--- _id: '10403' abstract: - lang: eng text: Synaptic transmission, connectivity, and dendritic morphology mature in parallel during brain development and are often disrupted in neurodevelopmental disorders. Yet how these changes influence the neuronal computations necessary for normal brain function are not well understood. To identify cellular mechanisms underlying the maturation of synaptic integration in interneurons, we combined patch-clamp recordings of excitatory inputs in mouse cerebellar stellate cells (SCs), three-dimensional reconstruction of SC morphology with excitatory synapse location, and biophysical modeling. We found that postnatal maturation of postsynaptic strength was homogeneously reduced along the somatodendritic axis, but dendritic integration was always sublinear. However, dendritic branching increased without changes in synapse density, leading to a substantial gain in distal inputs. Thus, changes in synapse distribution, rather than dendrite cable properties, are the dominant mechanism underlying the maturation of neuronal computation. These mechanisms favor the emergence of a spatially compartmentalized two-stage integration model promoting location-dependent integration within dendritic subunits. acknowledgement: This study was supported by the Centre National de la Recherche Scientifique and the Agence Nationale de la Recherche (ANR-13-BSV4-00166, to LC and DAD). TA was supported by fellowships from the Fondation pour la Recherche Medicale and the Swedish Research Council. We thank Dmitry Ershov from the Image Analysis Hub of the Institut Pasteur, Elodie Le Monnier, Elena Hollergschwandtner, Vanessa Zheden, and Corinne Nantet for technical support and Haining Zhong for providing the Venus-tagged PSD95 mouse line. We would like to thank Alberto Bacci, Ann Lohof, and Nelson Rebola for comments on the manuscript. article_number: e65954 article_processing_charge: No article_type: original author: - first_name: Celia full_name: Biane, Celia last_name: Biane - first_name: Florian full_name: Rückerl, Florian last_name: Rückerl - first_name: Therese full_name: Abrahamsson, Therese last_name: Abrahamsson - first_name: Cécile full_name: Saint-Cloment, Cécile last_name: Saint-Cloment - first_name: Jean full_name: Mariani, Jean last_name: Mariani - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: David A. full_name: Digregorio, David A. last_name: Digregorio - first_name: Rachel M. full_name: Sherrard, Rachel M. last_name: Sherrard - first_name: Laurence full_name: Cathala, Laurence last_name: Cathala citation: ama: Biane C, Rückerl F, Abrahamsson T, et al. Developmental emergence of two-stage nonlinear synaptic integration in cerebellar interneurons. eLife. 2021;10. doi:10.7554/eLife.65954 apa: Biane, C., Rückerl, F., Abrahamsson, T., Saint-Cloment, C., Mariani, J., Shigemoto, R., … Cathala, L. (2021). Developmental emergence of two-stage nonlinear synaptic integration in cerebellar interneurons. ELife. eLife Sciences Publications. https://doi.org/10.7554/eLife.65954 chicago: Biane, Celia, Florian Rückerl, Therese Abrahamsson, Cécile Saint-Cloment, Jean Mariani, Ryuichi Shigemoto, David A. Digregorio, Rachel M. Sherrard, and Laurence Cathala. “Developmental Emergence of Two-Stage Nonlinear Synaptic Integration in Cerebellar Interneurons.” ELife. eLife Sciences Publications, 2021. https://doi.org/10.7554/eLife.65954. ieee: C. Biane et al., “Developmental emergence of two-stage nonlinear synaptic integration in cerebellar interneurons,” eLife, vol. 10. eLife Sciences Publications, 2021. ista: Biane C, Rückerl F, Abrahamsson T, Saint-Cloment C, Mariani J, Shigemoto R, Digregorio DA, Sherrard RM, Cathala L. 2021. Developmental emergence of two-stage nonlinear synaptic integration in cerebellar interneurons. eLife. 10, e65954. mla: Biane, Celia, et al. “Developmental Emergence of Two-Stage Nonlinear Synaptic Integration in Cerebellar Interneurons.” ELife, vol. 10, e65954, eLife Sciences Publications, 2021, doi:10.7554/eLife.65954. short: C. Biane, F. Rückerl, T. Abrahamsson, C. Saint-Cloment, J. Mariani, R. Shigemoto, D.A. Digregorio, R.M. Sherrard, L. Cathala, ELife 10 (2021). date_created: 2021-12-05T23:01:40Z date_published: 2021-11-03T00:00:00Z date_updated: 2023-08-14T13:12:07Z day: '03' ddc: - '570' department: - _id: RySh doi: 10.7554/eLife.65954 external_id: isi: - '000715789500001' file: - access_level: open_access checksum: c7c33c3319428d56e332e22349c50ed3 content_type: application/pdf creator: cchlebak date_created: 2021-12-10T08:31:41Z date_updated: 2021-12-10T08:31:41Z file_id: '10528' file_name: 2021_eLife_Biane.pdf file_size: 13131322 relation: main_file success: 1 file_date_updated: 2021-12-10T08:31:41Z has_accepted_license: '1' intvolume: ' 10' isi: 1 language: - iso: eng month: '11' oa: 1 oa_version: Published Version publication: eLife publication_identifier: eissn: - 2050-084X publication_status: published publisher: eLife Sciences Publications quality_controlled: '1' scopus_import: '1' status: public title: Developmental emergence of two-stage nonlinear synaptic integration in cerebellar interneurons 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: 10 year: '2021' ... --- _id: '9756' abstract: - lang: eng text: High-resolution visualization and quantification of membrane proteins contribute to the understanding of their functions and the roles they play in physiological and pathological conditions. Sodium dodecyl sulfate-digested freeze-fracture replica labeling (SDS-FRL) is a powerful electron microscopy method to study quantitatively the two-dimensional distribution of transmembrane proteins and their tightly associated proteins. During treatment with SDS, intracellular organelles and proteins not anchored to the replica are dissolved, whereas integral membrane proteins captured and stabilized by carbon/platinum deposition remain on the replica. Their intra- and extracellular domains become exposed on the surface of the replica, facilitating the accessibility of antibodies and, therefore, providing higher labeling efficiency than those obtained with other immunoelectron microscopy techniques. In this chapter, we describe the protocols of SDS-FRL adapted for mammalian brain samples, and optimization of the SDS treatment to increase the labeling efficiency for quantification of Cav2.1, the alpha subunit of P/Q-type voltage-dependent calcium channels utilizing deep learning algorithms. acknowledgement: This work was supported by the European Union (European Research Council Advanced grant no. 694539 and Human Brain Project Ref. 720270 to R. S.) and the Austrian Academy of Sciences (DOC fellowship to D.K.). alternative_title: - Neuromethods article_processing_charge: No author: - first_name: Walter full_name: Kaufmann, Walter id: 3F99E422-F248-11E8-B48F-1D18A9856A87 last_name: Kaufmann orcid: 0000-0001-9735-5315 - first_name: David full_name: Kleindienst, David id: 42E121A4-F248-11E8-B48F-1D18A9856A87 last_name: Kleindienst - first_name: Harumi full_name: Harada, Harumi id: 2E55CDF2-F248-11E8-B48F-1D18A9856A87 last_name: Harada orcid: 0000-0001-7429-7896 - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 citation: ama: 'Kaufmann W, Kleindienst D, Harada H, Shigemoto R. High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In: Receptor and Ion Channel Detection in the Brain. Vol 169. Neuromethods. New York: Humana; 2021:267-283. doi:10.1007/978-1-0716-1522-5_19' apa: 'Kaufmann, W., Kleindienst, D., Harada, H., & Shigemoto, R. (2021). High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In Receptor and Ion Channel Detection in the Brain (Vol. 169, pp. 267–283). New York: Humana. https://doi.org/10.1007/978-1-0716-1522-5_19' chicago: 'Kaufmann, Walter, David Kleindienst, Harumi Harada, and Ryuichi Shigemoto. “High-Resolution Localization and Quantitation of Membrane Proteins by SDS-Digested Freeze-Fracture Replica Labeling (SDS-FRL).” In Receptor and Ion Channel Detection in the Brain, 169:267–83. Neuromethods. New York: Humana, 2021. https://doi.org/10.1007/978-1-0716-1522-5_19.' ieee: 'W. Kaufmann, D. Kleindienst, H. Harada, and R. Shigemoto, “High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL),” in Receptor and Ion Channel Detection in the Brain, vol. 169, New York: Humana, 2021, pp. 267–283.' ista: 'Kaufmann W, Kleindienst D, Harada H, Shigemoto R. 2021.High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In: Receptor and Ion Channel Detection in the Brain. Neuromethods, vol. 169, 267–283.' mla: Kaufmann, Walter, et al. “High-Resolution Localization and Quantitation of Membrane Proteins by SDS-Digested Freeze-Fracture Replica Labeling (SDS-FRL).” Receptor and Ion Channel Detection in the Brain, vol. 169, Humana, 2021, pp. 267–83, doi:10.1007/978-1-0716-1522-5_19. short: W. Kaufmann, D. Kleindienst, H. Harada, R. Shigemoto, in:, Receptor and Ion Channel Detection in the Brain, Humana, New York, 2021, pp. 267–283. date_created: 2021-07-30T09:34:56Z date_published: 2021-07-27T00:00:00Z date_updated: 2024-03-25T23:30:16Z day: '27' ddc: - '573' department: - _id: RySh - _id: EM-Fac doi: 10.1007/978-1-0716-1522-5_19 ec_funded: 1 has_accepted_license: '1' intvolume: ' 169' keyword: - 'Freeze-fracture replica: Deep learning' - Immunogold labeling - Integral membrane protein - Electron microscopy language: - iso: eng month: '07' oa_version: None page: 267-283 place: New York 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: ' Receptor and Ion Channel Detection in the Brain' publication_identifier: eisbn: - '9781071615225' isbn: - '9781071615218' publication_status: published publisher: Humana quality_controlled: '1' related_material: record: - id: '9562' relation: dissertation_contains status: public series_title: Neuromethods status: public title: High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL) type: book_chapter user_id: D865714E-FA4E-11E9-B85B-F5C5E5697425 volume: 169 year: '2021' ... --- _id: '7878' abstract: - lang: eng text: Type 1 metabotropic glutamate receptors (mGluR1s) are key elements in neuronal signaling. While their function is well documented in slices, requirements for their activation in vivo are poorly understood. We examine this question in adult mice in vivo using 2-photon imaging of cerebellar molecular layer interneurons (MLIs) expressing GCaMP. In anesthetized mice, parallel fiber activation evokes beam-like Cai rises in postsynaptic MLIs which depend on co-activation of mGluR1s and ionotropic glutamate receptors (iGluRs). In awake mice, blocking mGluR1 decreases Cai rises associated with locomotion. In vitro studies and freeze-fracture electron microscopy show that the iGluR-mGluR1 interaction is synergistic and favored by close association of the two classes of receptors. Altogether our results suggest that mGluR1s, acting in synergy with iGluRs, potently contribute to processing cerebellar neuronal signaling under physiological conditions. article_number: e56839 article_processing_charge: No article_type: original author: - first_name: Jin full_name: Bao, Jin last_name: Bao - first_name: Michael full_name: Graupner, Michael last_name: Graupner - first_name: Guadalupe full_name: Astorga, Guadalupe last_name: Astorga - first_name: Thibault full_name: Collin, Thibault last_name: Collin - first_name: Abdelali full_name: Jalil, Abdelali last_name: Jalil - first_name: Dwi Wahyu full_name: Indriati, Dwi Wahyu last_name: Indriati - first_name: Jonathan full_name: Bradley, Jonathan last_name: Bradley - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Isabel full_name: Llano, Isabel last_name: Llano citation: ama: Bao J, Graupner M, Astorga G, et al. Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo. eLife. 2020;9. doi:10.7554/eLife.56839 apa: Bao, J., Graupner, M., Astorga, G., Collin, T., Jalil, A., Indriati, D. W., … Llano, I. (2020). Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo. ELife. eLife Sciences Publications. https://doi.org/10.7554/eLife.56839 chicago: Bao, Jin, Michael Graupner, Guadalupe Astorga, Thibault Collin, Abdelali Jalil, Dwi Wahyu Indriati, Jonathan Bradley, Ryuichi Shigemoto, and Isabel Llano. “Synergism of Type 1 Metabotropic and Ionotropic Glutamate Receptors in Cerebellar Molecular Layer Interneurons in Vivo.” ELife. eLife Sciences Publications, 2020. https://doi.org/10.7554/eLife.56839. ieee: J. Bao et al., “Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo,” eLife, vol. 9. eLife Sciences Publications, 2020. ista: Bao J, Graupner M, Astorga G, Collin T, Jalil A, Indriati DW, Bradley J, Shigemoto R, Llano I. 2020. Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo. eLife. 9, e56839. mla: Bao, Jin, et al. “Synergism of Type 1 Metabotropic and Ionotropic Glutamate Receptors in Cerebellar Molecular Layer Interneurons in Vivo.” ELife, vol. 9, e56839, eLife Sciences Publications, 2020, doi:10.7554/eLife.56839. short: J. Bao, M. Graupner, G. Astorga, T. Collin, A. Jalil, D.W. Indriati, J. Bradley, R. Shigemoto, I. Llano, ELife 9 (2020). date_created: 2020-05-24T22:00:58Z date_published: 2020-05-13T00:00:00Z date_updated: 2023-08-21T06:26:50Z day: '13' ddc: - '570' department: - _id: RySh doi: 10.7554/eLife.56839 external_id: isi: - '000535191600001' pmid: - '32401196' file: - access_level: open_access checksum: 8ea99bb6660cc407dbdb00c173b01683 content_type: application/pdf creator: dernst date_created: 2020-05-26T09:34:54Z date_updated: 2020-07-14T12:48:04Z file_id: '7891' file_name: 2020_eLife_Bao.pdf file_size: 4832050 relation: main_file file_date_updated: 2020-07-14T12:48:04Z has_accepted_license: '1' intvolume: ' 9' isi: 1 language: - iso: eng month: '05' oa: 1 oa_version: Published Version pmid: 1 publication: eLife publication_identifier: eissn: - 2050084X publication_status: published publisher: eLife Sciences Publications quality_controlled: '1' scopus_import: '1' status: public title: Synergism of type 1 metabotropic and ionotropic glutamate receptors in cerebellar molecular layer interneurons in vivo 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: 9 year: '2020' ... --- _id: '7908' abstract: - lang: eng text: Volatile anesthetics are widely used for surgery, but neuronal mechanisms of anesthesia remain unidentified. At the calyx of Held in brainstem slices from rats of either sex, isoflurane at clinical doses attenuated EPSCs by decreasing the release probability and the number of readily releasable vesicles. In presynaptic recordings of Ca2+ currents and exocytic capacitance changes, isoflurane attenuated exocytosis by inhibiting Ca2+ currents evoked by a short presynaptic depolarization, whereas it inhibited exocytosis evoked by a prolonged depolarization via directly blocking exocytic machinery downstream of Ca2+ influx. Since the length of presynaptic depolarization can simulate the frequency of synaptic inputs, isoflurane anesthesia is likely mediated by distinct dual mechanisms, depending on input frequencies. In simultaneous presynaptic and postsynaptic action potential recordings, isoflurane impaired the fidelity of repetitive spike transmission, more strongly at higher frequencies. Furthermore, in the cerebrum of adult mice, isoflurane inhibited monosynaptic corticocortical spike transmission, preferentially at a higher frequency. We conclude that dual presynaptic mechanisms operate for the anesthetic action of isoflurane, of which direct inhibition of exocytic machinery plays a low-pass filtering role in spike transmission at central excitatory synapses. article_processing_charge: No article_type: original author: - first_name: Han Ying full_name: Wang, Han Ying last_name: Wang - first_name: Kohgaku full_name: Eguchi, Kohgaku id: 2B7846DC-F248-11E8-B48F-1D18A9856A87 last_name: Eguchi orcid: 0000-0002-6170-2546 - first_name: Takayuki full_name: Yamashita, Takayuki last_name: Yamashita - first_name: Tomoyuki full_name: Takahashi, Tomoyuki last_name: Takahashi citation: ama: Wang HY, Eguchi K, Yamashita T, Takahashi T. Frequency-dependent block of excitatory neurotransmission by isoflurane via dual presynaptic mechanisms. Journal of Neuroscience. 2020;40(21):4103-4115. doi:10.1523/JNEUROSCI.2946-19.2020 apa: Wang, H. Y., Eguchi, K., Yamashita, T., & Takahashi, T. (2020). Frequency-dependent block of excitatory neurotransmission by isoflurane via dual presynaptic mechanisms. Journal of Neuroscience. Society for Neuroscience. https://doi.org/10.1523/JNEUROSCI.2946-19.2020 chicago: Wang, Han Ying, Kohgaku Eguchi, Takayuki Yamashita, and Tomoyuki Takahashi. “Frequency-Dependent Block of Excitatory Neurotransmission by Isoflurane via Dual Presynaptic Mechanisms.” Journal of Neuroscience. Society for Neuroscience, 2020. https://doi.org/10.1523/JNEUROSCI.2946-19.2020. ieee: H. Y. Wang, K. Eguchi, T. Yamashita, and T. Takahashi, “Frequency-dependent block of excitatory neurotransmission by isoflurane via dual presynaptic mechanisms,” Journal of Neuroscience, vol. 40, no. 21. Society for Neuroscience, pp. 4103–4115, 2020. ista: Wang HY, Eguchi K, Yamashita T, Takahashi T. 2020. Frequency-dependent block of excitatory neurotransmission by isoflurane via dual presynaptic mechanisms. Journal of Neuroscience. 40(21), 4103–4115. mla: Wang, Han Ying, et al. “Frequency-Dependent Block of Excitatory Neurotransmission by Isoflurane via Dual Presynaptic Mechanisms.” Journal of Neuroscience, vol. 40, no. 21, Society for Neuroscience, 2020, pp. 4103–15, doi:10.1523/JNEUROSCI.2946-19.2020. short: H.Y. Wang, K. Eguchi, T. Yamashita, T. Takahashi, Journal of Neuroscience 40 (2020) 4103–4115. date_created: 2020-05-31T22:00:48Z date_published: 2020-05-20T00:00:00Z date_updated: 2023-08-21T06:31:25Z day: '20' ddc: - '570' department: - _id: RySh doi: 10.1523/JNEUROSCI.2946-19.2020 external_id: isi: - '000535694700004' file: - access_level: open_access checksum: 6571607ea9036154b67cc78e848a7f7d content_type: application/pdf creator: dernst date_created: 2020-06-02T09:12:16Z date_updated: 2020-07-14T12:48:05Z file_id: '7912' file_name: 2020_JourNeuroscience_Wang.pdf file_size: 3817360 relation: main_file file_date_updated: 2020-07-14T12:48:05Z has_accepted_license: '1' intvolume: ' 40' isi: 1 issue: '21' language: - iso: eng month: '05' oa: 1 oa_version: Published Version page: 4103-4115 publication: Journal of Neuroscience publication_identifier: eissn: - '15292401' publication_status: published publisher: Society for Neuroscience quality_controlled: '1' scopus_import: '1' status: public title: Frequency-dependent block of excitatory neurotransmission by isoflurane via dual presynaptic mechanisms 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: 40 year: '2020' ... --- _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: 2024-03-25T23:30:16Z 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: '7148' abstract: - lang: eng text: In the cerebellum, GluD2 is exclusively expressed in Purkinje cells, where it regulates synapse formation and regeneration, synaptic plasticity, and motor learning. Delayed cognitive development in humans with GluD2 gene mutations suggests extracerebellar functions of GluD2. However, extracerebellar expression of GluD2 and its relationship with that of GluD1 are poorly understood. GluD2 mRNA and protein were widely detected, with relatively high levels observed in the olfactory glomerular layer, medial prefrontal cortex, cingulate cortex, retrosplenial granular cortex, olfactory tubercle, subiculum, striatum, lateral septum, anterodorsal thalamic nucleus, and arcuate hypothalamic nucleus. These regions were also enriched for GluD1, and many individual neurons coexpressed the two GluDs. In the retrosplenial granular cortex, GluD1 and GluD2 were selectively expressed at PSD‐95‐expressing glutamatergic synapses, and their coexpression on the same synapses was shown by SDS‐digested freeze‐fracture replica labeling. Biochemically, GluD1 and GluD2 formed coimmunoprecipitable complex formation in HEK293T cells and in the cerebral cortex and hippocampus. We further estimated the relative protein amount by quantitative immunoblotting using GluA2/GluD2 and GluA2/GluD1 chimeric proteins as standards for titration of GluD1 and GluD2 antibodies. Intriguingly, the relative amount of GluD2 was almost comparable to that of GluD1 in the postsynaptic density fraction prepared from the cerebral cortex and hippocampus. In contrast, GluD2 was overwhelmingly predominant in the cerebellum. Thus, we have determined the relative extracerebellar expression of GluD1 and GluD2 at regional, neuronal, and synaptic levels. These data provide a molecular–anatomical basis for possible competitive and cooperative interactions of GluD family members at synapses in various brain regions. acknowledgement: This study was supported by Grants-in-Aid for Scientific Research to K.K. (18K06813), Y.M. (17K08503, 17H0631319), and K.S. (16H04650) and a grant for Scientific Research on Innovative Areas to K.S (16H06276) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT). We thank K. Akashi, I. Watanabe-Iida, Y. Suzuki, and H. Azechi for technical assistance and advice, and H. Uchida for valuable discussions. We thank E. Kushiya,I. Yabe, C. Ohori, Y. Mochizuki, Y. Ishikawa, and N. Ishimoto for technical assistance in generating GluD1-KO mice. article_processing_charge: No article_type: original author: - first_name: Chihiro full_name: Nakamoto, Chihiro last_name: Nakamoto - first_name: Kohtarou full_name: Konno, Kohtarou last_name: Konno - first_name: Taisuke full_name: Miyazaki, Taisuke last_name: Miyazaki - first_name: Ena full_name: Nakatsukasa, Ena last_name: Nakatsukasa - first_name: Rie full_name: Natsume, Rie last_name: Natsume - first_name: Manabu full_name: Abe, Manabu last_name: Abe - first_name: Meiko full_name: Kawamura, Meiko last_name: Kawamura - 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 - first_name: Miwako full_name: Yamasaki, Miwako last_name: Yamasaki - first_name: Kenji full_name: Sakimura, Kenji last_name: Sakimura - first_name: Masahiko full_name: Watanabe, Masahiko last_name: Watanabe citation: ama: Nakamoto C, Konno K, Miyazaki T, et al. Expression mapping, quantification, and complex formation of GluD1 and GluD2 glutamate receptors in adult mouse brain. Journal of Comparative Neurology. 2020;528(6):1003-1027. doi:10.1002/cne.24792 apa: Nakamoto, C., Konno, K., Miyazaki, T., Nakatsukasa, E., Natsume, R., Abe, M., … Watanabe, M. (2020). Expression mapping, quantification, and complex formation of GluD1 and GluD2 glutamate receptors in adult mouse brain. Journal of Comparative Neurology. Wiley. https://doi.org/10.1002/cne.24792 chicago: Nakamoto, Chihiro, Kohtarou Konno, Taisuke Miyazaki, Ena Nakatsukasa, Rie Natsume, Manabu Abe, Meiko Kawamura, et al. “Expression Mapping, Quantification, and Complex Formation of GluD1 and GluD2 Glutamate Receptors in Adult Mouse Brain.” Journal of Comparative Neurology. Wiley, 2020. https://doi.org/10.1002/cne.24792. ieee: C. Nakamoto et al., “Expression mapping, quantification, and complex formation of GluD1 and GluD2 glutamate receptors in adult mouse brain,” Journal of Comparative Neurology, vol. 528, no. 6. Wiley, pp. 1003–1027, 2020. ista: Nakamoto C, Konno K, Miyazaki T, Nakatsukasa E, Natsume R, Abe M, Kawamura M, Fukazawa Y, Shigemoto R, Yamasaki M, Sakimura K, Watanabe M. 2020. Expression mapping, quantification, and complex formation of GluD1 and GluD2 glutamate receptors in adult mouse brain. Journal of Comparative Neurology. 528(6), 1003–1027. mla: Nakamoto, Chihiro, et al. “Expression Mapping, Quantification, and Complex Formation of GluD1 and GluD2 Glutamate Receptors in Adult Mouse Brain.” Journal of Comparative Neurology, vol. 528, no. 6, Wiley, 2020, pp. 1003–27, doi:10.1002/cne.24792. short: C. Nakamoto, K. Konno, T. Miyazaki, E. Nakatsukasa, R. Natsume, M. Abe, M. Kawamura, Y. Fukazawa, R. Shigemoto, M. Yamasaki, K. Sakimura, M. Watanabe, Journal of Comparative Neurology 528 (2020) 1003–1027. date_created: 2019-12-04T16:09:29Z date_published: 2020-04-01T00:00:00Z date_updated: 2023-08-17T14:06:50Z day: '01' ddc: - '571' - '599' department: - _id: RySh doi: 10.1002/cne.24792 external_id: isi: - '000496410200001' pmid: - '31625608' has_accepted_license: '1' intvolume: ' 528' isi: 1 issue: '6' language: - iso: eng month: '04' oa_version: None page: 1003-1027 pmid: 1 publication: Journal of Comparative Neurology publication_identifier: eissn: - 1096-9861 issn: - 0021-9967 publication_status: published publisher: Wiley quality_controlled: '1' scopus_import: '1' status: public title: Expression mapping, quantification, and complex formation of GluD1 and GluD2 glutamate receptors in adult mouse brain type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 528 year: '2020' ... --- _id: '7207' abstract: - lang: eng text: The hippocampus plays key roles in learning and memory and is a main target of Alzheimer's disease (AD), which causes progressive memory impairments. Despite numerous investigations about the processes required for the normal hippocampal functions, the neurotransmitter receptors involved in the synaptic deficits by which AD disables the hippocampus are not yet characterized. By combining histoblots, western blots, immunohistochemistry and high‐resolution immunoelectron microscopic methods for GABAB receptors, this study provides a quantitative description of the expression and the subcellular localization of GABAB1 in the hippocampus in a mouse model of AD at 1, 6 and 12 months of age. Western blots and histoblots showed that the total amount of protein and the laminar expression pattern of GABAB1 were similar in APP/PS1 mice and in age‐matched wild‐type mice. In contrast, immunoelectron microscopic techniques showed that the subcellular localization of GABAB1 subunit did not change significantly in APP/PS1 mice at 1 month of age, was significantly reduced in the stratum lacunosum‐moleculare of CA1 pyramidal cells at 6 months of age and significantly reduced at the membrane surface of CA1 pyramidal cells at 12 months of age. This reduction of plasma membrane GABAB1 was paralleled by a significant increase of the subunit at the intracellular sites. We further observed a decrease of membrane‐targeted GABAB receptors in axon terminals contacting CA1 pyramidal cells. Our data demonstrate compartment‐ and age‐dependent reduction of plasma membrane‐targeted GABAB receptors in the CA1 region of the hippocampus, suggesting that this decrease might be enough to alter the GABAB‐mediated synaptic transmission taking place in AD. article_processing_charge: No article_type: original author: - first_name: Alejandro full_name: Martín-Belmonte, Alejandro last_name: Martín-Belmonte - first_name: Carolina full_name: Aguado, Carolina last_name: Aguado - first_name: Rocío full_name: Alfaro-Ruíz, Rocío last_name: Alfaro-Ruíz - first_name: Ana Esther full_name: Moreno-Martínez, Ana Esther last_name: Moreno-Martínez - first_name: Luis full_name: De La Ossa, Luis last_name: De La Ossa - first_name: José full_name: Martínez-Hernández, José last_name: Martínez-Hernández - first_name: Alain full_name: Buisson, Alain last_name: Buisson - first_name: Simon full_name: Früh, Simon last_name: Früh - first_name: Bernhard full_name: Bettler, Bernhard last_name: Bettler - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Yugo full_name: Fukazawa, Yugo last_name: Fukazawa - first_name: Rafael full_name: Luján, Rafael last_name: Luján citation: ama: Martín-Belmonte A, Aguado C, Alfaro-Ruíz R, et al. Reduction in the neuronal surface of post and presynaptic GABA>B< receptors in the hippocampus in a mouse model of Alzheimer’s disease. Brain Pathology. 2020;30(3):554-575. doi:10.1111/bpa.12802 apa: Martín-Belmonte, A., Aguado, C., Alfaro-Ruíz, R., Moreno-Martínez, A. E., De La Ossa, L., Martínez-Hernández, J., … Luján, R. (2020). Reduction in the neuronal surface of post and presynaptic GABA>B< receptors in the hippocampus in a mouse model of Alzheimer’s disease. Brain Pathology. Wiley. https://doi.org/10.1111/bpa.12802 chicago: Martín-Belmonte, Alejandro, Carolina Aguado, Rocío Alfaro-Ruíz, Ana Esther Moreno-Martínez, Luis De La Ossa, José Martínez-Hernández, Alain Buisson, et al. “Reduction in the Neuronal Surface of Post and Presynaptic GABA>B< Receptors in the Hippocampus in a Mouse Model of Alzheimer’s Disease.” Brain Pathology. Wiley, 2020. https://doi.org/10.1111/bpa.12802. ieee: A. Martín-Belmonte et al., “Reduction in the neuronal surface of post and presynaptic GABA>B< receptors in the hippocampus in a mouse model of Alzheimer’s disease,” Brain Pathology, vol. 30, no. 3. Wiley, pp. 554–575, 2020. ista: Martín-Belmonte A, Aguado C, Alfaro-Ruíz R, Moreno-Martínez AE, De La Ossa L, Martínez-Hernández J, Buisson A, Früh S, Bettler B, Shigemoto R, Fukazawa Y, Luján R. 2020. Reduction in the neuronal surface of post and presynaptic GABA>B< receptors in the hippocampus in a mouse model of Alzheimer’s disease. Brain Pathology. 30(3), 554–575. mla: Martín-Belmonte, Alejandro, et al. “Reduction in the Neuronal Surface of Post and Presynaptic GABA>B< Receptors in the Hippocampus in a Mouse Model of Alzheimer’s Disease.” Brain Pathology, vol. 30, no. 3, Wiley, 2020, pp. 554–75, doi:10.1111/bpa.12802. short: A. Martín-Belmonte, C. Aguado, R. Alfaro-Ruíz, A.E. Moreno-Martínez, L. De La Ossa, J. Martínez-Hernández, A. Buisson, S. Früh, B. Bettler, R. Shigemoto, Y. Fukazawa, R. Luján, Brain Pathology 30 (2020) 554–575. date_created: 2019-12-22T23:00:43Z date_published: 2020-05-01T00:00:00Z date_updated: 2023-09-06T14:48:01Z day: '01' ddc: - '570' department: - _id: RySh doi: 10.1111/bpa.12802 ec_funded: 1 external_id: isi: - '000502270900001' pmid: - '31729777' file: - access_level: open_access checksum: 549cc1b18f638a21d17a939ba5563fa9 content_type: application/pdf creator: dernst date_created: 2020-09-22T09:47:19Z date_updated: 2020-09-22T09:47:19Z file_id: '8554' file_name: 2020_BrainPathology_MartinBelmonte.pdf file_size: 4220935 relation: main_file success: 1 file_date_updated: 2020-09-22T09:47:19Z has_accepted_license: '1' intvolume: ' 30' isi: 1 issue: '3' language: - iso: eng month: '05' oa: 1 oa_version: Published Version page: 554-575 pmid: 1 project: - _id: 25CBA828-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '720270' name: Human Brain Project Specific Grant Agreement 1 (HBP SGA 1) - _id: 26436750-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '785907' name: Human Brain Project Specific Grant Agreement 2 (HBP SGA 2) publication: Brain Pathology publication_identifier: eissn: - '17503639' issn: - '10156305' publication_status: published publisher: Wiley quality_controlled: '1' scopus_import: '1' status: public title: Reduction in the neuronal surface of post and presynaptic GABA>B< receptors in the hippocampus in a mouse model of Alzheimer's disease 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: 30 year: '2020' ... --- _id: '7339' abstract: - lang: eng text: Cytoskeletal filaments such as microtubules (MTs) and filamentous actin (F-actin) dynamically support cell structure and functions. In central presynaptic terminals, F-actin is expressed along the release edge and reportedly plays diverse functional roles, but whether axonal MTs extend deep into terminals and play any physiological role remains controversial. At the calyx of Held in rats of either sex, confocal and high-resolution microscopy revealed that MTs enter deep into presynaptic terminal swellings and partially colocalize with a subset of synaptic vesicles (SVs). Electrophysiological analysis demonstrated that depolymerization of MTs specifically prolonged the slow-recovery time component of EPSCs from short-term depression induced by a train of high-frequency stimulation, whereas depolymerization of F-actin specifically prolonged the fast-recovery component. In simultaneous presynaptic and postsynaptic action potential recordings, depolymerization of MTs or F-actin significantly impaired the fidelity of high-frequency neurotransmission. We conclude that MTs and F-actin differentially contribute to slow and fast SV replenishment, thereby maintaining high-frequency neurotransmission. article_processing_charge: No article_type: original author: - first_name: Lashmi full_name: Piriya Ananda Babu, Lashmi last_name: Piriya Ananda Babu - first_name: Han Ying full_name: Wang, Han Ying last_name: Wang - first_name: Kohgaku full_name: Eguchi, Kohgaku id: 2B7846DC-F248-11E8-B48F-1D18A9856A87 last_name: Eguchi orcid: 0000-0002-6170-2546 - first_name: Laurent full_name: Guillaud, Laurent last_name: Guillaud - first_name: Tomoyuki full_name: Takahashi, Tomoyuki last_name: Takahashi citation: ama: Piriya Ananda Babu L, Wang HY, Eguchi K, Guillaud L, Takahashi T. Microtubule and actin differentially regulate synaptic vesicle cycling to maintain high-frequency neurotransmission. Journal of neuroscience. 2020;40(1):131-142. doi:10.1523/JNEUROSCI.1571-19.2019 apa: Piriya Ananda Babu, L., Wang, H. Y., Eguchi, K., Guillaud, L., & Takahashi, T. (2020). Microtubule and actin differentially regulate synaptic vesicle cycling to maintain high-frequency neurotransmission. Journal of Neuroscience. Society for Neuroscience. https://doi.org/10.1523/JNEUROSCI.1571-19.2019 chicago: Piriya Ananda Babu, Lashmi, Han Ying Wang, Kohgaku Eguchi, Laurent Guillaud, and Tomoyuki Takahashi. “Microtubule and Actin Differentially Regulate Synaptic Vesicle Cycling to Maintain High-Frequency Neurotransmission.” Journal of Neuroscience. Society for Neuroscience, 2020. https://doi.org/10.1523/JNEUROSCI.1571-19.2019. ieee: L. Piriya Ananda Babu, H. Y. Wang, K. Eguchi, L. Guillaud, and T. Takahashi, “Microtubule and actin differentially regulate synaptic vesicle cycling to maintain high-frequency neurotransmission,” Journal of neuroscience, vol. 40, no. 1. Society for Neuroscience, pp. 131–142, 2020. ista: Piriya Ananda Babu L, Wang HY, Eguchi K, Guillaud L, Takahashi T. 2020. Microtubule and actin differentially regulate synaptic vesicle cycling to maintain high-frequency neurotransmission. Journal of neuroscience. 40(1), 131–142. mla: Piriya Ananda Babu, Lashmi, et al. “Microtubule and Actin Differentially Regulate Synaptic Vesicle Cycling to Maintain High-Frequency Neurotransmission.” Journal of Neuroscience, vol. 40, no. 1, Society for Neuroscience, 2020, pp. 131–42, doi:10.1523/JNEUROSCI.1571-19.2019. short: L. Piriya Ananda Babu, H.Y. Wang, K. Eguchi, L. Guillaud, T. Takahashi, Journal of Neuroscience 40 (2020) 131–142. date_created: 2020-01-19T23:00:38Z date_published: 2020-01-02T00:00:00Z date_updated: 2023-08-17T14:25:23Z day: '02' ddc: - '570' department: - _id: RySh doi: 10.1523/JNEUROSCI.1571-19.2019 external_id: isi: - '000505167600013' pmid: - '31767677' file: - access_level: open_access checksum: 92f5e8a47f454fc131fb94cd7f106e60 content_type: application/pdf creator: dernst date_created: 2020-01-20T14:44:10Z date_updated: 2020-07-14T12:47:56Z file_id: '7345' file_name: 2020_JourNeuroscience_Piriya.pdf file_size: 4460781 relation: main_file file_date_updated: 2020-07-14T12:47:56Z has_accepted_license: '1' intvolume: ' 40' isi: 1 issue: '1' language: - iso: eng month: '01' oa: 1 oa_version: Published Version page: 131-142 pmid: 1 publication: Journal of neuroscience publication_identifier: eissn: - '15292401' publication_status: published publisher: Society for Neuroscience quality_controlled: '1' scopus_import: '1' status: public title: Microtubule and actin differentially regulate synaptic vesicle cycling to maintain high-frequency neurotransmission 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: 40 year: '2020' ... --- _id: '7525' abstract: - lang: eng text: "The medial habenula (MHb) is an evolutionary conserved epithalamic structure important for the modulation of emotional memory. It is involved in regulation of anxiety, compulsive behavior, addiction (nicotinic and opioid), sexual and feeding behavior. MHb receives inputs from septal regions and projects exclusively to the interpeduncular nucleus (IPN). Distinct sub-regions of the septum project to different subnuclei of MHb: the bed nucleus of anterior commissure projects to dorsal MHb and the triangular septum projects to ventral MHb. Furthermore, the dorsal and ventral MHb project to the lateral and rostral/central IPN, respectively. Importantly, these projections have unique features of prominent co-release of different neurotransmitters and requirement of a peculiar type of calcium channel for release. In general, synaptic neurotransmission requires an activity-dependent influx of Ca2+ into the presynaptic terminal through voltage-gated calcium channels. The calcium channel family most commonly involved in neurotransmitter release comprises three members, P/Q-, N- and R-type with Cav2.1, Cav2.2 and Cav2.3 subunits, respectively. In contrast to most CNS synapses that mainly express Cav2.1 and/or Cav2.2, MHb terminals in the IPN exclusively express Cav2.3. In other parts of the brain, such as the hippocampus, Cav2.3 is mostly located to postsynaptic elements. This unusual presynaptic location of Cav2.3 in the MHb-IPN pathway implies unique mechanisms of glutamate release in this pathway. One potential example of such uniqueness is the facilitation of release by GABAB receptor (GBR) activation. Presynaptic GBRs usually inhibit the release of neurotransmitters by inhibiting presynaptic calcium channels. MHb shows the highest expression levels of GBR in the brain. GBRs comprise two subunits, GABAB1 (GB1) and GABAB2 (GB2), and are associated with auxiliary subunits, called potassium channel tetramerization domain containing proteins (KCTD) 8, 12, 12b and 16. Among these four subunits, KCTD12b is exclusively expressed in ventral MHb, and KCTD8 shows the strongest expression in the whole MHb among other brain regions, indicating that KCTD8 and KCTD12b may be involved in the unique mechanisms of neurotransmitter release mediated by Cav2.3 and regulated by GBRs in this pathway. \r\nIn the present study, we first verified that neurotransmission in both dorsal and ventral MHb-IPN pathways is mainly mediated by Cav2.3 using a selective blocker of R-type channels, SNX-482. We next found that baclofen, a GBR agonist, has facilitatory effects on release from ventral MHb terminal in rostral IPN, whereas it has inhibitory effects on release from dorsal MHb terminals in lateral IPN, indicating that KCTD12b expressed exclusively in ventral MHb may have a role in the facilitatory effects of GBR activation. In a heterologous expression system using HEK cells, we found that KCTD8 and KCTD12b but not KCTD12 directly bind with Cav2.3. Pre-embedding immunogold electron microscopy data show that Cav2.3 and KCTD12b are distributed most densely in presynaptic active zone in IPN with KCTD12b being present only in rostral/central but not lateral IPN, whereas GABAB, KCTD8 and KCTD12 are distributed most densely in perisynaptic sites with KCTD12 present more frequently in postsynaptic elements and only in rostral/central IPN. In freeze-fracture replica labelling, Cav2.3, KCTD8 and KCTD12b are co-localized with each other in the same active zone indicating that they may form complexes regulating vesicle release in rostral IPN. \r\nOn electrophysiological studies of wild type (WT) mice, we found that paired-pulse ratio in rostral IPN of KCTD12b knock-out (KO) mice is lower than those of WT and KCTD8 KO mice. Consistent with this finding, in mean variance analysis, release probability in rostral IPN of KCTD12b KO mice is higher than that of WT and KCTD8 KO mice. Although paired-pulse ratios are not different between WT and KCTD8 KO mice, the mean variance analysis revealed significantly lower release probability in rostral IPN of KCTD8 KO than WT mice. These results demonstrate bidirectional regulation of Cav2.3-mediated release by KCTD8 and KCTD12b without GBR activation in rostral IPN. Finally, we examined the baclofen effects in rostral IPN of KCTD8 and KCTD12b KO mice, and found the facilitation of release remained in both KO mice, indicating that the peculiar effects of the GBR activation in this pathway do not depend on the selective expression of these KCTD subunits in ventral MHb. However, we found that presynaptic potentiation of evoked EPSC amplitude by baclofen falls to baseline after washout faster in KCTD12b KO mice than WT, KCTD8 KO and KCTD8/12b double KO mice. This result indicates that KCTD12b is involved in sustained potentiation of vesicle release by GBR activation, whereas KCTD8 is involved in its termination in the absence of KCTD12b. Consistent with these functional findings, replica labelling revealed an increase in density of KCTD8, but not Cav2.3 or GBR at active zone in rostral IPN of KCTD12b KO mice compared with that of WT mice, suggesting that increased association of KCTD8 with Cav2.3 facilitates the release probability and termination of the GBR effect in the absence of KCTD12b.\r\nIn summary, our study provided new insights into the physiological roles of presynaptic Cav2.3, GBRs and their auxiliary subunits KCTDs at an evolutionary conserved neuronal circuit. Future studies will be required to identify the exact molecular mechanism underlying the GBR-mediated presynaptic potentiation on ventral MHb terminals. It remains to be determined whether the prominent presence of presynaptic KCTDs at active zone could exert similar neuromodulatory functions in different pathways of the brain.\r\n" acknowledged_ssus: - _id: EM-Fac alternative_title: - ISTA Thesis article_processing_charge: No author: - first_name: Pradeep full_name: Bhandari, Pradeep id: 45EDD1BC-F248-11E8-B48F-1D18A9856A87 last_name: Bhandari orcid: 0000-0003-0863-4481 citation: ama: Bhandari P. Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway. 2020. doi:10.15479/AT:ISTA:7525 apa: Bhandari, P. (2020). Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:7525 chicago: Bhandari, Pradeep. “Localization and Functional Role of Cav2.3 in the Medial Habenula to Interpeduncular Nucleus Pathway.” Institute of Science and Technology Austria, 2020. https://doi.org/10.15479/AT:ISTA:7525. ieee: P. Bhandari, “Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway,” Institute of Science and Technology Austria, 2020. ista: Bhandari P. 2020. Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway. Institute of Science and Technology Austria. mla: Bhandari, Pradeep. Localization and Functional Role of Cav2.3 in the Medial Habenula to Interpeduncular Nucleus Pathway. Institute of Science and Technology Austria, 2020, doi:10.15479/AT:ISTA:7525. short: P. Bhandari, Localization and Functional Role of Cav2.3 in the Medial Habenula to Interpeduncular Nucleus Pathway, Institute of Science and Technology Austria, 2020. date_created: 2020-02-26T10:56:37Z date_published: 2020-02-28T00:00:00Z date_updated: 2023-09-07T13:20:03Z day: '28' ddc: - '570' degree_awarded: PhD department: - _id: RySh doi: 10.15479/AT:ISTA:7525 file: - access_level: open_access checksum: 4589234fdb12b4ad72273b311723a7b4 content_type: application/pdf creator: pbhandari date_created: 2020-02-28T08:37:53Z date_updated: 2021-03-01T23:30:04Z embargo: 2021-02-28 file_id: '7538' file_name: Pradeep Bhandari Thesis.pdf file_size: 9646346 relation: main_file title: Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway - access_level: closed checksum: aa79490553ca0a5c9b6fbcd152e93928 content_type: application/vnd.openxmlformats-officedocument.wordprocessingml.document creator: pbhandari date_created: 2020-02-28T08:47:14Z date_updated: 2021-03-01T23:30:04Z embargo_to: open_access file_id: '7539' file_name: Pradeep Bhandari Thesis.docx file_size: 35252164 relation: source_file title: Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway file_date_updated: 2021-03-01T23:30:04Z has_accepted_license: '1' keyword: - Cav2.3 - medial habenula (MHb) - interpeduncular nucleus (IPN) language: - iso: eng month: '02' oa: 1 oa_version: Published Version page: '79' publication_identifier: issn: - 2663-337X publication_status: published publisher: Institute of Science and Technology Austria 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: Localization and functional role of Cav2.3 in the medial habenula to interpeduncular nucleus pathway type: dissertation user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 year: '2020' ... --- _id: '7664' abstract: - lang: eng text: Metabotropic γ-aminobutyric acid (GABAB) receptors contribute to the control of network activity and information processing in hippocampal circuits by regulating neuronal excitability and synaptic transmission. The dysfunction in the dentate gyrus (DG) has been implicated in Alzheimer´s disease (AD). Given the involvement of GABAB receptors in AD, to determine their subcellular localisation and possible alteration in granule cells of the DG in a mouse model of AD at 12 months of age, we used high-resolution immunoelectron microscopic analysis. Immunohistochemistry at the light microscopic level showed that the regional and cellular expression pattern of GABAB1 was similar in an AD model mouse expressing mutated human amyloid precursor protein and presenilin1 (APP/PS1) and in age-matched wild type mice. High-resolution immunoelectron microscopy revealed a distance-dependent gradient of immunolabelling for GABAB receptors, increasing from proximal to distal dendrites in both wild type and APP/PS1 mice. However, the overall density of GABAB receptors at the neuronal surface of these postsynaptic compartments of granule cells was significantly reduced in APP/PS1 mice. Parallel to this reduction in surface receptors, we found a significant increase in GABAB1 at cytoplasmic sites. GABAB receptors were also detected at presynaptic sites in the molecular layer of the DG. We also found a decrease in plasma membrane GABAB receptors in axon terminals contacting dendritic spines of granule cells, which was more pronounced in the outer than in the inner molecular layer. Altogether, our data showing post- and presynaptic reduction in surface GABAB receptors in the DG suggest the alteration of the GABAB-mediated modulation of excitability and synaptic transmission in granule cells, which may contribute to the cognitive dysfunctions in the APP/PS1 model of AD article_number: '2459' article_processing_charge: No article_type: original author: - first_name: Alejandro full_name: Martín-Belmonte, Alejandro last_name: Martín-Belmonte - first_name: Carolina full_name: Aguado, Carolina last_name: Aguado - first_name: Rocío full_name: Alfaro-Ruíz, Rocío last_name: Alfaro-Ruíz - first_name: Ana Esther full_name: Moreno-Martínez, Ana Esther last_name: Moreno-Martínez - first_name: Luis full_name: De La Ossa, Luis last_name: De La Ossa - first_name: José full_name: Martínez-Hernández, José last_name: Martínez-Hernández - first_name: Alain full_name: Buisson, Alain last_name: Buisson - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Yugo full_name: Fukazawa, Yugo last_name: Fukazawa - first_name: Rafael full_name: Luján, Rafael last_name: Luján citation: ama: Martín-Belmonte A, Aguado C, Alfaro-Ruíz R, et al. Density of GABAB receptors is reduced in granule cells of the hippocampus in a mouse model of Alzheimer’s disease. International journal of molecular sciences. 2020;21(7). doi:10.3390/ijms21072459 apa: Martín-Belmonte, A., Aguado, C., Alfaro-Ruíz, R., Moreno-Martínez, A. E., De La Ossa, L., Martínez-Hernández, J., … Luján, R. (2020). Density of GABAB receptors is reduced in granule cells of the hippocampus in a mouse model of Alzheimer’s disease. International Journal of Molecular Sciences. MDPI. https://doi.org/10.3390/ijms21072459 chicago: Martín-Belmonte, Alejandro, Carolina Aguado, Rocío Alfaro-Ruíz, Ana Esther Moreno-Martínez, Luis De La Ossa, José Martínez-Hernández, Alain Buisson, Ryuichi Shigemoto, Yugo Fukazawa, and Rafael Luján. “Density of GABAB Receptors Is Reduced in Granule Cells of the Hippocampus in a Mouse Model of Alzheimer’s Disease.” International Journal of Molecular Sciences. MDPI, 2020. https://doi.org/10.3390/ijms21072459. ieee: A. Martín-Belmonte et al., “Density of GABAB receptors is reduced in granule cells of the hippocampus in a mouse model of Alzheimer’s disease,” International journal of molecular sciences, vol. 21, no. 7. MDPI, 2020. ista: Martín-Belmonte A, Aguado C, Alfaro-Ruíz R, Moreno-Martínez AE, De La Ossa L, Martínez-Hernández J, Buisson A, Shigemoto R, Fukazawa Y, Luján R. 2020. Density of GABAB receptors is reduced in granule cells of the hippocampus in a mouse model of Alzheimer’s disease. International journal of molecular sciences. 21(7), 2459. mla: Martín-Belmonte, Alejandro, et al. “Density of GABAB Receptors Is Reduced in Granule Cells of the Hippocampus in a Mouse Model of Alzheimer’s Disease.” International Journal of Molecular Sciences, vol. 21, no. 7, 2459, MDPI, 2020, doi:10.3390/ijms21072459. short: A. Martín-Belmonte, C. Aguado, R. Alfaro-Ruíz, A.E. Moreno-Martínez, L. De La Ossa, J. Martínez-Hernández, A. Buisson, R. Shigemoto, Y. Fukazawa, R. Luján, International Journal of Molecular Sciences 21 (2020). date_created: 2020-04-19T22:00:55Z date_published: 2020-04-02T00:00:00Z date_updated: 2023-08-21T06:13:19Z day: '02' ddc: - '570' department: - _id: RySh doi: 10.3390/ijms21072459 external_id: isi: - '000535574200201' pmid: - '32252271' file: - access_level: open_access checksum: b9d2f1657d8c4a74b01a62b474d009b0 content_type: application/pdf creator: dernst date_created: 2020-04-20T11:43:18Z date_updated: 2020-07-14T12:48:01Z file_id: '7669' file_name: 2020_JournMolecSciences_Martin_Belmonte.pdf file_size: 2941197 relation: main_file file_date_updated: 2020-07-14T12:48:01Z has_accepted_license: '1' intvolume: ' 21' isi: 1 issue: '7' language: - iso: eng month: '04' oa: 1 oa_version: Published Version pmid: 1 publication: International journal of molecular sciences publication_identifier: eissn: - '14220067' publication_status: published publisher: MDPI quality_controlled: '1' scopus_import: '1' status: public title: Density of GABAB receptors is reduced in granule cells of the hippocampus in a mouse model of Alzheimer's disease 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: '7665' abstract: - lang: eng text: Acute brain slice preparation is a powerful experimental model for investigating the characteristics of synaptic function in the brain. Although brain tissue is usually cut at ice-cold temperature (CT) to facilitate slicing and avoid neuronal damage, exposure to CT causes molecular and architectural changes of synapses. To address these issues, we investigated ultrastructural and electrophysiological features of synapses in mouse acute cerebellar slices prepared at ice-cold and physiological temperature (PT). In the slices prepared at CT, we found significant spine loss and reconstruction, synaptic vesicle rearrangement and decrease in synaptic proteins, all of which were not detected in slices prepared at PT. Consistent with these structural findings, slices prepared at PT showed higher release probability. Furthermore, preparation at PT allows electrophysiological recording immediately after slicing resulting in higher detectability of long-term depression (LTD) after motor learning compared with that at CT. These results indicate substantial advantages of the slice preparation at PT for investigating synaptic functions in different physiological conditions. article_number: '63' article_processing_charge: Yes (via OA deal) article_type: original author: - first_name: Kohgaku full_name: Eguchi, Kohgaku id: 2B7846DC-F248-11E8-B48F-1D18A9856A87 last_name: Eguchi orcid: 0000-0002-6170-2546 - first_name: Philipp full_name: Velicky, Philipp id: 39BDC62C-F248-11E8-B48F-1D18A9856A87 last_name: Velicky orcid: 0000-0002-2340-7431 - first_name: Elena full_name: Hollergschwandtner, Elena id: 3C054040-F248-11E8-B48F-1D18A9856A87 last_name: Hollergschwandtner - first_name: Makoto full_name: Itakura, Makoto last_name: Itakura - first_name: Yugo full_name: Fukazawa, Yugo last_name: Fukazawa - first_name: Johann G full_name: Danzl, Johann G id: 42EFD3B6-F248-11E8-B48F-1D18A9856A87 last_name: Danzl orcid: 0000-0001-8559-3973 - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 citation: ama: Eguchi K, Velicky P, Saeckl E, et al. Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions. Frontiers in Cellular Neuroscience. 2020;14. doi:10.3389/fncel.2020.00063 apa: Eguchi, K., Velicky, P., Saeckl, E., Itakura, M., Fukazawa, Y., Danzl, J. G., & Shigemoto, R. (2020). Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions. Frontiers in Cellular Neuroscience. Frontiers Media. https://doi.org/10.3389/fncel.2020.00063 chicago: Eguchi, Kohgaku, Philipp Velicky, Elena Saeckl, Makoto Itakura, Yugo Fukazawa, Johann G Danzl, and Ryuichi Shigemoto. “Advantages of Acute Brain Slices Prepared at Physiological Temperature in the Characterization of Synaptic Functions.” Frontiers in Cellular Neuroscience. Frontiers Media, 2020. https://doi.org/10.3389/fncel.2020.00063. ieee: K. Eguchi et al., “Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions,” Frontiers in Cellular Neuroscience, vol. 14. Frontiers Media, 2020. ista: Eguchi K, Velicky P, Saeckl E, Itakura M, Fukazawa Y, Danzl JG, Shigemoto R. 2020. Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions. Frontiers in Cellular Neuroscience. 14, 63. mla: Eguchi, Kohgaku, et al. “Advantages of Acute Brain Slices Prepared at Physiological Temperature in the Characterization of Synaptic Functions.” Frontiers in Cellular Neuroscience, vol. 14, 63, Frontiers Media, 2020, doi:10.3389/fncel.2020.00063. short: K. Eguchi, P. Velicky, E. Saeckl, M. Itakura, Y. Fukazawa, J.G. Danzl, R. Shigemoto, Frontiers in Cellular Neuroscience 14 (2020). date_created: 2020-04-19T22:00:55Z date_published: 2020-03-19T00:00:00Z date_updated: 2023-08-21T06:12:48Z day: '19' ddc: - '570' department: - _id: JoDa - _id: RySh doi: 10.3389/fncel.2020.00063 ec_funded: 1 external_id: isi: - '000525582200001' file: - access_level: open_access checksum: 1c145123c6f8dc3e2e4bd5a66a1ad60e content_type: application/pdf creator: dernst date_created: 2020-04-20T10:59:49Z date_updated: 2020-07-14T12:48:01Z file_id: '7668' file_name: 2020_FrontiersCellularNeurosc_Eguchi.pdf file_size: 9227283 relation: main_file file_date_updated: 2020-07-14T12:48:01Z has_accepted_license: '1' intvolume: ' 14' isi: 1 language: - iso: eng month: '03' oa: 1 oa_version: Published Version project: - _id: 2659CC84-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '793482' name: 'Ultrastructural analysis of phosphoinositides in nerve terminals: distribution, dynamics and physiological roles in synaptic transmission' - _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: 265CB4D0-B435-11E9-9278-68D0E5697425 call_identifier: FWF grant_number: I03600 name: Optical control of synaptic function via adhesion molecules - _id: B67AFEDC-15C9-11EA-A837-991A96BB2854 name: IST Austria Open Access Fund publication: Frontiers in Cellular Neuroscience publication_identifier: issn: - '16625102' publication_status: published publisher: Frontiers Media quality_controlled: '1' scopus_import: '1' status: public title: Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions 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: 14 year: '2020' ... --- _id: '6659' abstract: - lang: eng text: Chemical labeling of proteins with synthetic molecular probes offers the possibility to probe the functions of proteins of interest in living cells. However, the methods for covalently labeling targeted proteins using complementary peptide tag-probe pairs are still limited, irrespective of the versatility of such pairs in biological research. Herein, we report the new CysHis tag-Ni(II) probe pair for the specific covalent labeling of proteins. A broad-range evaluation of the reactivity profiles of the probe and the CysHis peptide tag afforded a tag-probe pair with an optimized and high labeling selectivity and reactivity. In particular, the labeling specificity of this pair was notably improved compared to the previously reported one. This pair was successfully utilized for the fluorescence imaging of membrane proteins on the surfaces of living cells, demonstrating its potential utility in biological research. acknowledgement: his work was supported by the Grant-in-Aid for Scientific Research B (JSPS KAKENHI grant no. JP17H03090 to A. O.); the Scientific Research on Innovative Areas “Chemistry for Multimolecular Crowding Biosystems” (JSPS KAKENHI grant no. JP17H06349 to A. O.); and the European Union (European Research Council Advanced grant no. 694539 and Human Brain Project Ref. 720270 to R. S.). A. O. acknowledges the financial support of the Takeda Science Foundation. article_processing_charge: No article_type: original author: - first_name: Naoki full_name: Zenmyo, Naoki last_name: Zenmyo - first_name: Hiroki full_name: Tokumaru, Hiroki last_name: Tokumaru - first_name: Shohei full_name: Uchinomiya, Shohei last_name: Uchinomiya - first_name: Hirokazu full_name: Fuchida, Hirokazu last_name: Fuchida - first_name: Shigekazu full_name: Tabata, Shigekazu id: 4427179E-F248-11E8-B48F-1D18A9856A87 last_name: Tabata - 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: Zenmyo N, Tokumaru H, Uchinomiya S, et al. Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins. Bulletin of the Chemical Society of Japan. 2019;92(5):995-1000. doi:10.1246/bcsj.20190034 apa: Zenmyo, N., Tokumaru, H., Uchinomiya, S., Fuchida, H., Tabata, S., Hamachi, I., … Ojida, A. (2019). Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins. Bulletin of the Chemical Society of Japan. Bulletin of the Chemical Society of Japan. https://doi.org/10.1246/bcsj.20190034 chicago: Zenmyo, Naoki, Hiroki Tokumaru, Shohei Uchinomiya, Hirokazu Fuchida, Shigekazu Tabata, Itaru Hamachi, Ryuichi Shigemoto, and Akio Ojida. “Optimized Reaction Pair of the CysHis Tag and Ni(II)-NTA Probe for Highly Selective Chemical Labeling of Membrane Proteins.” Bulletin of the Chemical Society of Japan. Bulletin of the Chemical Society of Japan, 2019. https://doi.org/10.1246/bcsj.20190034. ieee: N. Zenmyo et al., “Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins,” Bulletin of the Chemical Society of Japan, vol. 92, no. 5. Bulletin of the Chemical Society of Japan, pp. 995–1000, 2019. ista: Zenmyo N, Tokumaru H, Uchinomiya S, Fuchida H, Tabata S, Hamachi I, Shigemoto R, Ojida A. 2019. Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins. Bulletin of the Chemical Society of Japan. 92(5), 995–1000. mla: Zenmyo, Naoki, et al. “Optimized Reaction Pair of the CysHis Tag and Ni(II)-NTA Probe for Highly Selective Chemical Labeling of Membrane Proteins.” Bulletin of the Chemical Society of Japan, vol. 92, no. 5, Bulletin of the Chemical Society of Japan, 2019, pp. 995–1000, doi:10.1246/bcsj.20190034. short: N. Zenmyo, H. Tokumaru, S. Uchinomiya, H. Fuchida, S. Tabata, I. Hamachi, R. Shigemoto, A. Ojida, Bulletin of the Chemical Society of Japan 92 (2019) 995–1000. date_created: 2019-07-21T21:59:16Z date_published: 2019-05-15T00:00:00Z date_updated: 2021-01-12T08:08:26Z day: '15' ddc: - '570' department: - _id: RySh doi: 10.1246/bcsj.20190034 ec_funded: 1 file: - access_level: open_access checksum: 186de511d6e0ca93f5d981e2443eb8cd content_type: application/pdf creator: dernst date_created: 2020-10-02T08:49:58Z date_updated: 2020-10-02T08:49:58Z file_id: '8594' file_name: 2019_BCSJ_Zenmyo.pdf file_size: 2464903 relation: main_file success: 1 file_date_updated: 2020-10-02T08:49:58Z has_accepted_license: '1' intvolume: ' 92' issue: '5' language: - iso: eng month: '05' oa: 1 oa_version: Published Version page: 995-1000 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' publication: Bulletin of the Chemical Society of Japan publication_identifier: issn: - '00092673' publication_status: published publisher: Bulletin of the Chemical Society of Japan quality_controlled: '1' scopus_import: '1' status: public title: Optimized reaction pair of the CysHis tag and Ni(II)-NTA probe for highly selective chemical labeling of membrane proteins type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 92 year: '2019' ... --- _id: '6868' abstract: - lang: eng text: "Hyperpolarization-activated cyclic-nucleotide-gated (HCN) channels control electrical rhythmicity and excitability in the heart and brain, but the function of HCN channels at the subcellular level in axons remains poorly understood. Here, we show that the action potential conduction velocity in both myelinated and unmyelinated central axons can be bidirectionally modulated by a HCN channel blocker, cyclic adenosine monophosphate (cAMP), and neuromodulators. Recordings from mouse cerebellar mossy fiber boutons show that HCN channels ensure reliable high-frequency firing and are strongly modulated by cAMP (EC50 40 mM; estimated endogenous cAMP concentration 13 mM). In addition, immunogold-electron microscopy revealed HCN2 as the dominating subunit in cerebellar mossy fibers. Computational modeling indicated that HCN2 channels control conduction velocity primarily by altering the resting membrane potential\r\nand are associated with significant metabolic costs. These results suggest that the cAMP-HCN pathway provides neuromodulators with an opportunity to finely tune energy consumption and temporal delays across axons in the brain." article_number: e42766 article_processing_charge: No article_type: original author: - first_name: Niklas full_name: Byczkowicz, Niklas last_name: Byczkowicz - first_name: Abdelmoneim full_name: Eshra, Abdelmoneim last_name: Eshra - first_name: Jacqueline-Claire full_name: Montanaro-Punzengruber, Jacqueline-Claire id: 3786AB44-F248-11E8-B48F-1D18A9856A87 last_name: Montanaro-Punzengruber - first_name: Andrea full_name: Trevisiol, Andrea last_name: Trevisiol - first_name: Johannes full_name: Hirrlinger, Johannes last_name: Hirrlinger - first_name: Maarten Hp full_name: Kole, Maarten Hp last_name: Kole - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Stefan full_name: Hallermann, Stefan last_name: Hallermann citation: ama: Byczkowicz N, Eshra A, Montanaro-Punzengruber J-C, et al. HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons. eLife. 2019;8. doi:10.7554/eLife.42766 apa: Byczkowicz, N., Eshra, A., Montanaro-Punzengruber, J.-C., Trevisiol, A., Hirrlinger, J., Kole, M. H., … Hallermann, S. (2019). HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons. ELife. eLife Sciences Publications. https://doi.org/10.7554/eLife.42766 chicago: Byczkowicz, Niklas, Abdelmoneim Eshra, Jacqueline-Claire Montanaro-Punzengruber, Andrea Trevisiol, Johannes Hirrlinger, Maarten Hp Kole, Ryuichi Shigemoto, and Stefan Hallermann. “HCN Channel-Mediated Neuromodulation Can Control Action Potential Velocity and Fidelity in Central Axons.” ELife. eLife Sciences Publications, 2019. https://doi.org/10.7554/eLife.42766. ieee: N. Byczkowicz et al., “HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons,” eLife, vol. 8. eLife Sciences Publications, 2019. ista: Byczkowicz N, Eshra A, Montanaro-Punzengruber J-C, Trevisiol A, Hirrlinger J, Kole MH, Shigemoto R, Hallermann S. 2019. HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons. eLife. 8, e42766. mla: Byczkowicz, Niklas, et al. “HCN Channel-Mediated Neuromodulation Can Control Action Potential Velocity and Fidelity in Central Axons.” ELife, vol. 8, e42766, eLife Sciences Publications, 2019, doi:10.7554/eLife.42766. short: N. Byczkowicz, A. Eshra, J.-C. Montanaro-Punzengruber, A. Trevisiol, J. Hirrlinger, M.H. Kole, R. Shigemoto, S. Hallermann, ELife 8 (2019). date_created: 2019-09-15T22:00:43Z date_published: 2019-09-09T00:00:00Z date_updated: 2023-08-30T06:17:06Z day: '09' ddc: - '570' department: - _id: RySh doi: 10.7554/eLife.42766 external_id: isi: - '000485663900001' file: - access_level: open_access checksum: c350b7861ef0fb537cae8a3232aec016 content_type: application/pdf creator: dernst date_created: 2019-09-16T13:14:33Z date_updated: 2020-07-14T12:47:42Z file_id: '6880' file_name: 2019_eLife_Byczkowicz.pdf file_size: 4008137 relation: main_file file_date_updated: 2020-07-14T12:47:42Z has_accepted_license: '1' intvolume: ' 8' isi: 1 language: - iso: eng month: '09' oa: 1 oa_version: Published Version publication: eLife publication_identifier: eissn: - 2050084X publication_status: published publisher: eLife Sciences Publications quality_controlled: '1' scopus_import: '1' status: public title: HCN channel-mediated neuromodulation can control action potential velocity and fidelity in central axons 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: 8 year: '2019' ... --- _id: '7099' acknowledgement: "The authors thank Gabi Schmid for excellent technical support. We also thank\r\nDr. H. Harada, Dr. W. Kaufmann, and Dr. B. Kapelari for testing the specificity\r\nof some of the antibodies used in this study on replicas. Funding was provided\r\nby the Austrian Science Fund (Fonds zur Fo¨ rderung der Wissenschaftlichen\r\nForschung) Sonderforschungsbereich grants F44-17 (to F.jF.), F44-10 and\r\nP25375-B24 (to N.S.), and P26680 (to G.S.) and by the Novartis Research\r\nFoundation and the Swiss National Science Foundation (to A.L). We also thank\r\nProf. M. Capogna for reading a previous version of the manuscript." article_processing_charge: No article_type: original author: - first_name: Yu full_name: Kasugai, Yu last_name: Kasugai - first_name: Elisabeth full_name: Vogel, Elisabeth last_name: Vogel - first_name: Heide full_name: Hörtnagl, Heide last_name: Hörtnagl - first_name: Sabine full_name: Schönherr, Sabine last_name: Schönherr - first_name: Enrica full_name: Paradiso, Enrica last_name: Paradiso - first_name: Markus full_name: Hauschild, Markus last_name: Hauschild - first_name: Georg full_name: Göbel, Georg last_name: Göbel - first_name: Ivan full_name: Milenkovic, Ivan last_name: Milenkovic - first_name: Yvan full_name: Peterschmitt, Yvan last_name: Peterschmitt - first_name: Ramon full_name: Tasan, Ramon last_name: Tasan - first_name: Günther full_name: Sperk, Günther last_name: Sperk - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Werner full_name: Sieghart, Werner last_name: Sieghart - first_name: Nicolas full_name: Singewald, Nicolas last_name: Singewald - first_name: Andreas full_name: Lüthi, Andreas last_name: Lüthi - first_name: Francesco full_name: Ferraguti, Francesco last_name: Ferraguti citation: ama: Kasugai Y, Vogel E, Hörtnagl H, et al. Structural and functional remodeling of amygdala GABAergic synapses in associative fear learning. Neuron. 2019;104(4):781-794.e4. doi:10.1016/j.neuron.2019.08.013 apa: Kasugai, Y., Vogel, E., Hörtnagl, H., Schönherr, S., Paradiso, E., Hauschild, M., … Ferraguti, F. (2019). Structural and functional remodeling of amygdala GABAergic synapses in associative fear learning. Neuron. Elsevier. https://doi.org/10.1016/j.neuron.2019.08.013 chicago: Kasugai, Yu, Elisabeth Vogel, Heide Hörtnagl, Sabine Schönherr, Enrica Paradiso, Markus Hauschild, Georg Göbel, et al. “Structural and Functional Remodeling of Amygdala GABAergic Synapses in Associative Fear Learning.” Neuron. Elsevier, 2019. https://doi.org/10.1016/j.neuron.2019.08.013. ieee: Y. Kasugai et al., “Structural and functional remodeling of amygdala GABAergic synapses in associative fear learning,” Neuron, vol. 104, no. 4. Elsevier, p. 781–794.e4, 2019. ista: Kasugai Y, Vogel E, Hörtnagl H, Schönherr S, Paradiso E, Hauschild M, Göbel G, Milenkovic I, Peterschmitt Y, Tasan R, Sperk G, Shigemoto R, Sieghart W, Singewald N, Lüthi A, Ferraguti F. 2019. Structural and functional remodeling of amygdala GABAergic synapses in associative fear learning. Neuron. 104(4), 781–794.e4. mla: Kasugai, Yu, et al. “Structural and Functional Remodeling of Amygdala GABAergic Synapses in Associative Fear Learning.” Neuron, vol. 104, no. 4, Elsevier, 2019, p. 781–794.e4, doi:10.1016/j.neuron.2019.08.013. short: Y. Kasugai, E. Vogel, H. Hörtnagl, S. Schönherr, E. Paradiso, M. Hauschild, G. Göbel, I. Milenkovic, Y. Peterschmitt, R. Tasan, G. Sperk, R. Shigemoto, W. Sieghart, N. Singewald, A. Lüthi, F. Ferraguti, Neuron 104 (2019) 781–794.e4. date_created: 2019-11-25T08:02:39Z date_published: 2019-11-20T00:00:00Z date_updated: 2023-08-30T07:28:22Z day: '20' ddc: - '571' - '599' department: - _id: RySh doi: 10.1016/j.neuron.2019.08.013 external_id: isi: - '000497963500017' pmid: - '31543297' has_accepted_license: '1' intvolume: ' 104' isi: 1 issue: '4' language: - iso: eng main_file_link: - open_access: '1' url: https://doi.org/10.1016/j.neuron.2019.08.013 month: '11' oa: 1 oa_version: Published Version page: 781-794.e4 pmid: 1 publication: Neuron publication_identifier: issn: - 0896-6273 publication_status: published publisher: Elsevier quality_controlled: '1' scopus_import: '1' status: public title: Structural and functional remodeling of amygdala GABAergic synapses in associative fear learning type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 104 year: '2019' ... --- _id: '7179' abstract: - lang: eng text: Glutamate is the major excitatory neurotransmitter in the CNS binding to a variety of glutamate receptors. Metabotropic glutamate receptors (mGluR1 to mGluR8) can act excitatory or inhibitory, depending on associated signal cascades. Expression and localization of inhibitory acting mGluRs at inner hair cells (IHCs) in the cochlea are largely unknown. Here, we analyzed expression of mGluR2, mGluR3, mGluR4, mGluR6, mGluR7, and mGluR8 and investigated their localization with respect to the presynaptic ribbon of IHC synapses. We detected transcripts for mGluR2, mGluR3, and mGluR4 as well as for mGluR7a, mGluR7b, mGluR8a, and mGluR8b splice variants. Using receptor-specific antibodies in cochlear wholemounts, we found expression of mGluR2, mGluR4, and mGluR8b close to presynaptic ribbons. Super resolution and confocal microscopy in combination with 3-dimensional reconstructions indicated a postsynaptic localization of mGluR2 that overlaps with postsynaptic density protein 95 on dendrites of afferent type I spiral ganglion neurons. In contrast, mGluR4 and mGluR8b were expressed at the presynapse close to IHC ribbons. In summary, we localized in detail 3 mGluR types at IHC ribbon synapses, providing a fundament for new therapeutical strategies that could protect the cochlea against noxious stimuli and excitotoxicity. article_processing_charge: No article_type: original author: - first_name: Lisa full_name: Klotz, Lisa last_name: Klotz - first_name: Olaf full_name: Wendler, Olaf last_name: Wendler - first_name: Renato full_name: Frischknecht, Renato last_name: Frischknecht - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Holger full_name: Schulze, Holger last_name: Schulze - first_name: Ralf full_name: Enz, Ralf last_name: Enz citation: ama: Klotz L, Wendler O, Frischknecht R, Shigemoto R, Schulze H, Enz R. Localization of group II and III metabotropic glutamate receptors at pre- and postsynaptic sites of inner hair cell ribbon synapses. FASEB Journal. 2019;33(12):13734-13746. doi:10.1096/fj.201901543R apa: Klotz, L., Wendler, O., Frischknecht, R., Shigemoto, R., Schulze, H., & Enz, R. (2019). Localization of group II and III metabotropic glutamate receptors at pre- and postsynaptic sites of inner hair cell ribbon synapses. FASEB Journal. FASEB. https://doi.org/10.1096/fj.201901543R chicago: Klotz, Lisa, Olaf Wendler, Renato Frischknecht, Ryuichi Shigemoto, Holger Schulze, and Ralf Enz. “Localization of Group II and III Metabotropic Glutamate Receptors at Pre- and Postsynaptic Sites of Inner Hair Cell Ribbon Synapses.” FASEB Journal. FASEB, 2019. https://doi.org/10.1096/fj.201901543R. ieee: L. Klotz, O. Wendler, R. Frischknecht, R. Shigemoto, H. Schulze, and R. Enz, “Localization of group II and III metabotropic glutamate receptors at pre- and postsynaptic sites of inner hair cell ribbon synapses,” FASEB Journal, vol. 33, no. 12. FASEB, pp. 13734–13746, 2019. ista: Klotz L, Wendler O, Frischknecht R, Shigemoto R, Schulze H, Enz R. 2019. Localization of group II and III metabotropic glutamate receptors at pre- and postsynaptic sites of inner hair cell ribbon synapses. FASEB Journal. 33(12), 13734–13746. mla: Klotz, Lisa, et al. “Localization of Group II and III Metabotropic Glutamate Receptors at Pre- and Postsynaptic Sites of Inner Hair Cell Ribbon Synapses.” FASEB Journal, vol. 33, no. 12, FASEB, 2019, pp. 13734–46, doi:10.1096/fj.201901543R. short: L. Klotz, O. Wendler, R. Frischknecht, R. Shigemoto, H. Schulze, R. Enz, FASEB Journal 33 (2019) 13734–13746. date_created: 2019-12-15T23:00:42Z date_published: 2019-12-01T00:00:00Z date_updated: 2023-09-06T14:34:36Z day: '01' ddc: - '571' - '599' department: - _id: RySh doi: 10.1096/fj.201901543R external_id: isi: - '000507466100054' pmid: - '31585509' file: - access_level: open_access checksum: 79e3b72481dc32489911121cf3b7d8d0 content_type: application/pdf creator: shigemot date_created: 2020-12-06T17:30:09Z date_updated: 2020-12-06T17:30:09Z file_id: '8922' file_name: Klotz et al 2019 EMBO Reports.pdf file_size: 4766789 relation: main_file success: 1 file_date_updated: 2020-12-06T17:30:09Z has_accepted_license: '1' intvolume: ' 33' isi: 1 issue: '12' language: - iso: eng month: '12' oa: 1 oa_version: Submitted Version page: 13734-13746 pmid: 1 publication: FASEB Journal publication_identifier: eissn: - '15306860' publication_status: published publisher: FASEB quality_controlled: '1' scopus_import: '1' status: public title: Localization of group II and III metabotropic glutamate receptors at pre- and postsynaptic sites of inner hair cell ribbon synapses type: journal_article user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 volume: 33 year: '2019' ... --- _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: 2024-03-25T23:30:07Z 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 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' ... --- _id: '7398' abstract: - lang: eng text: 'Transporters of the solute carrier 6 (SLC6) family translocate their cognate substrate together with Na+ and Cl−. Detailed kinetic models exist for the transporters of GABA (GAT1/SLC6A1) and the monoamines dopamine (DAT/SLC6A3) and serotonin (SERT/SLC6A4). Here, we posited that the transport cycle of individual SLC6 transporters reflects the physiological requirements they operate under. We tested this hypothesis by analyzing the transport cycle of glycine transporter 1 (GlyT1/SLC6A9) and glycine transporter 2 (GlyT2/SLC6A5). GlyT2 is the only SLC6 family member known to translocate glycine, Na+, and Cl− in a 1:3:1 stoichiometry. We analyzed partial reactions in real time by electrophysiological recordings. Contrary to monoamine transporters, both GlyTs were found to have a high transport capacity driven by rapid return of the empty transporter after release of Cl− on the intracellular side. Rapid cycling of both GlyTs was further supported by highly cooperative binding of cosubstrate ions and substrate such that their forward transport mode was maintained even under conditions of elevated intracellular Na+ or Cl−. The most important differences in the transport cycle of GlyT1 and GlyT2 arose from the kinetics of charge movement and the resulting voltage-dependent rate-limiting reactions: the kinetics of GlyT1 were governed by transition of the substrate-bound transporter from outward- to inward-facing conformations, whereas the kinetics of GlyT2 were governed by Na+ binding (or a related conformational change). Kinetic modeling showed that the kinetics of GlyT1 are ideally suited for supplying the extracellular glycine levels required for NMDA receptor activation.' article_processing_charge: No article_type: original author: - first_name: Fatma Asli full_name: Erdem, Fatma Asli last_name: Erdem - first_name: Marija full_name: Ilic, Marija last_name: Ilic - first_name: Peter full_name: Koppensteiner, Peter id: 3B8B25A8-F248-11E8-B48F-1D18A9856A87 last_name: Koppensteiner orcid: 0000-0002-3509-1948 - first_name: Jakub full_name: Gołacki, Jakub last_name: Gołacki - first_name: Gert full_name: Lubec, Gert last_name: Lubec - first_name: Michael full_name: Freissmuth, Michael last_name: Freissmuth - first_name: Walter full_name: Sandtner, Walter last_name: Sandtner citation: ama: Erdem FA, Ilic M, Koppensteiner P, et al. A comparison of the transport kinetics of glycine transporter 1 and glycine transporter 2. The Journal of General Physiology. 2019;151(8):1035-1050. doi:10.1085/jgp.201912318 apa: Erdem, F. A., Ilic, M., Koppensteiner, P., Gołacki, J., Lubec, G., Freissmuth, M., & Sandtner, W. (2019). A comparison of the transport kinetics of glycine transporter 1 and glycine transporter 2. The Journal of General Physiology. Rockefeller University Press. https://doi.org/10.1085/jgp.201912318 chicago: Erdem, Fatma Asli, Marija Ilic, Peter Koppensteiner, Jakub Gołacki, Gert Lubec, Michael Freissmuth, and Walter Sandtner. “A Comparison of the Transport Kinetics of Glycine Transporter 1 and Glycine Transporter 2.” The Journal of General Physiology. Rockefeller University Press, 2019. https://doi.org/10.1085/jgp.201912318. ieee: F. A. Erdem et al., “A comparison of the transport kinetics of glycine transporter 1 and glycine transporter 2,” The Journal of General Physiology, vol. 151, no. 8. Rockefeller University Press, pp. 1035–1050, 2019. ista: Erdem FA, Ilic M, Koppensteiner P, Gołacki J, Lubec G, Freissmuth M, Sandtner W. 2019. A comparison of the transport kinetics of glycine transporter 1 and glycine transporter 2. The Journal of General Physiology. 151(8), 1035–1050. mla: Erdem, Fatma Asli, et al. “A Comparison of the Transport Kinetics of Glycine Transporter 1 and Glycine Transporter 2.” The Journal of General Physiology, vol. 151, no. 8, Rockefeller University Press, 2019, pp. 1035–50, doi:10.1085/jgp.201912318. short: F.A. Erdem, M. Ilic, P. Koppensteiner, J. Gołacki, G. Lubec, M. Freissmuth, W. Sandtner, The Journal of General Physiology 151 (2019) 1035–1050. date_created: 2020-01-29T16:06:29Z date_published: 2019-07-03T00:00:00Z date_updated: 2023-09-07T14:52:23Z day: '03' ddc: - '570' department: - _id: RySh doi: 10.1085/jgp.201912318 external_id: isi: - '000478792500008' pmid: - '31270129' file: - access_level: open_access checksum: 5706b4ccd74ee3e50bf7ecb2a203df71 content_type: application/pdf creator: dernst date_created: 2020-02-05T07:20:32Z date_updated: 2020-07-14T12:47:57Z file_id: '7450' file_name: 2019_JGP_Erdem.pdf file_size: 2641297 relation: main_file file_date_updated: 2020-07-14T12:47:57Z has_accepted_license: '1' intvolume: ' 151' isi: 1 issue: '8' language: - iso: eng license: https://creativecommons.org/licenses/by-nc-sa/4.0/ month: '07' oa: 1 oa_version: Published Version page: 1035-1050 pmid: 1 publication: The Journal of General Physiology publication_identifier: eissn: - 1540-7748 issn: - 0022-1295 publication_status: published publisher: Rockefeller University Press quality_controlled: '1' scopus_import: '1' status: public title: A comparison of the transport kinetics of glycine transporter 1 and glycine transporter 2 tmp: image: /images/cc_by_nc_sa.png legal_code_url: https://creativecommons.org/licenses/by-nc-sa/4.0/legalcode name: Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0) short: CC BY-NC-SA (4.0) type: journal_article user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 volume: 151 year: '2019' ... --- _id: '326' abstract: - lang: eng text: Three-dimensional (3D) super-resolution microscopy technique structured illumination microscopy (SIM) imaging of dendritic spines along the dendrite has not been previously performed in fixed tissues, mainly due to deterioration of the stripe pattern of the excitation laser induced by light scattering and optical aberrations. To address this issue and solve these optical problems, we applied a novel clearing reagent, LUCID, to fixed brains. In SIM imaging, the penetration depth and the spatial resolution were improved in LUCID-treated slices, and 160-nm spatial resolution was obtained in a large portion of the imaging volume on a single apical dendrite. Furthermore, in a morphological analysis of spine heads of layer V pyramidal neurons (L5PNs) in the medial prefrontal cortex (mPFC) of chronic dexamethasone (Dex)-treated mice, SIM imaging revealed an altered distribution of spine forms that could not be detected by high-NA confocal imaging. Thus, super-resolution SIM imaging represents a promising high-throughput method for revealing spine morphologies in single dendrites. acknowledged_ssus: - _id: EM-Fac article_processing_charge: No author: - first_name: Kazuaki full_name: Sawada, Kazuaki last_name: Sawada - first_name: Ryosuke full_name: Kawakami, Ryosuke last_name: Kawakami - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Tomomi full_name: Nemoto, Tomomi last_name: Nemoto citation: ama: Sawada K, Kawakami R, Shigemoto R, Nemoto T. Super resolution structural analysis of dendritic spines using three-dimensional structured illumination microscopy in cleared mouse brain slices. European Journal of Neuroscience. 2018;47(9):1033-1042. doi:10.1111/ejn.13901 apa: Sawada, K., Kawakami, R., Shigemoto, R., & Nemoto, T. (2018). Super resolution structural analysis of dendritic spines using three-dimensional structured illumination microscopy in cleared mouse brain slices. European Journal of Neuroscience. Wiley. https://doi.org/10.1111/ejn.13901 chicago: Sawada, Kazuaki, Ryosuke Kawakami, Ryuichi Shigemoto, and Tomomi Nemoto. “Super Resolution Structural Analysis of Dendritic Spines Using Three-Dimensional Structured Illumination Microscopy in Cleared Mouse Brain Slices.” European Journal of Neuroscience. Wiley, 2018. https://doi.org/10.1111/ejn.13901. ieee: K. Sawada, R. Kawakami, R. Shigemoto, and T. Nemoto, “Super resolution structural analysis of dendritic spines using three-dimensional structured illumination microscopy in cleared mouse brain slices,” European Journal of Neuroscience, vol. 47, no. 9. Wiley, pp. 1033–1042, 2018. ista: Sawada K, Kawakami R, Shigemoto R, Nemoto T. 2018. Super resolution structural analysis of dendritic spines using three-dimensional structured illumination microscopy in cleared mouse brain slices. European Journal of Neuroscience. 47(9), 1033–1042. mla: Sawada, Kazuaki, et al. “Super Resolution Structural Analysis of Dendritic Spines Using Three-Dimensional Structured Illumination Microscopy in Cleared Mouse Brain Slices.” European Journal of Neuroscience, vol. 47, no. 9, Wiley, 2018, pp. 1033–42, doi:10.1111/ejn.13901. short: K. Sawada, R. Kawakami, R. Shigemoto, T. Nemoto, European Journal of Neuroscience 47 (2018) 1033–1042. date_created: 2018-12-11T11:45:50Z date_published: 2018-03-07T00:00:00Z date_updated: 2023-09-19T09:58:40Z day: '07' ddc: - '570' department: - _id: RySh doi: 10.1111/ejn.13901 external_id: isi: - '000431496400001' file: - access_level: open_access checksum: 98e901d8229e44aa8f3b51d248dedd09 content_type: application/pdf creator: dernst date_created: 2018-12-17T16:16:50Z date_updated: 2020-07-14T12:46:06Z file_id: '5721' file_name: 2018_EJN_Sawada.pdf file_size: 4850261 relation: main_file file_date_updated: 2020-07-14T12:46:06Z has_accepted_license: '1' intvolume: ' 47' isi: 1 issue: '9' language: - iso: eng license: https://creativecommons.org/licenses/by-nc/4.0/ month: '03' oa: 1 oa_version: Published Version page: 1033 - 1042 publication: European Journal of Neuroscience publication_status: published publisher: Wiley publist_id: '7539' quality_controlled: '1' scopus_import: '1' status: public title: Super resolution structural analysis of dendritic spines using three-dimensional structured illumination microscopy in cleared mouse brain slices tmp: image: /images/cc_by_nc.png legal_code_url: https://creativecommons.org/licenses/by-nc/4.0/legalcode name: Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) short: CC BY-NC (4.0) type: journal_article user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 volume: 47 year: '2018' ... --- _id: '163' abstract: - lang: eng text: For ultrafast fixation of biological samples to avoid artifacts, high-pressure freezing (HPF) followed by freeze substitution (FS) is preferred over chemical fixation at room temperature. After HPF, samples are maintained at low temperature during dehydration and fixation, while avoiding damaging recrystallization. This is a notoriously slow process. McDonald and Webb demonstrated, in 2011, that sample agitation during FS dramatically reduces the necessary time. Then, in 2015, we (H.G. and S.R.) introduced an agitation module into the cryochamber of an automated FS unit and demonstrated that the preparation of algae could be shortened from days to a couple of hours. We argued that variability in the processing, reproducibility, and safety issues are better addressed using automated FS units. For dissemination, we started low-cost manufacturing of agitation modules for two of the most widely used FS units, the Automatic Freeze Substitution Systems, AFS(1) and AFS2, from Leica Microsystems, using three dimensional (3D)-printing of the major components. To test them, several labs independently used the modules on a wide variety of specimens that had previously been processed by manual agitation, or without agitation. We demonstrate that automated processing with sample agitation saves time, increases flexibility with respect to sample requirements and protocols, and produces data of at least as good quality as other approaches. article_processing_charge: No article_type: original author: - first_name: Siegfried full_name: Reipert, Siegfried last_name: Reipert - first_name: Helmuth full_name: Goldammer, Helmuth last_name: Goldammer - first_name: Christine full_name: Richardson, Christine last_name: Richardson - first_name: Martin full_name: Goldberg, Martin last_name: Goldberg - first_name: Timothy full_name: Hawkins, Timothy last_name: Hawkins - first_name: Elena full_name: Hollergschwandtner, Elena id: 3C054040-F248-11E8-B48F-1D18A9856A87 last_name: Hollergschwandtner - first_name: Walter full_name: Kaufmann, Walter id: 3F99E422-F248-11E8-B48F-1D18A9856A87 last_name: Kaufmann orcid: 0000-0001-9735-5315 - first_name: Sebastian full_name: Antreich, Sebastian last_name: Antreich - first_name: York full_name: Stierhof, York last_name: Stierhof citation: ama: 'Reipert S, Goldammer H, Richardson C, et al. Agitation modules: Flexible means to accelerate automated freeze substitution. Journal of Histochemistry and Cytochemistry. 2018;66(12):903-921. doi:10.1369/0022155418786698' apa: 'Reipert, S., Goldammer, H., Richardson, C., Goldberg, M., Hawkins, T., Saeckl, E., … Stierhof, Y. (2018). Agitation modules: Flexible means to accelerate automated freeze substitution. Journal of Histochemistry and Cytochemistry. SAGE Publications. https://doi.org/10.1369/0022155418786698' chicago: 'Reipert, Siegfried, Helmuth Goldammer, Christine Richardson, Martin Goldberg, Timothy Hawkins, Elena Saeckl, Walter Kaufmann, Sebastian Antreich, and York Stierhof. “Agitation Modules: Flexible Means to Accelerate Automated Freeze Substitution.” Journal of Histochemistry and Cytochemistry. SAGE Publications, 2018. https://doi.org/10.1369/0022155418786698.' ieee: 'S. Reipert et al., “Agitation modules: Flexible means to accelerate automated freeze substitution,” Journal of Histochemistry and Cytochemistry, vol. 66, no. 12. SAGE Publications, pp. 903–921, 2018.' ista: 'Reipert S, Goldammer H, Richardson C, Goldberg M, Hawkins T, Saeckl E, Kaufmann W, Antreich S, Stierhof Y. 2018. Agitation modules: Flexible means to accelerate automated freeze substitution. Journal of Histochemistry and Cytochemistry. 66(12), 903–921.' mla: 'Reipert, Siegfried, et al. “Agitation Modules: Flexible Means to Accelerate Automated Freeze Substitution.” Journal of Histochemistry and Cytochemistry, vol. 66, no. 12, SAGE Publications, 2018, pp. 903–21, doi:10.1369/0022155418786698.' short: S. Reipert, H. Goldammer, C. Richardson, M. Goldberg, T. Hawkins, E. Saeckl, W. Kaufmann, S. Antreich, Y. Stierhof, Journal of Histochemistry and Cytochemistry 66 (2018) 903–921. date_created: 2018-12-11T11:44:57Z date_published: 2018-12-01T00:00:00Z date_updated: 2023-10-17T08:42:24Z day: '01' department: - _id: RySh - _id: EM-Fac doi: 10.1369/0022155418786698 external_id: isi: - '000452277700005' pmid: - '29969056' intvolume: ' 66' isi: 1 issue: '12' language: - iso: eng main_file_link: - open_access: '1' url: https://doi.org/10.1369/0022155418786698 month: '12' oa: 1 oa_version: Published Version page: 903-921 pmid: 1 publication: Journal of Histochemistry and Cytochemistry publication_identifier: issn: - 0022-1554 publication_status: published publisher: SAGE Publications quality_controlled: '1' scopus_import: '1' status: public title: 'Agitation modules: Flexible means to accelerate automated freeze substitution' type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 66 year: '2018' ... --- _id: '705' abstract: - lang: eng text: Although dopamine receptors D1 and D2 play key roles in hippocampal function, their synaptic localization within the hippocampus has not been fully elucidated. In order to understand precise functions of pre- or postsynaptic dopamine receptors (DRs), the development of protocols to differentiate pre- and postsynaptic DRs is essential. So far, most studies on determination and quantification of DRs did not discriminate between subsynaptic localization. Therefore, the aim of the study was to generate a robust workflow for the localization of DRs. This work provides the basis for future work on hippocampal DRs, in light that DRs may have different functions at pre- or postsynaptic sites. Synaptosomes from rat hippocampi isolated by a sucrose gradient protocol were prepared for super-resolution direct stochastic optical reconstruction microscopy (dSTORM) using Bassoon as a presynaptic zone and Homer1 as postsynaptic density marker. Direct labeling of primary validated antibodies against dopamine receptors D1 (D1R) and D2 (D2R) with Alexa Fluor 594 enabled unequivocal assignment of D1R and D2R to both, pre- and postsynaptic sites. D1R immunoreactivity clusters were observed within the presynaptic active zone as well as at perisynaptic sites at the edge of the presynaptic active zone. The results may be useful for the interpretation of previous studies and the design of future work on DRs in the hippocampus. Moreover, the reduction of the complexity of brain tissue by the use of synaptosomal preparations and dSTORM technology may represent a useful tool for synaptic localization of brain proteins. article_processing_charge: No author: - first_name: Andras full_name: Miklosi, Andras last_name: Miklosi - first_name: Giorgia full_name: Del Favero, Giorgia last_name: Del Favero - first_name: Tanja full_name: Bulat, Tanja last_name: Bulat - first_name: Harald full_name: Höger, Harald last_name: Höger - first_name: Ryuichi full_name: Shigemoto, Ryuichi id: 499F3ABC-F248-11E8-B48F-1D18A9856A87 last_name: Shigemoto orcid: 0000-0001-8761-9444 - first_name: Doris full_name: Marko, Doris last_name: Marko - first_name: Gert full_name: Lubec, Gert last_name: Lubec citation: ama: Miklosi A, Del Favero G, Bulat T, et al. Super resolution microscopical localization of dopamine receptors 1 and 2 in rat hippocampal synaptosomes. Molecular Neurobiology. 2018;55(6):4857 – 4869. doi:10.1007/s12035-017-0688-y apa: Miklosi, A., Del Favero, G., Bulat, T., Höger, H., Shigemoto, R., Marko, D., & Lubec, G. (2018). Super resolution microscopical localization of dopamine receptors 1 and 2 in rat hippocampal synaptosomes. Molecular Neurobiology. Springer. https://doi.org/10.1007/s12035-017-0688-y chicago: Miklosi, Andras, Giorgia Del Favero, Tanja Bulat, Harald Höger, Ryuichi Shigemoto, Doris Marko, and Gert Lubec. “Super Resolution Microscopical Localization of Dopamine Receptors 1 and 2 in Rat Hippocampal Synaptosomes.” Molecular Neurobiology. Springer, 2018. https://doi.org/10.1007/s12035-017-0688-y. ieee: A. Miklosi et al., “Super resolution microscopical localization of dopamine receptors 1 and 2 in rat hippocampal synaptosomes,” Molecular Neurobiology, vol. 55, no. 6. Springer, pp. 4857 – 4869, 2018. ista: Miklosi A, Del Favero G, Bulat T, Höger H, Shigemoto R, Marko D, Lubec G. 2018. Super resolution microscopical localization of dopamine receptors 1 and 2 in rat hippocampal synaptosomes. Molecular Neurobiology. 55(6), 4857 – 4869. mla: Miklosi, Andras, et al. “Super Resolution Microscopical Localization of Dopamine Receptors 1 and 2 in Rat Hippocampal Synaptosomes.” Molecular Neurobiology, vol. 55, no. 6, Springer, 2018, pp. 4857 – 4869, doi:10.1007/s12035-017-0688-y. short: A. Miklosi, G. Del Favero, T. Bulat, H. Höger, R. Shigemoto, D. Marko, G. Lubec, Molecular Neurobiology 55 (2018) 4857 – 4869. date_created: 2018-12-11T11:48:02Z date_published: 2018-06-01T00:00:00Z date_updated: 2023-09-19T09:58:11Z day: '01' department: - _id: RySh doi: 10.1007/s12035-017-0688-y external_id: isi: - '000431991500025' intvolume: ' 55' isi: 1 issue: '6' language: - iso: eng month: '06' oa_version: None page: 4857 – 4869 publication: Molecular Neurobiology publication_status: published publisher: Springer publist_id: '6991' quality_controlled: '1' scopus_import: '1' status: public title: Super resolution microscopical localization of dopamine receptors 1 and 2 in rat hippocampal synaptosomes type: journal_article user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 volume: 55 year: '2018' ...