: Aino, Shotaro last_name: Aino - first_name: Masahiro full_name: Miyabe, Masahiro last_name: Miyabe - first_name: Ken full_name: Wakita, Ken last_name: Wakita - first_name: Ryohei full_name: Oogi, Ryohei last_name: Oogi - first_name: Michiyo full_name: Kasayuki, Michiyo last_name: Kasayuki - first_name: Shihomi full_name: Kawashima, Shihomi last_name: Kawashima - first_name: Shunichi full_name: Sugimoto, Shunichi last_name: Sugimoto - first_name: Kanako full_name: Chikamatsu, Kanako last_name: Chikamatsu - first_name: Noritaka full_name: Nitta, Noritaka last_name: Nitta - first_name: Tsuneyuki full_name: Koga, Tsuneyuki last_name: Koga - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Toshiyuki full_name: Takai, Toshiyuki last_name: Takai - first_name: Isao full_name: Ito, Isao last_name: Ito citation: ama: Ukai H, Kawahara A, Hirayama K, et al. PirB regulates asymmetries in hippocampal circuitry. PLoS One. 2017;12(6). doi:10.1371/journal.pone.0179377 apa: Ukai, H., Kawahara, A., Hirayama, K., Case, M. J., Aino, S., Miyabe, M., … Ito, I. (2017). PirB regulates asymmetries in hippocampal circuitry. PLoS One. Public Library of Science. https://doi.org/10.1371/journal.pone.0179377 chicago: Ukai, Hikari, Aiko Kawahara, Keiko Hirayama, Matthew J Case, Shotaro Aino, Masahiro Miyabe, Ken Wakita, et al. “PirB Regulates Asymmetries in Hippocampal Circuitry.” PLoS One. Public Library of Science, 2017. https://doi.org/10.1371/journal.pone.0179377. ieee: H. Ukai et al., “PirB regulates asymmetries in hippocampal circuitry,” PLoS One, vol. 12, no. 6. Public Library of Science, 2017. ista: Ukai H, Kawahara A, Hirayama K, Case MJ, Aino S, Miyabe M, Wakita K, Oogi R, Kasayuki M, Kawashima S, Sugimoto S, Chikamatsu K, Nitta N, Koga T, Shigemoto R, Takai T, Ito I. 2017. PirB regulates asymmetries in hippocampal circuitry. PLoS One. 12(6), e0179377. mla: Ukai, Hikari, et al. “PirB Regulates Asymmetries in Hippocampal Circuitry.” PLoS One, vol. 12, no. 6, e0179377, Public Library of Science, 2017, doi:10.1371/journal.pone.0179377. short: H. Ukai, A. Kawahara, K. Hirayama, M.J. Case, S. Aino, M. Miyabe, K. Wakita, R. Oogi, M. Kasayuki, S. Kawashima, S. Sugimoto, K. Chikamatsu, N. Nitta, T. Koga, R. Shigemoto, T. Takai, I. Ito, PLoS One 12 (2017). date_created: 2018-12-11T11:47:54Z date_published: 2017-06-01T00:00:00Z date_updated: 2024-03-27T23:30:12Z day: '01' ddc: - '571' department: - _id: RySh doi: 10.1371/journal.pone.0179377 file: - access_level: open_access checksum: 24dd19c46fb1c761b0bcbbcd1025a3a8 content_type: application/pdf creator: system date_created: 2018-12-12T10:12:16Z date_updated: 2020-07-14T12:47:40Z file_id: '4934' file_name: IST-2017-897-v1+1_journal.pone.0179377.pdf file_size: 5798454 relation: main_file file_date_updated: 2020-07-14T12:47:40Z has_accepted_license: '1' intvolume: ' 12' issue: '6' language: - iso: eng month: '06' oa: 1 oa_version: Published Version publication: PLoS One publication_identifier: issn: - '19326203' publication_status: published publisher: Public Library of Science publist_id: '7034' pubrep_id: '897' quality_controlled: '1' related_material: record: - id: '51' relation: dissertation_contains status: public scopus_import: 1 status: public title: PirB regulates asymmetries in hippocampal circuitry 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: 12 year: '2017' ...