[{"month":"08","department":[{"_id":"PeJo"},{"_id":"GaNo"},{"_id":"BeBi"},{"_id":"JoDa"},{"_id":"Bio"}],"language":[{"iso":"eng"}],"oa":1,"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","citation":{"apa":"Velicky, P., Miguel Villalba, E., Michalska, J. M., Lyudchik, J., Wei, D., Lin, Z., … Danzl, J. G. (2023). Dense 4D nanoscale reconstruction of living brain tissue. <i>Nature Methods</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41592-023-01936-6\">https://doi.org/10.1038/s41592-023-01936-6</a>","mla":"Velicky, Philipp, et al. “Dense 4D Nanoscale Reconstruction of Living Brain Tissue.” <i>Nature Methods</i>, vol. 20, Springer Nature, 2023, pp. 1256–65, doi:<a href=\"https://doi.org/10.1038/s41592-023-01936-6\">10.1038/s41592-023-01936-6</a>.","chicago":"Velicky, Philipp, Eder Miguel Villalba, Julia M Michalska, Julia Lyudchik, Donglai Wei, Zudi Lin, Jake Watson, et al. “Dense 4D Nanoscale Reconstruction of Living Brain Tissue.” <i>Nature Methods</i>. Springer Nature, 2023. <a href=\"https://doi.org/10.1038/s41592-023-01936-6\">https://doi.org/10.1038/s41592-023-01936-6</a>.","ista":"Velicky P, Miguel Villalba E, Michalska JM, Lyudchik J, Wei D, Lin Z, Watson J, Troidl J, Beyer J, Ben Simon Y, Sommer CM, Jahr W, Cenameri A, Broichhagen J, Grant SGN, Jonas PM, Novarino G, Pfister H, Bickel B, Danzl JG. 2023. Dense 4D nanoscale reconstruction of living brain tissue. Nature Methods. 20, 1256–1265.","ieee":"P. Velicky <i>et al.</i>, “Dense 4D nanoscale reconstruction of living brain tissue,” <i>Nature Methods</i>, vol. 20. Springer Nature, pp. 1256–1265, 2023.","short":"P. Velicky, E. Miguel Villalba, J.M. Michalska, J. Lyudchik, D. Wei, Z. Lin, J. Watson, J. Troidl, J. Beyer, Y. Ben Simon, C.M. Sommer, W. Jahr, A. Cenameri, J. Broichhagen, S.G.N. Grant, P.M. Jonas, G. Novarino, H. Pfister, B. Bickel, J.G. Danzl, Nature Methods 20 (2023) 1256–1265.","ama":"Velicky P, Miguel Villalba E, Michalska JM, et al. Dense 4D nanoscale reconstruction of living brain tissue. <i>Nature Methods</i>. 2023;20:1256-1265. doi:<a href=\"https://doi.org/10.1038/s41592-023-01936-6\">10.1038/s41592-023-01936-6</a>"},"title":"Dense 4D nanoscale reconstruction of living brain tissue","oa_version":"Published Version","author":[{"last_name":"Velicky","full_name":"Velicky, Philipp","id":"39BDC62C-F248-11E8-B48F-1D18A9856A87","first_name":"Philipp","orcid":"0000-0002-2340-7431"},{"first_name":"Eder","orcid":"0000-0001-5665-0430","last_name":"Miguel Villalba","full_name":"Miguel Villalba, Eder","id":"3FB91342-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Julia M","orcid":"0000-0003-3862-1235","last_name":"Michalska","full_name":"Michalska, Julia M","id":"443DB6DE-F248-11E8-B48F-1D18A9856A87"},{"id":"46E28B80-F248-11E8-B48F-1D18A9856A87","full_name":"Lyudchik, Julia","last_name":"Lyudchik","first_name":"Julia"},{"full_name":"Wei, Donglai","last_name":"Wei","first_name":"Donglai"},{"first_name":"Zudi","last_name":"Lin","full_name":"Lin, Zudi"},{"last_name":"Watson","id":"63836096-4690-11EA-BD4E-32803DDC885E","full_name":"Watson, Jake","orcid":"0000-0002-8698-3823","first_name":"Jake"},{"first_name":"Jakob","last_name":"Troidl","full_name":"Troidl, Jakob"},{"first_name":"Johanna","last_name":"Beyer","full_name":"Beyer, Johanna"},{"last_name":"Ben Simon","full_name":"Ben Simon, Yoav","id":"43DF3136-F248-11E8-B48F-1D18A9856A87","first_name":"Yoav"},{"last_name":"Sommer","full_name":"Sommer, Christoph M","id":"4DF26D8C-F248-11E8-B48F-1D18A9856A87","first_name":"Christoph M","orcid":"0000-0003-1216-9105"},{"last_name":"Jahr","full_name":"Jahr, Wiebke","id":"425C1CE8-F248-11E8-B48F-1D18A9856A87","first_name":"Wiebke"},{"last_name":"Cenameri","full_name":"Cenameri, Alban","id":"9ac8f577-2357-11eb-997a-e566c5550886","first_name":"Alban"},{"first_name":"Johannes","last_name":"Broichhagen","full_name":"Broichhagen, Johannes"},{"full_name":"Grant, Seth G.N.","last_name":"Grant","first_name":"Seth G.N."},{"last_name":"Jonas","full_name":"Jonas, Peter M","id":"353C1B58-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-5001-4804","first_name":"Peter M"},{"id":"3E57A680-F248-11E8-B48F-1D18A9856A87","full_name":"Novarino, Gaia","last_name":"Novarino","first_name":"Gaia","orcid":"0000-0002-7673-7178"},{"full_name":"Pfister, Hanspeter","last_name":"Pfister","first_name":"Hanspeter"},{"first_name":"Bernd","orcid":"0000-0001-6511-9385","last_name":"Bickel","full_name":"Bickel, Bernd","id":"49876194-F248-11E8-B48F-1D18A9856A87"},{"id":"42EFD3B6-F248-11E8-B48F-1D18A9856A87","full_name":"Danzl, Johann G","last_name":"Danzl","first_name":"Johann G","orcid":"0000-0001-8559-3973"}],"day":"01","scopus_import":"1","article_type":"original","date_created":"2023-07-23T22:01:13Z","volume":20,"abstract":[{"lang":"eng","text":"Three-dimensional (3D) reconstruction of living brain tissue down to an individual synapse level would create opportunities for decoding the dynamics and structure–function relationships of the brain’s complex and dense information processing network; however, this has been hindered by insufficient 3D resolution, inadequate signal-to-noise ratio and prohibitive light burden in optical imaging, whereas electron microscopy is inherently static. Here we solved these challenges by developing an integrated optical/machine-learning technology, LIONESS (live information-optimized nanoscopy enabling saturated segmentation). This leverages optical modifications to stimulated emission depletion microscopy in comprehensively, extracellularly labeled tissue and previous information on sample structure via machine learning to simultaneously achieve isotropic super-resolution, high signal-to-noise ratio and compatibility with living tissue. This allows dense deep-learning-based instance segmentation and 3D reconstruction at a synapse level, incorporating molecular, activity and morphodynamic information. LIONESS opens up avenues for studying the dynamic functional (nano-)architecture of living brain tissue."}],"intvolume":"        20","acknowledged_ssus":[{"_id":"ScienComp"},{"_id":"Bio"},{"_id":"PreCl"},{"_id":"E-Lib"},{"_id":"LifeSc"},{"_id":"M-Shop"}],"publication_status":"published","publication_identifier":{"eissn":["1548-7105"],"issn":["1548-7091"]},"external_id":{"pmid":["37429995"],"isi":["001025621500001"]},"related_material":{"record":[{"id":"12817","status":"public","relation":"research_data"},{"status":"public","relation":"shorter_version","id":"14770"}],"link":[{"relation":"software","url":"https://github.com/danzllab/LIONESS"}]},"isi":1,"year":"2023","project":[{"_id":"265CB4D0-B435-11E9-9278-68D0E5697425","grant_number":"I03600","name":"Optical control of synaptic function via adhesion molecules","call_identifier":"FWF"},{"_id":"2548AE96-B435-11E9-9278-68D0E5697425","call_identifier":"FWF","grant_number":"W1232-B24","name":"Molecular Drug Targets"},{"_id":"25C5A090-B435-11E9-9278-68D0E5697425","call_identifier":"FWF","name":"The Wittgenstein Prize","grant_number":"Z00312"},{"name":"High content imaging to decode human immune cell interactions in health and allergic disease","_id":"23889792-32DE-11EA-91FC-C7463DDC885E"},{"name":"International IST Doctoral Program","grant_number":"665385","call_identifier":"H2020","_id":"2564DBCA-B435-11E9-9278-68D0E5697425"},{"name":"MATERIALIZABLE: Intelligent fabrication-oriented Computational Design and Modeling","grant_number":"715767","call_identifier":"H2020","_id":"24F9549A-B435-11E9-9278-68D0E5697425"},{"name":"Probing the Reversibility of Autism Spectrum Disorders by Employing in vivo and in vitro Models","grant_number":"715508","call_identifier":"H2020","_id":"25444568-B435-11E9-9278-68D0E5697425"},{"_id":"25B7EB9E-B435-11E9-9278-68D0E5697425","grant_number":"692692","name":"Biophysics and circuit function of a giant cortical glumatergic synapse","call_identifier":"H2020"},{"_id":"fc2be41b-9c52-11eb-aca3-faa90aa144e9","name":"Synaptic computations of the hippocampal CA3 circuitry","grant_number":"101026635","call_identifier":"H2020"},{"grant_number":"LT00057","name":"High-speed 3D-nanoscopy to study the role of adhesion during 3D cell migration","_id":"2668BFA0-B435-11E9-9278-68D0E5697425"}],"publication":"Nature Methods","status":"public","acknowledgement":"We thank J. Vorlaufer, N. Agudelo and A. Wartak for microscope maintenance and troubleshooting, C. Kreuzinger and A. Freeman for technical assistance, M. Šuplata for hardware control support and M. Cunha dos Santos for initial exploration of software. We\r\nthank P. Henderson for advice on deep-learning training and M. Sixt, S. Boyd and T. Weiss for discussions and critical reading of the manuscript. L. Lavis (Janelia Research Campus) generously provided the JF585-HaloTag ligand. We acknowledge expert support by IST\r\nAustria’s scientific computing, imaging and optics, preclinical, library and laboratory support facilities and by the Miba machine shop. We gratefully acknowledge funding by the following sources: Austrian Science Fund (F.W.F.) grant no. I3600-B27 (J.G.D.), grant no. DK W1232\r\n(J.G.D. and J.M.M.) and grant no. Z 312-B27, Wittgenstein award (P.J.); the Gesellschaft für Forschungsförderung NÖ grant no. LSC18-022 (J.G.D.); an ISTA Interdisciplinary project grant (J.G.D. and B.B.); the European Union’s Horizon 2020 research and innovation programme,\r\nMarie-Skłodowska Curie grant 665385 (J.M.M. and J.L.); the European Union’s Horizon 2020 research and innovation programme, European Research Council grant no. 715767, MATERIALIZABLE (B.B.); grant no. 715508, REVERSEAUTISM (G.N.); grant no. 695568, SYNNOVATE (S.G.N.G.); and grant no. 692692, GIANTSYN (P.J.); the Simons\r\nFoundation Autism Research Initiative grant no. 529085 (S.G.N.G.); the Wellcome Trust Technology Development grant no. 202932 (S.G.N.G.); the Marie Skłodowska-Curie Actions Individual Fellowship no. 101026635 under the EU Horizon 2020 program (J.F.W.);\r\nthe Human Frontier Science Program postdoctoral fellowship LT000557/2018 (W.J.); and the National Science Foundation grant no. IIS-1835231 (H.P.) and NCS-FO-2124179 (H.P.).","date_published":"2023-08-01T00:00:00Z","pmid":1,"ec_funded":1,"publisher":"Springer Nature","doi":"10.1038/s41592-023-01936-6","article_processing_charge":"Yes","type":"journal_article","date_updated":"2024-01-10T08:37:48Z","_id":"13267","page":"1256-1265","quality_controlled":"1","main_file_link":[{"url":"https://doi.org/10.1038/s41592-023-01936-6","open_access":"1"}]},{"oa":1,"language":[{"iso":"eng"}],"citation":{"ama":"Michalska JM, Lyudchik J, Velicky P, et al. Imaging brain tissue architecture across millimeter to nanometer scales. <i>Nature Biotechnology</i>. 2023. doi:<a href=\"https://doi.org/10.1038/s41587-023-01911-8\">10.1038/s41587-023-01911-8</a>","ieee":"J. M. Michalska <i>et al.</i>, “Imaging brain tissue architecture across millimeter to nanometer scales,” <i>Nature Biotechnology</i>. Springer Nature, 2023.","short":"J.M. Michalska, J. Lyudchik, P. Velicky, H. Korinkova, J. Watson, A. Cenameri, C.M. Sommer, N. Amberg, A. Venturino, K. Roessler, T. Czech, R. Höftberger, S. Siegert, G. Novarino, P.M. Jonas, J.G. Danzl, Nature Biotechnology (2023).","ista":"Michalska JM, Lyudchik J, Velicky P, Korinkova H, Watson J, Cenameri A, Sommer CM, Amberg N, Venturino A, Roessler K, Czech T, Höftberger R, Siegert S, Novarino G, Jonas PM, Danzl JG. 2023. Imaging brain tissue architecture across millimeter to nanometer scales. Nature Biotechnology.","chicago":"Michalska, Julia M, Julia Lyudchik, Philipp Velicky, Hana Korinkova, Jake Watson, Alban Cenameri, Christoph M Sommer, et al. “Imaging Brain Tissue Architecture across Millimeter to Nanometer Scales.” <i>Nature Biotechnology</i>. Springer Nature, 2023. <a href=\"https://doi.org/10.1038/s41587-023-01911-8\">https://doi.org/10.1038/s41587-023-01911-8</a>.","apa":"Michalska, J. M., Lyudchik, J., Velicky, P., Korinkova, H., Watson, J., Cenameri, A., … Danzl, J. G. (2023). Imaging brain tissue architecture across millimeter to nanometer scales. <i>Nature Biotechnology</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41587-023-01911-8\">https://doi.org/10.1038/s41587-023-01911-8</a>","mla":"Michalska, Julia M., et al. “Imaging Brain Tissue Architecture across Millimeter to Nanometer Scales.” <i>Nature Biotechnology</i>, Springer Nature, 2023, doi:<a href=\"https://doi.org/10.1038/s41587-023-01911-8\">10.1038/s41587-023-01911-8</a>."},"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","month":"08","department":[{"_id":"SaSi"},{"_id":"GaNo"},{"_id":"PeJo"},{"_id":"JoDa"},{"_id":"Bio"},{"_id":"RySh"}],"abstract":[{"lang":"eng","text":"Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanometer synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS uses fixation-compatible extracellular labeling and optical imaging, including stimulated emission depletion or expansion microscopy, to comprehensively delineate cellular structures. It enables three-dimensional reconstruction of single synapses and mapping of synaptic connectivity by identification and analysis of putative synaptic cleft regions. Applying CATS to the mouse hippocampal mossy fiber circuitry, we reconstructed and quantified the synaptic input and output structure of identified neurons. We furthermore demonstrate applicability to clinically derived human tissue samples, including formalin-fixed paraffin-embedded routine diagnostic specimens, for visualizing the cellular architecture of brain tissue in health and disease."}],"acknowledged_ssus":[{"_id":"ScienComp"},{"_id":"Bio"},{"_id":"PreCl"},{"_id":"LifeSc"},{"_id":"M-Shop"},{"_id":"E-Lib"}],"publication_identifier":{"issn":["1087-0156"],"eissn":["1546-1696"]},"publication_status":"epub_ahead","author":[{"first_name":"Julia M","orcid":"0000-0003-3862-1235","id":"443DB6DE-F248-11E8-B48F-1D18A9856A87","full_name":"Michalska, Julia M","last_name":"Michalska"},{"id":"46E28B80-F248-11E8-B48F-1D18A9856A87","full_name":"Lyudchik, Julia","last_name":"Lyudchik","first_name":"Julia"},{"full_name":"Velicky, Philipp","id":"39BDC62C-F248-11E8-B48F-1D18A9856A87","last_name":"Velicky","orcid":"0000-0002-2340-7431","first_name":"Philipp"},{"first_name":"Hana","last_name":"Korinkova","id":"ee3cb6ca-ec98-11ea-ae11-ff703e2254ed","full_name":"Korinkova, Hana"},{"orcid":"0000-0002-8698-3823","first_name":"Jake","last_name":"Watson","full_name":"Watson, Jake","id":"63836096-4690-11EA-BD4E-32803DDC885E"},{"full_name":"Cenameri, Alban","id":"9ac8f577-2357-11eb-997a-e566c5550886","last_name":"Cenameri","first_name":"Alban"},{"full_name":"Sommer, Christoph M","id":"4DF26D8C-F248-11E8-B48F-1D18A9856A87","last_name":"Sommer","first_name":"Christoph M","orcid":"0000-0003-1216-9105"},{"last_name":"Amberg","id":"4CD6AAC6-F248-11E8-B48F-1D18A9856A87","full_name":"Amberg, Nicole","orcid":"0000-0002-3183-8207","first_name":"Nicole"},{"first_name":"Alessandro","orcid":"0000-0003-2356-9403","id":"41CB84B2-F248-11E8-B48F-1D18A9856A87","full_name":"Venturino, Alessandro","last_name":"Venturino"},{"first_name":"Karl","full_name":"Roessler, Karl","last_name":"Roessler"},{"last_name":"Czech","full_name":"Czech, Thomas","first_name":"Thomas"},{"last_name":"Höftberger","full_name":"Höftberger, Romana","first_name":"Romana"},{"full_name":"Siegert, Sandra","id":"36ACD32E-F248-11E8-B48F-1D18A9856A87","last_name":"Siegert","first_name":"Sandra","orcid":"0000-0001-8635-0877"},{"full_name":"Novarino, Gaia","id":"3E57A680-F248-11E8-B48F-1D18A9856A87","last_name":"Novarino","orcid":"0000-0002-7673-7178","first_name":"Gaia"},{"first_name":"Peter M","orcid":"0000-0001-5001-4804","last_name":"Jonas","id":"353C1B58-F248-11E8-B48F-1D18A9856A87","full_name":"Jonas, Peter M"},{"last_name":"Danzl","full_name":"Danzl, Johann G","id":"42EFD3B6-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-8559-3973","first_name":"Johann G"}],"scopus_import":"1","day":"31","title":"Imaging brain tissue architecture across millimeter to nanometer scales","oa_version":"Published Version","article_type":"original","date_created":"2023-09-03T22:01:15Z","project":[{"_id":"265CB4D0-B435-11E9-9278-68D0E5697425","name":"Optical control of synaptic function via adhesion molecules","grant_number":"I03600","call_identifier":"FWF"},{"_id":"2548AE96-B435-11E9-9278-68D0E5697425","call_identifier":"FWF","name":"Molecular Drug Targets","grant_number":"W1232-B24"},{"call_identifier":"FWF","grant_number":"Z00312","name":"The Wittgenstein Prize","_id":"25C5A090-B435-11E9-9278-68D0E5697425"},{"name":"High content imaging to decode human immune cell interactions in health and allergic disease","_id":"23889792-32DE-11EA-91FC-C7463DDC885E"},{"_id":"25444568-B435-11E9-9278-68D0E5697425","call_identifier":"H2020","name":"Probing the Reversibility of Autism Spectrum Disorders by Employing in vivo and in vitro Models","grant_number":"715508"},{"call_identifier":"H2020","name":"Biophysics and circuit function of a giant cortical glumatergic synapse","grant_number":"692692","_id":"25B7EB9E-B435-11E9-9278-68D0E5697425"},{"_id":"2564DBCA-B435-11E9-9278-68D0E5697425","grant_number":"665385","name":"International IST Doctoral Program","call_identifier":"H2020"},{"_id":"fc2be41b-9c52-11eb-aca3-faa90aa144e9","name":"Synaptic computations of the hippocampal CA3 circuitry","grant_number":"101026635","call_identifier":"H2020"}],"status":"public","publication":"Nature Biotechnology","ec_funded":1,"acknowledgement":"We thank J. Vorlaufer, N. Agudelo-Dueñas, W. Jahr and A. Wartak for microscope maintenance and troubleshooting; C. Kreuzinger, A. Freeman and I. Erber for technical assistance; and M. Tomschik for support with obtaining human samples. We gratefully acknowledge E. Miguel for setting up webKnossos and M. Šuplata for computational support and hardware control. We are grateful to R. Shigemoto and B. Bickel for generous support and M. Sixt and S. Boyd (Stanford University) for discussions and critical reading of the paper. PSD95-HaloTag mice were kindly provided by S. Grant (University of Edinburgh). We acknowledge expert support by Institute of Science and Technology Austria’s scientific computing, imaging and optics, preclinical and lab support facilities and by the Miba machine shop and library. We gratefully acknowledge funding by the following sources: Austrian Science Fund (FWF) grant I3600-B27 (J.G.D.); Austrian Science Fund (FWF) grant DK W1232 (J.G.D. and J.M.M.); Austrian Science Fund (FWF) grant Z 312-B27, Wittgenstein award (P.J.); Austrian Science Fund (FWF) projects I4685-B, I6565-B (SYNABS) and DOC 33-B27 (R.H.); Gesellschaft für Forschungsförderung NÖ (NFB) grant LSC18-022 (J.G.D.); European Union’s Horizon 2020 research and innovation programme, European Research Council (ERC) grant 715508 – REVERSEAUTISM (G.N.); European Union’s Horizon 2020 research and innovation programme, European Research Council (ERC) grant 692692 – GIANTSYN (P.J.); Marie Skłodowska-Curie Actions Fellowship GA no. 665385 under the EU Horizon 2020 program (J.M.M. and J.L.); and Marie Skłodowska-Curie Actions Individual Fellowship no. 101026635 under the EU Horizon 2020 program (J.F.W.).","date_published":"2023-08-31T00:00:00Z","year":"2023","isi":1,"external_id":{"isi":["001065254200001"]},"related_material":{"link":[{"url":"https://github.com/danzllab/CATS","relation":"software"}],"record":[{"status":"public","relation":"research_data","id":"13126"}]},"main_file_link":[{"url":"https://doi.org/10.1038/s41587-023-01911-8","open_access":"1"}],"quality_controlled":"1","doi":"10.1038/s41587-023-01911-8","article_processing_charge":"Yes (in subscription journal)","publisher":"Springer Nature","date_updated":"2024-02-21T12:18:18Z","_id":"14257","type":"journal_article"},{"doi":"10.1007/978-1-0716-3004-4_3","article_processing_charge":"No","alternative_title":["Methods in Molecular Biology"],"editor":[{"first_name":"Garry","last_name":"Scarlett","full_name":"Scarlett, Garry"}],"publisher":"Springer Nature","date_updated":"2023-03-16T08:34:24Z","_id":"12720","type":"book_chapter","series_title":"MIMB","page":"33-44","quality_controlled":"1","year":"2023","external_id":{"pmid":["36853454"]},"publication":"DNA Manipulation and Analysis","status":"public","pmid":1,"date_published":"2023-03-01T00:00:00Z","author":[{"last_name":"Arroyo-Urea","full_name":"Arroyo-Urea, Sandra","first_name":"Sandra"},{"last_name":"Watson","id":"63836096-4690-11EA-BD4E-32803DDC885E","full_name":"Watson, Jake","orcid":"0000-0002-8698-3823","first_name":"Jake"},{"first_name":"Javier","full_name":"García-Nafría, Javier","last_name":"García-Nafría"}],"scopus_import":"1","day":"01","title":"Molecular Cloning Using In Vivo DNA Assembly","oa_version":"None","volume":2633,"date_created":"2023-03-12T23:01:02Z","abstract":[{"text":"Here we describe the in vivo DNA assembly approach, where molecular cloning procedures are performed using an E. coli recA-independent recombination pathway, which assembles linear fragments of DNA with short homologous termini. This pathway is present in all standard laboratory E. coli strains and, by bypassing the need for in vitro DNA assembly, allows simplified molecular cloning to be performed without the plasmid instability issues associated with specialized recombination-cloning bacterial strains. The methodology requires specific primer design and can perform all standard plasmid modifications (insertions, deletions, mutagenesis, and sub-cloning) in a rapid, simple, and cost-efficient manner, as it does not require commercial kits or specialized bacterial strains. Additionally, this approach can be used to perform complex procedures such as multiple modifications to a plasmid, as up to 6 linear fragments can be assembled in vivo by this recombination pathway. Procedures generally require less than 3 h, involving PCR amplification, DpnI digestion of template DNA, and transformation, upon which circular plasmids are assembled. In this chapter we describe the requirements, procedure, and potential pitfalls when using this technique, as well as protocol variations to overcome the most common issues.","lang":"eng"}],"intvolume":"      2633","publication_identifier":{"eissn":["1940-6029"],"eisbn":["978-1-0716-3004-4"],"issn":["1064-3745"],"isbn":["978-1-0716-3003-7"]},"publication_status":"published","month":"03","department":[{"_id":"PeJo"}],"language":[{"iso":"eng"}],"place":"New York, NY, United States","citation":{"apa":"Arroyo-Urea, S., Watson, J., &#38; García-Nafría, J. (2023). Molecular Cloning Using In Vivo DNA Assembly. In G. Scarlett (Ed.), <i>DNA Manipulation and Analysis</i> (Vol. 2633, pp. 33–44). New York, NY, United States: Springer Nature. <a href=\"https://doi.org/10.1007/978-1-0716-3004-4_3\">https://doi.org/10.1007/978-1-0716-3004-4_3</a>","mla":"Arroyo-Urea, Sandra, et al. “Molecular Cloning Using In Vivo DNA Assembly.” <i>DNA Manipulation and Analysis</i>, edited by Garry Scarlett, vol. 2633, Springer Nature, 2023, pp. 33–44, doi:<a href=\"https://doi.org/10.1007/978-1-0716-3004-4_3\">10.1007/978-1-0716-3004-4_3</a>.","ista":"Arroyo-Urea S, Watson J, García-Nafría J. 2023.Molecular Cloning Using In Vivo DNA Assembly. In: DNA Manipulation and Analysis. Methods in Molecular Biology, vol. 2633, 33–44.","chicago":"Arroyo-Urea, Sandra, Jake Watson, and Javier García-Nafría. “Molecular Cloning Using In Vivo DNA Assembly.” In <i>DNA Manipulation and Analysis</i>, edited by Garry Scarlett, 2633:33–44. MIMB. New York, NY, United States: Springer Nature, 2023. <a href=\"https://doi.org/10.1007/978-1-0716-3004-4_3\">https://doi.org/10.1007/978-1-0716-3004-4_3</a>.","ieee":"S. Arroyo-Urea, J. Watson, and J. García-Nafría, “Molecular Cloning Using In Vivo DNA Assembly,” in <i>DNA Manipulation and Analysis</i>, vol. 2633, G. Scarlett, Ed. New York, NY, United States: Springer Nature, 2023, pp. 33–44.","short":"S. Arroyo-Urea, J. Watson, J. García-Nafría, in:, G. Scarlett (Ed.), DNA Manipulation and Analysis, Springer Nature, New York, NY, United States, 2023, pp. 33–44.","ama":"Arroyo-Urea S, Watson J, García-Nafría J. Molecular Cloning Using In Vivo DNA Assembly. In: Scarlett G, ed. <i>DNA Manipulation and Analysis</i>. Vol 2633. MIMB. New York, NY, United States: Springer Nature; 2023:33-44. doi:<a href=\"https://doi.org/10.1007/978-1-0716-3004-4_3\">10.1007/978-1-0716-3004-4_3</a>"},"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87"},{"year":"2023","isi":1,"external_id":{"isi":["001066658700003"]},"publication":"Nature Communications","status":"public","acknowledgement":"We thank James Krieger for generating the ‘proDy’ interaction maps in Fig. 5B and S7C, and Jan-Niklas Dohrke for critically reading the manuscript. We thank members of the Greger lab for insightful comments during this study. We acknowledge Trevor Rutherford for confirming ligand integrity by NMR. We are also grateful to LMB scientific computing and the EM facility for their support. This research was funded in part by the Wellcome Trust (223194/Z/21/Z) to I.H.G. For the purpose of Open Access, the MRC Laboratory of Molecular Biology has applied a CC BY public copyright licence to any Author Accepted Manuscript (AAM) version arising from this submission. Further funding came from the Medical Research Council (MRU105174197) to I.H.G, and NIH grant (R56/R01MH123474) to T.N.","date_published":"2023-03-25T00:00:00Z","article_processing_charge":"No","doi":"10.1038/s41467-023-37259-5","publisher":"Springer Nature","_id":"12786","date_updated":"2023-12-13T11:15:58Z","type":"journal_article","ddc":["570"],"quality_controlled":"1","month":"03","department":[{"_id":"PeJo"}],"article_number":"1659","file":[{"file_id":"12797","creator":"dernst","date_updated":"2023-04-03T06:38:56Z","file_size":2613996,"date_created":"2023-04-03T06:38:56Z","checksum":"0a97b31191432dae5853bbb5ccb7698d","relation":"main_file","access_level":"open_access","content_type":"application/pdf","success":1,"file_name":"2023_NatureComm_Zhang.pdf"}],"oa":1,"language":[{"iso":"eng"}],"citation":{"ama":"Zhang D, Lape R, Shaikh SA, et al. Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics. <i>Nature Communications</i>. 2023;14. doi:<a href=\"https://doi.org/10.1038/s41467-023-37259-5\">10.1038/s41467-023-37259-5</a>","ieee":"D. Zhang <i>et al.</i>, “Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics,” <i>Nature Communications</i>, vol. 14. Springer Nature, 2023.","short":"D. Zhang, R. Lape, S.A. Shaikh, B.K. Kohegyi, J. Watson, O. Cais, T. Nakagawa, I.H. Greger, Nature Communications 14 (2023).","chicago":"Zhang, Danyang, Remigijus Lape, Saher A. Shaikh, Bianka K. Kohegyi, Jake Watson, Ondrej Cais, Terunaga Nakagawa, and Ingo H. Greger. “Modulatory Mechanisms of TARP Γ8-Selective AMPA Receptor Therapeutics.” <i>Nature Communications</i>. Springer Nature, 2023. <a href=\"https://doi.org/10.1038/s41467-023-37259-5\">https://doi.org/10.1038/s41467-023-37259-5</a>.","ista":"Zhang D, Lape R, Shaikh SA, Kohegyi BK, Watson J, Cais O, Nakagawa T, Greger IH. 2023. Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics. Nature Communications. 14, 1659.","apa":"Zhang, D., Lape, R., Shaikh, S. A., Kohegyi, B. K., Watson, J., Cais, O., … Greger, I. H. (2023). Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics. <i>Nature Communications</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41467-023-37259-5\">https://doi.org/10.1038/s41467-023-37259-5</a>","mla":"Zhang, Danyang, et al. “Modulatory Mechanisms of TARP Γ8-Selective AMPA Receptor Therapeutics.” <i>Nature Communications</i>, vol. 14, 1659, Springer Nature, 2023, doi:<a href=\"https://doi.org/10.1038/s41467-023-37259-5\">10.1038/s41467-023-37259-5</a>."},"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","scopus_import":"1","day":"25","author":[{"first_name":"Danyang","last_name":"Zhang","full_name":"Zhang, Danyang"},{"first_name":"Remigijus","full_name":"Lape, Remigijus","last_name":"Lape"},{"full_name":"Shaikh, Saher A.","last_name":"Shaikh","first_name":"Saher A."},{"first_name":"Bianka K.","full_name":"Kohegyi, Bianka K.","last_name":"Kohegyi"},{"first_name":"Jake","orcid":"0000-0002-8698-3823","id":"63836096-4690-11EA-BD4E-32803DDC885E","full_name":"Watson, Jake","last_name":"Watson"},{"first_name":"Ondrej","full_name":"Cais, Ondrej","last_name":"Cais"},{"first_name":"Terunaga","last_name":"Nakagawa","full_name":"Nakagawa, Terunaga"},{"first_name":"Ingo H.","last_name":"Greger","full_name":"Greger, Ingo H."}],"oa_version":"Published Version","title":"Modulatory mechanisms of TARP γ8-selective AMPA receptor therapeutics","volume":14,"date_created":"2023-04-02T22:01:09Z","article_type":"original","has_accepted_license":"1","tmp":{"name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","short":"CC BY (4.0)"},"abstract":[{"text":"AMPA glutamate receptors (AMPARs) mediate excitatory neurotransmission throughout the brain. Their signalling is uniquely diversified by brain region-specific auxiliary subunits, providing an opportunity for the development of selective therapeutics. AMPARs associated with TARP γ8 are enriched in the hippocampus, and are targets of emerging anti-epileptic drugs. To understand their therapeutic activity, we determined cryo-EM structures of the GluA1/2-γ8 receptor associated with three potent, chemically diverse ligands. We find that despite sharing a lipid-exposed and water-accessible binding pocket, drug action is differentially affected by binding-site mutants. Together with patch-clamp recordings and MD simulations we also demonstrate that ligand-triggered reorganisation of the AMPAR-TARP interface contributes to modulation. Unexpectedly, one ligand (JNJ-61432059) acts bifunctionally, negatively affecting GluA1 but exerting positive modulatory action on GluA2-containing AMPARs, in a TARP stoichiometry-dependent manner. These results further illuminate the action of TARPs, demonstrate the sensitive balance between positive and negative modulatory action, and provide a mechanistic platform for development of both positive and negative selective AMPAR modulators.","lang":"eng"}],"intvolume":"        14","file_date_updated":"2023-04-03T06:38:56Z","publication_identifier":{"eissn":["2041-1723"]},"publication_status":"published"},{"year":"2022","isi":1,"external_id":{"isi":["000757297200008"],"pmid":["35136046"]},"pmid":1,"acknowledgement":"We thank Ondrej Cais for critical reading of the manuscript. We are grateful to LMB\r\nscientific computing and the EM facility for support, Paul Emsley for help with model\r\nbuilding and Takanori Nakane for helpful comments with Relion 3.1. This work was\r\nsupported by grants from the Medical Research Council (MC_U105174197) and BBSRC\r\n(BB/N002113/1) to I.H.G, and grants from the MCIN/AEI/ 10.13039/501100011033 and\r\n“ESF Investing in your future” to B.H (PID2019-106284GA-I00 and RYC2018-025720-I).","date_published":"2022-02-08T00:00:00Z","publication":"Nature Communications","status":"public","date_updated":"2023-08-02T14:25:33Z","_id":"10763","type":"journal_article","doi":"10.1038/s41467-022-28404-7","article_processing_charge":"No","publisher":"Springer Nature","quality_controlled":"1","ddc":["570"],"department":[{"_id":"PeJo"}],"article_number":"734","file":[{"file_size":2625540,"date_created":"2022-02-21T07:59:32Z","creator":"dernst","date_updated":"2022-02-21T07:59:32Z","file_id":"10778","file_name":"2022_NatureCommunications_Herguedas.pdf","success":1,"content_type":"application/pdf","access_level":"open_access","relation":"main_file","checksum":"d86ee8eabe8b794730729ffbb1a8832e"}],"month":"02","citation":{"ama":"Herguedas B, Kohegyi BK, Dohrke JN, et al. Mechanisms underlying TARP modulation of the GluA1/2-γ8 AMPA receptor. <i>Nature Communications</i>. 2022;13. doi:<a href=\"https://doi.org/10.1038/s41467-022-28404-7\">10.1038/s41467-022-28404-7</a>","short":"B. Herguedas, B.K. Kohegyi, J.N. Dohrke, J. Watson, D. Zhang, H. Ho, S.A. Shaikh, R. Lape, J.M. Krieger, I.H. Greger, Nature Communications 13 (2022).","ieee":"B. Herguedas <i>et al.</i>, “Mechanisms underlying TARP modulation of the GluA1/2-γ8 AMPA receptor,” <i>Nature Communications</i>, vol. 13. Springer Nature, 2022.","chicago":"Herguedas, Beatriz, Bianka K. Kohegyi, Jan Niklas Dohrke, Jake Watson, Danyang Zhang, Hinze Ho, Saher A. Shaikh, Remigijus Lape, James M. Krieger, and Ingo H. Greger. “Mechanisms Underlying TARP Modulation of the GluA1/2-Γ8 AMPA Receptor.” <i>Nature Communications</i>. Springer Nature, 2022. <a href=\"https://doi.org/10.1038/s41467-022-28404-7\">https://doi.org/10.1038/s41467-022-28404-7</a>.","ista":"Herguedas B, Kohegyi BK, Dohrke JN, Watson J, Zhang D, Ho H, Shaikh SA, Lape R, Krieger JM, Greger IH. 2022. Mechanisms underlying TARP modulation of the GluA1/2-γ8 AMPA receptor. Nature Communications. 13, 734.","mla":"Herguedas, Beatriz, et al. “Mechanisms Underlying TARP Modulation of the GluA1/2-Γ8 AMPA Receptor.” <i>Nature Communications</i>, vol. 13, 734, Springer Nature, 2022, doi:<a href=\"https://doi.org/10.1038/s41467-022-28404-7\">10.1038/s41467-022-28404-7</a>.","apa":"Herguedas, B., Kohegyi, B. K., Dohrke, J. N., Watson, J., Zhang, D., Ho, H., … Greger, I. H. (2022). Mechanisms underlying TARP modulation of the GluA1/2-γ8 AMPA receptor. <i>Nature Communications</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41467-022-28404-7\">https://doi.org/10.1038/s41467-022-28404-7</a>"},"user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","oa":1,"language":[{"iso":"eng"}],"volume":13,"article_type":"original","date_created":"2022-02-20T23:01:30Z","author":[{"first_name":"Beatriz","full_name":"Herguedas, Beatriz","last_name":"Herguedas"},{"last_name":"Kohegyi","full_name":"Kohegyi, Bianka K.","first_name":"Bianka K."},{"full_name":"Dohrke, Jan Niklas","last_name":"Dohrke","first_name":"Jan Niklas"},{"full_name":"Watson, Jake","id":"63836096-4690-11EA-BD4E-32803DDC885E","last_name":"Watson","orcid":"0000-0002-8698-3823","first_name":"Jake"},{"last_name":"Zhang","full_name":"Zhang, Danyang","first_name":"Danyang"},{"first_name":"Hinze","full_name":"Ho, Hinze","last_name":"Ho"},{"first_name":"Saher A.","last_name":"Shaikh","full_name":"Shaikh, Saher A."},{"first_name":"Remigijus","last_name":"Lape","full_name":"Lape, Remigijus"},{"first_name":"James M.","full_name":"Krieger, James M.","last_name":"Krieger"},{"first_name":"Ingo H.","full_name":"Greger, Ingo H.","last_name":"Greger"}],"day":"08","scopus_import":"1","title":"Mechanisms underlying TARP modulation of the GluA1/2-γ8 AMPA receptor","oa_version":"Published Version","publication_status":"published","publication_identifier":{"eissn":["20411723"]},"file_date_updated":"2022-02-21T07:59:32Z","has_accepted_license":"1","abstract":[{"text":"AMPA-type glutamate receptors (AMPARs) mediate rapid signal transmission at excitatory\r\nsynapses in the brain. Glutamate binding to the receptor’s ligand-binding domains (LBDs)\r\nleads to ion channel activation and desensitization. Gating kinetics shape synaptic transmission\r\nand are strongly modulated by transmembrane AMPAR regulatory proteins (TARPs)\r\nthrough currently incompletely resolved mechanisms. Here, electron cryo-microscopy\r\nstructures of the GluA1/2 TARP-γ8 complex, in both open and desensitized states\r\n(at 3.5 Å), reveal state-selective engagement of the LBDs by the large TARP-γ8 loop (‘β1’),\r\nelucidating how this TARP stabilizes specific gating states. We further show how TARPs alter\r\nchannel rectification, by interacting with the pore helix of the selectivity filter. Lastly, we\r\nreveal that the Q/R-editing site couples the channel constriction at the filter entrance to the\r\ngate, and forms the major cation binding site in the conduction path. Our results provide a\r\nmechanistic framework of how TARPs modulate AMPAR gating and conductance.","lang":"eng"}],"tmp":{"name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","short":"CC BY (4.0)"},"intvolume":"        13"},{"user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","date_published":"2022-05-09T00:00:00Z","citation":{"mla":"Velicky, Philipp, et al. “Saturated Reconstruction of Living Brain Tissue.” <i>BioRxiv</i>, Cold Spring Harbor Laboratory, doi:<a href=\"https://doi.org/10.1101/2022.03.16.484431\">10.1101/2022.03.16.484431</a>.","apa":"Velicky, P., Miguel Villalba, E., Michalska, J. M., Wei, D., Lin, Z., Watson, J., … Danzl, J. G. (n.d.). Saturated reconstruction of living brain tissue. <i>bioRxiv</i>. Cold Spring Harbor Laboratory. <a href=\"https://doi.org/10.1101/2022.03.16.484431\">https://doi.org/10.1101/2022.03.16.484431</a>","chicago":"Velicky, Philipp, Eder Miguel Villalba, Julia M Michalska, Donglai Wei, Zudi Lin, Jake Watson, Jakob Troidl, et al. “Saturated Reconstruction of Living Brain Tissue.” <i>BioRxiv</i>. Cold Spring Harbor Laboratory, n.d. <a href=\"https://doi.org/10.1101/2022.03.16.484431\">https://doi.org/10.1101/2022.03.16.484431</a>.","ista":"Velicky P, Miguel Villalba E, Michalska JM, Wei D, Lin Z, Watson J, Troidl J, Beyer J, Ben Simon Y, Sommer CM, Jahr W, Cenameri A, Broichhagen J, Grant SGN, Jonas PM, Novarino G, Pfister H, Bickel B, Danzl JG. Saturated reconstruction of living brain tissue. bioRxiv, <a href=\"https://doi.org/10.1101/2022.03.16.484431\">10.1101/2022.03.16.484431</a>.","short":"P. Velicky, E. Miguel Villalba, J.M. Michalska, D. Wei, Z. Lin, J. Watson, J. Troidl, J. Beyer, Y. Ben Simon, C.M. Sommer, W. Jahr, A. Cenameri, J. Broichhagen, S.G.N. Grant, P.M. Jonas, G. Novarino, H. Pfister, B. Bickel, J.G. Danzl, BioRxiv (n.d.).","ieee":"P. Velicky <i>et al.</i>, “Saturated reconstruction of living brain tissue,” <i>bioRxiv</i>. Cold Spring Harbor Laboratory.","ama":"Velicky P, Miguel Villalba E, Michalska JM, et al. Saturated reconstruction of living brain tissue. <i>bioRxiv</i>. doi:<a href=\"https://doi.org/10.1101/2022.03.16.484431\">10.1101/2022.03.16.484431</a>"},"publication":"bioRxiv","status":"public","language":[{"iso":"eng"}],"oa":1,"department":[{"_id":"PeJo"},{"_id":"GaNo"},{"_id":"BeBi"},{"_id":"JoDa"}],"related_material":{"record":[{"id":"12470","status":"public","relation":"dissertation_contains"}]},"month":"05","year":"2022","publication_status":"submitted","main_file_link":[{"url":"https://doi.org/10.1101/2022.03.16.484431","open_access":"1"}],"abstract":[{"lang":"eng","text":"Complex wiring between neurons underlies the information-processing network enabling all brain functions, including cognition and memory. For understanding how the network is structured, processes information, and changes over time, comprehensive visualization of the architecture of living brain tissue with its cellular and molecular components would open up major opportunities. However, electron microscopy (EM) provides nanometre-scale resolution required for full <jats:italic>in-silico</jats:italic> reconstruction<jats:sup>1–5</jats:sup>, yet is limited to fixed specimens and static representations. Light microscopy allows live observation, with super-resolution approaches<jats:sup>6–12</jats:sup> facilitating nanoscale visualization, but comprehensive 3D-reconstruction of living brain tissue has been hindered by tissue photo-burden, photobleaching, insufficient 3D-resolution, and inadequate signal-to-noise ratio (SNR). Here we demonstrate saturated reconstruction of living brain tissue. We developed an integrated imaging and analysis technology, adapting stimulated emission depletion (STED) microscopy<jats:sup>6,13</jats:sup> in extracellularly labelled tissue<jats:sup>14</jats:sup> for high SNR and near-isotropic resolution. Centrally, a two-stage deep-learning approach leveraged previously obtained information on sample structure to drastically reduce photo-burden and enable automated volumetric reconstruction down to single synapse level. Live reconstruction provides unbiased analysis of tissue architecture across time in relation to functional activity and targeted activation, and contextual understanding of molecular labelling. This adoptable technology will facilitate novel insights into the dynamic functional architecture of living brain tissue."}],"date_created":"2022-08-23T11:07:59Z","type":"preprint","_id":"11943","date_updated":"2024-03-25T23:30:11Z","oa_version":"Preprint","publisher":"Cold Spring Harbor Laboratory","title":"Saturated reconstruction of living brain tissue","day":"09","article_processing_charge":"No","doi":"10.1101/2022.03.16.484431","author":[{"orcid":"0000-0002-2340-7431","first_name":"Philipp","full_name":"Velicky, Philipp","id":"39BDC62C-F248-11E8-B48F-1D18A9856A87","last_name":"Velicky"},{"id":"3FB91342-F248-11E8-B48F-1D18A9856A87","full_name":"Miguel Villalba, Eder","last_name":"Miguel Villalba","first_name":"Eder","orcid":"0000-0001-5665-0430"},{"full_name":"Michalska, Julia M","id":"443DB6DE-F248-11E8-B48F-1D18A9856A87","last_name":"Michalska","orcid":"0000-0003-3862-1235","first_name":"Julia M"},{"last_name":"Wei","full_name":"Wei, Donglai","first_name":"Donglai"},{"last_name":"Lin","full_name":"Lin, Zudi","first_name":"Zudi"},{"orcid":"0000-0002-8698-3823","first_name":"Jake","last_name":"Watson","full_name":"Watson, Jake","id":"63836096-4690-11EA-BD4E-32803DDC885E"},{"last_name":"Troidl","full_name":"Troidl, Jakob","first_name":"Jakob"},{"last_name":"Beyer","full_name":"Beyer, Johanna","first_name":"Johanna"},{"full_name":"Ben Simon, Yoav","id":"43DF3136-F248-11E8-B48F-1D18A9856A87","last_name":"Ben Simon","first_name":"Yoav"},{"orcid":"0000-0003-1216-9105","first_name":"Christoph M","last_name":"Sommer","full_name":"Sommer, Christoph M","id":"4DF26D8C-F248-11E8-B48F-1D18A9856A87"},{"last_name":"Jahr","full_name":"Jahr, Wiebke","id":"425C1CE8-F248-11E8-B48F-1D18A9856A87","first_name":"Wiebke"},{"last_name":"Cenameri","full_name":"Cenameri, Alban","id":"9ac8f577-2357-11eb-997a-e566c5550886","first_name":"Alban"},{"first_name":"Johannes","last_name":"Broichhagen","full_name":"Broichhagen, Johannes"},{"full_name":"Grant, Seth G. N.","last_name":"Grant","first_name":"Seth G. N."},{"first_name":"Peter M","orcid":"0000-0001-5001-4804","last_name":"Jonas","full_name":"Jonas, Peter M","id":"353C1B58-F248-11E8-B48F-1D18A9856A87"},{"first_name":"Gaia","orcid":"0000-0002-7673-7178","last_name":"Novarino","id":"3E57A680-F248-11E8-B48F-1D18A9856A87","full_name":"Novarino, Gaia"},{"first_name":"Hanspeter","last_name":"Pfister","full_name":"Pfister, Hanspeter"},{"full_name":"Bickel, Bernd","id":"49876194-F248-11E8-B48F-1D18A9856A87","last_name":"Bickel","orcid":"0000-0001-6511-9385","first_name":"Bernd"},{"last_name":"Danzl","id":"42EFD3B6-F248-11E8-B48F-1D18A9856A87","full_name":"Danzl, Johann G","orcid":"0000-0001-8559-3973","first_name":"Johann G"}]},{"date_updated":"2024-03-25T23:30:11Z","_id":"11950","type":"preprint","date_created":"2022-08-24T08:24:52Z","doi":"10.1101/2022.08.17.504272","author":[{"first_name":"Julia M","orcid":"0000-0003-3862-1235","last_name":"Michalska","id":"443DB6DE-F248-11E8-B48F-1D18A9856A87","full_name":"Michalska, Julia M"},{"full_name":"Lyudchik, Julia","id":"46E28B80-F248-11E8-B48F-1D18A9856A87","last_name":"Lyudchik","first_name":"Julia"},{"first_name":"Philipp","orcid":"0000-0002-2340-7431","full_name":"Velicky, Philipp","id":"39BDC62C-F248-11E8-B48F-1D18A9856A87","last_name":"Velicky"},{"full_name":"Korinkova, Hana","id":"ee3cb6ca-ec98-11ea-ae11-ff703e2254ed","last_name":"Korinkova","first_name":"Hana"},{"last_name":"Watson","id":"63836096-4690-11EA-BD4E-32803DDC885E","full_name":"Watson, Jake","orcid":"0000-0002-8698-3823","first_name":"Jake"},{"first_name":"Alban","full_name":"Cenameri, Alban","id":"9ac8f577-2357-11eb-997a-e566c5550886","last_name":"Cenameri"},{"orcid":"0000-0003-1216-9105","first_name":"Christoph M","full_name":"Sommer, Christoph M","id":"4DF26D8C-F248-11E8-B48F-1D18A9856A87","last_name":"Sommer"},{"first_name":"Alessandro","orcid":"0000-0003-2356-9403","id":"41CB84B2-F248-11E8-B48F-1D18A9856A87","full_name":"Venturino, Alessandro","last_name":"Venturino"},{"first_name":"Karl","full_name":"Roessler, Karl","last_name":"Roessler"},{"full_name":"Czech, Thomas","last_name":"Czech","first_name":"Thomas"},{"orcid":"0000-0001-8635-0877","first_name":"Sandra","last_name":"Siegert","id":"36ACD32E-F248-11E8-B48F-1D18A9856A87","full_name":"Siegert, Sandra"},{"id":"3E57A680-F248-11E8-B48F-1D18A9856A87","full_name":"Novarino, Gaia","last_name":"Novarino","orcid":"0000-0002-7673-7178","first_name":"Gaia"},{"last_name":"Jonas","full_name":"Jonas, Peter M","id":"353C1B58-F248-11E8-B48F-1D18A9856A87","orcid":"0000-0001-5001-4804","first_name":"Peter M"},{"id":"42EFD3B6-F248-11E8-B48F-1D18A9856A87","full_name":"Danzl, Johann G","last_name":"Danzl","first_name":"Johann G","orcid":"0000-0001-8559-3973"}],"day":"18","article_processing_charge":"No","title":"Uncovering brain tissue architecture across scales with super-resolution light microscopy","publisher":"Cold Spring Harbor Laboratory","oa_version":"Preprint","main_file_link":[{"open_access":"1","url":"https://doi.org/10.1101/2022.08.17.504272"}],"publication_status":"submitted","abstract":[{"text":"Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanoscopic synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS leverages fixation-compatible extracellular labeling and advanced optical readout, in particular stimulated-emission depletion and expansion microscopy, to comprehensively delineate cellular structures. It enables 3D-reconstructing single synapses and mapping synaptic connectivity by identification and tailored analysis of putative synaptic cleft regions. Applying CATS to the hippocampal mossy fiber circuitry, we demonstrate its power to reveal the system’s molecularly informed ultrastructure across spatial scales and assess local connectivity by reconstructing and quantifying the synaptic input and output structure of identified neurons.","lang":"eng"}],"department":[{"_id":"SaSi"},{"_id":"GaNo"},{"_id":"PeJo"},{"_id":"JoDa"}],"year":"2022","related_material":{"record":[{"id":"12470","relation":"dissertation_contains","status":"public"}]},"month":"08","citation":{"chicago":"Michalska, Julia M, Julia Lyudchik, Philipp Velicky, Hana Korinkova, Jake Watson, Alban Cenameri, Christoph M Sommer, et al. “Uncovering Brain Tissue Architecture across Scales with Super-Resolution Light Microscopy.” <i>BioRxiv</i>. Cold Spring Harbor Laboratory, n.d. <a href=\"https://doi.org/10.1101/2022.08.17.504272\">https://doi.org/10.1101/2022.08.17.504272</a>.","ista":"Michalska JM, Lyudchik J, Velicky P, Korinkova H, Watson J, Cenameri A, Sommer CM, Venturino A, Roessler K, Czech T, Siegert S, Novarino G, Jonas PM, Danzl JG. Uncovering brain tissue architecture across scales with super-resolution light microscopy. bioRxiv, <a href=\"https://doi.org/10.1101/2022.08.17.504272\">10.1101/2022.08.17.504272</a>.","mla":"Michalska, Julia M., et al. “Uncovering Brain Tissue Architecture across Scales with Super-Resolution Light Microscopy.” <i>BioRxiv</i>, Cold Spring Harbor Laboratory, doi:<a href=\"https://doi.org/10.1101/2022.08.17.504272\">10.1101/2022.08.17.504272</a>.","apa":"Michalska, J. M., Lyudchik, J., Velicky, P., Korinkova, H., Watson, J., Cenameri, A., … Danzl, J. G. (n.d.). Uncovering brain tissue architecture across scales with super-resolution light microscopy. <i>bioRxiv</i>. Cold Spring Harbor Laboratory. <a href=\"https://doi.org/10.1101/2022.08.17.504272\">https://doi.org/10.1101/2022.08.17.504272</a>","ama":"Michalska JM, Lyudchik J, Velicky P, et al. Uncovering brain tissue architecture across scales with super-resolution light microscopy. <i>bioRxiv</i>. doi:<a href=\"https://doi.org/10.1101/2022.08.17.504272\">10.1101/2022.08.17.504272</a>","short":"J.M. Michalska, J. Lyudchik, P. Velicky, H. Korinkova, J. Watson, A. Cenameri, C.M. Sommer, A. Venturino, K. Roessler, T. Czech, S. Siegert, G. Novarino, P.M. Jonas, J.G. Danzl, BioRxiv (n.d.).","ieee":"J. M. Michalska <i>et al.</i>, “Uncovering brain tissue architecture across scales with super-resolution light microscopy,” <i>bioRxiv</i>. Cold Spring Harbor Laboratory."},"date_published":"2022-08-18T00:00:00Z","user_id":"2DF688A6-F248-11E8-B48F-1D18A9856A87","oa":1,"language":[{"iso":"eng"}],"status":"public","publication":"bioRxiv"},{"external_id":{"pmid":["34079129"],"isi":["000657238100003"]},"isi":1,"year":"2021","acknowledgement":"We thank members of the Greger laboratory, B. Herguedas, J. Krieger and J.-N. Dohrke for comments on the manuscript; J. Krieger and J.-N. Dohrke for discussion, J. Krieger for help with the normal mode analysis, B. Köhegyi for help with cryo-EM imaging, V. Chang and K. Suzuki for helping to generate the CNIH2-1D4-HA stable cell line, M. Carvalho for assistance at early stages of this project, the LMB scientific computing and the cryo-EM facility for support, P. Emsley for help with model building, T. Nakane for helpful comments with RELION 3.1 and R. Warshamanage for helping with EMDA cryo-EM-map processing. We acknowledge the Diamond Light Source for access and support of the Cryo-EM facilities at the UK national electron bio10 imaging centre (eBIC), proposal EM17434, funded by the Wellcome Trust, MRC and BBSRC. This work was supported by grants from the Medical Research Council, as part of United Kingdom Research and Innovation (also known as UK Research and Innovation) (MC_U105174197) and BBSRC (BB/N002113/1) to I.H.G.","date_published":"2021-06-02T00:00:00Z","pmid":1,"status":"public","publication":"Nature","type":"journal_article","_id":"9549","date_updated":"2023-08-08T13:59:51Z","publisher":"Springer Nature","article_processing_charge":"No","doi":"10.1038/s41586-021-03613-0","quality_controlled":"1","main_file_link":[{"url":"https://doi.org/10.1038/s41586-021-03613-0","open_access":"1"}],"page":"454-458","department":[{"_id":"PeJo"}],"month":"06","user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","citation":{"chicago":"Zhang, Danyang, Jake Watson, Peter M. Matthews, Ondrej Cais, and Ingo H. Greger. “Gating and Modulation of a Hetero-Octameric AMPA Glutamate Receptor.” <i>Nature</i>. Springer Nature, 2021. <a href=\"https://doi.org/10.1038/s41586-021-03613-0\">https://doi.org/10.1038/s41586-021-03613-0</a>.","ista":"Zhang D, Watson J, Matthews PM, Cais O, Greger IH. 2021. Gating and modulation of a hetero-octameric AMPA glutamate receptor. Nature. 594, 454–458.","apa":"Zhang, D., Watson, J., Matthews, P. M., Cais, O., &#38; Greger, I. H. (2021). Gating and modulation of a hetero-octameric AMPA glutamate receptor. <i>Nature</i>. Springer Nature. <a href=\"https://doi.org/10.1038/s41586-021-03613-0\">https://doi.org/10.1038/s41586-021-03613-0</a>","mla":"Zhang, Danyang, et al. “Gating and Modulation of a Hetero-Octameric AMPA Glutamate Receptor.” <i>Nature</i>, vol. 594, Springer Nature, 2021, pp. 454–58, doi:<a href=\"https://doi.org/10.1038/s41586-021-03613-0\">10.1038/s41586-021-03613-0</a>.","ama":"Zhang D, Watson J, Matthews PM, Cais O, Greger IH. Gating and modulation of a hetero-octameric AMPA glutamate receptor. <i>Nature</i>. 2021;594:454-458. doi:<a href=\"https://doi.org/10.1038/s41586-021-03613-0\">10.1038/s41586-021-03613-0</a>","ieee":"D. Zhang, J. Watson, P. M. Matthews, O. Cais, and I. H. Greger, “Gating and modulation of a hetero-octameric AMPA glutamate receptor,” <i>Nature</i>, vol. 594. Springer Nature, pp. 454–458, 2021.","short":"D. Zhang, J. Watson, P.M. Matthews, O. Cais, I.H. Greger, Nature 594 (2021) 454–458."},"language":[{"iso":"eng"}],"oa":1,"date_created":"2021-06-13T22:01:33Z","article_type":"original","volume":594,"oa_version":"Published Version","title":"Gating and modulation of a hetero-octameric AMPA glutamate receptor","scopus_import":"1","day":"02","author":[{"first_name":"Danyang","full_name":"Zhang, Danyang","last_name":"Zhang"},{"first_name":"Jake","orcid":"0000-0002-8698-3823","last_name":"Watson","full_name":"Watson, Jake","id":"63836096-4690-11EA-BD4E-32803DDC885E"},{"first_name":"Peter M.","last_name":"Matthews","full_name":"Matthews, Peter M."},{"first_name":"Ondrej","last_name":"Cais","full_name":"Cais, Ondrej"},{"full_name":"Greger, Ingo H.","last_name":"Greger","first_name":"Ingo H."}],"publication_identifier":{"issn":["0028-0836"],"eissn":["1476-4687"]},"publication_status":"published","intvolume":"       594","abstract":[{"text":"AMPA receptors (AMPARs) mediate the majority of excitatory transmission in the brain and enable the synaptic plasticity that underlies learning1. A diverse array of AMPAR signalling complexes are established by receptor auxiliary subunits, which associate with the AMPAR in various combinations to modulate trafficking, gating and synaptic strength2. However, their mechanisms of action are poorly understood. Here we determine cryo-electron microscopy structures of the heteromeric GluA1–GluA2 receptor assembled with both TARP-γ8 and CNIH2, the predominant AMPAR complex in the forebrain, in both resting and active states. Two TARP-γ8 and two CNIH2 subunits insert at distinct sites beneath the ligand-binding domains of the receptor, with site-specific lipids shaping each interaction and affecting the gating regulation of the AMPARs. Activation of the receptor leads to asymmetry between GluA1 and GluA2 along the ion conduction path and an outward expansion of the channel triggers counter-rotations of both auxiliary subunit pairs, promoting the active-state conformation. In addition, both TARP-γ8 and CNIH2 pivot towards the pore exit upon activation, extending their reach for cytoplasmic receptor elements. CNIH2 achieves this through its uniquely extended M2 helix, which has transformed this endoplasmic reticulum-export factor into a powerful AMPAR modulator that is capable of providing hippocampal pyramidal neurons with their integrative synaptic properties. ","lang":"eng"}]},{"date_created":"2021-09-05T22:01:23Z","article_type":"original","volume":12,"oa_version":"Published Version","title":"AMPA receptor anchoring at CA1 synapses is determined by N-terminal domain and TARP γ8 interactions","day":"23","scopus_import":"1","author":[{"orcid":"0000-0002-8698-3823","first_name":"Jake","id":"63836096-4690-11EA-BD4E-32803DDC885E","full_name":"Watson, Jake","last_name":"Watson"},{"first_name":"Alexandra","last_name":"Pinggera","full_name":"Pinggera, Alexandra"},{"first_name":"Hinze","full_name":"Ho, Hinze","last_name":"Ho"},{"first_name":"Ingo H.","last_name":"Greger","full_name":"Greger, Ingo H."}],"file_date_updated":"2021-09-08T12:57:06Z","publication_identifier":{"eissn":["2041-1723"]},"publication_status":"published","tmp":{"name":"Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)","legal_code_url":"https://creativecommons.org/licenses/by/4.0/legalcode","image":"/images/cc_by.png","short":"CC BY (4.0)"},"abstract":[{"lang":"eng","text":"AMPA receptor (AMPAR) abundance and positioning at excitatory synapses regulates the strength of transmission. Changes in AMPAR localisation can enact synaptic plasticity, allowing long-term information storage, and is therefore tightly controlled. Multiple mechanisms regulating AMPAR synaptic anchoring have been described, but with limited coherence or comparison between reports, our understanding of this process is unclear. Here, combining synaptic recordings from mouse hippocampal slices and super-resolution imaging in dissociated cultures, we compare the contributions of three AMPAR interaction domains controlling transmission at hippocampal CA1 synapses. We show that the AMPAR C-termini play only a modulatory role, whereas the extracellular N-terminal domain (NTD) and PDZ interactions of the auxiliary subunit TARP γ8 are both crucial, and each is sufficient to maintain transmission. Our data support a model in which γ8 accumulates AMPARs at the postsynaptic density, where the NTD further tunes their positioning. This interplay between cytosolic (TARP γ8) and synaptic cleft (NTD) interactions provides versatility to regulate synaptic transmission and plasticity."}],"intvolume":"        12","has_accepted_license":"1","article_number":"5083","file":[{"file_id":"9991","date_created":"2021-09-08T12:57:06Z","file_size":18310502,"date_updated":"2021-09-08T12:57:06Z","creator":"cchlebak","relation":"main_file","checksum":"1bf4f6a561f96bc426d754de9cb57710","success":1,"file_name":"2021_NatureCommunications_Watson.pdf","access_level":"open_access","content_type":"application/pdf"}],"department":[{"_id":"PeJo"}],"month":"08","user_id":"4359f0d1-fa6c-11eb-b949-802e58b17ae8","citation":{"apa":"Watson, J., Pinggera, A., Ho, H., &#38; Greger, I. H. (2021). AMPA receptor anchoring at CA1 synapses is determined by N-terminal domain and TARP γ8 interactions. <i>Nature Communications</i>. Nature Publishing Group. <a href=\"https://doi.org/10.1038/s41467-021-25281-4\">https://doi.org/10.1038/s41467-021-25281-4</a>","mla":"Watson, Jake, et al. “AMPA Receptor Anchoring at CA1 Synapses Is Determined by N-Terminal Domain and TARP Γ8 Interactions.” <i>Nature Communications</i>, vol. 12, no. 1, 5083, Nature Publishing Group, 2021, doi:<a href=\"https://doi.org/10.1038/s41467-021-25281-4\">10.1038/s41467-021-25281-4</a>.","chicago":"Watson, Jake, Alexandra Pinggera, Hinze Ho, and Ingo H. Greger. “AMPA Receptor Anchoring at CA1 Synapses Is Determined by N-Terminal Domain and TARP Γ8 Interactions.” <i>Nature Communications</i>. Nature Publishing Group, 2021. <a href=\"https://doi.org/10.1038/s41467-021-25281-4\">https://doi.org/10.1038/s41467-021-25281-4</a>.","ista":"Watson J, Pinggera A, Ho H, Greger IH. 2021. AMPA receptor anchoring at CA1 synapses is determined by N-terminal domain and TARP γ8 interactions. Nature Communications. 12(1), 5083.","ieee":"J. Watson, A. Pinggera, H. Ho, and I. H. Greger, “AMPA receptor anchoring at CA1 synapses is determined by N-terminal domain and TARP γ8 interactions,” <i>Nature Communications</i>, vol. 12, no. 1. Nature Publishing Group, 2021.","short":"J. Watson, A. Pinggera, H. Ho, I.H. Greger, Nature Communications 12 (2021).","ama":"Watson J, Pinggera A, Ho H, Greger IH. AMPA receptor anchoring at CA1 synapses is determined by N-terminal domain and TARP γ8 interactions. <i>Nature Communications</i>. 2021;12(1). doi:<a href=\"https://doi.org/10.1038/s41467-021-25281-4\">10.1038/s41467-021-25281-4</a>"},"issue":"1","language":[{"iso":"eng"}],"oa":1,"type":"journal_article","_id":"9985","date_updated":"2023-08-11T11:07:51Z","publisher":"Nature Publishing Group","article_processing_charge":"Yes","doi":"10.1038/s41467-021-25281-4","quality_controlled":"1","ddc":["612"],"external_id":{"isi":["000687672000006"],"pmid":["34426577 "]},"isi":1,"year":"2021","acknowledgement":"The authors are very grateful to Andrew Penn for advice and discussions on surface receptor labelling in slice tissue, dissociated culture transfection, and for providing tdTomato and BirAER expression plasmids. This work would not have been possible without support from the Biological Services teams at both the Laboratory of Molecular Biology and Ares facilities. We are also very grateful to Nick Barry and Jerome Boulanger of the LMB Light Microscopy facility for support with confocal and STORM imaging and analysis, Junichi Takagi for providing scFv-Clasp expression constructs, Veronica Chang for assistance with scFv-Clasp protein production, and Nejc Kejzar for assistance with cluster analysis. We would like to thank Teru Nakagawa and Ole Paulsen for critical reading of the manuscript and constructive feedback. This work was supported by grants from the Medical Research Council (MC_U105174197) and BBSRC (BB/N002113/1).","date_published":"2021-08-23T00:00:00Z","pmid":1,"status":"public","publication":"Nature Communications"}]