title: Nanoscale Molecular Reorganization of the Inhibitory Postsynaptic Density Is a Determinant of GABAergic Synaptic Potentiation.
abstract: Gephyrin is a key scaffold protein mediating the anchoring of GABAA receptors at inhibitory synapses. Here, we exploited superresolution techniques combined with proximity-based clustering analysis and model simulations to investigate the single-molecule gephyrin reorganization during plasticity of inhibitory synapses in mouse hippocampal cultured neurons. This approach revealed that, during the expression of inhibitory LTP, the increase of gephyrin density at postsynaptic sites is associated with the promoted formation of gephyrin nanodomains. We demonstrate that the gephyrin rearrangement in nanodomains stabilizes the amplitude of postsynaptic currents, indicating that, in addition to the number of synaptic GABAA receptors, the nanoscale distribution of GABAA receptors in the postsynaptic area is a crucial determinant for the expression of inhibitory synaptic plasticity. In addition, the methodology implemented here clears the way to the application of the graph-based theory to single-molecule data for the description and quantification of the spatial organization of the synapse at the single-molecule level.SIGNIFICANCE STATEMENTThe mechanisms of inhibitory synaptic plasticity are poorly understood, mainly because the size of the synapse is below the diffraction limit, thus reducing the effectiveness of conventional optical and imaging techniques. Here, we exploited superresolution approaches combined with clustering analysis to study at unprecedented resolution the distribution of the inhibitory scaffold protein gephyrin in response to protocols inducing LTP of inhibitory synaptic responses (iLTP). We found that, during the expression of iLTP, the increase of synaptic gephyrin is associated with the fragmentation of gephyrin in subsynaptic nanodomains. We demonstrate that such synaptic gephyrin nanodomains stabilize the amplitude of inhibitory postsynaptic responses, thus identifying the nanoscale gephyrin rearrangement as a key determinant for inhibitory synaptic plasticity.
authors.affiliation: Nanoscopy Department.|Nikon Imaging Center, Fondazione Istituto Italiano di Tecnologia, 16163 Genoa, Italy.|Università di Genova, Department of Physics, 16146 Genoa, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Pennacchietti
authors.affiliation: Pattern Analysis and Computer Vision Department.
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Vascon
authors.affiliation: Neuroscience and Brain Technologies Department, and.
authors.forename: Thierry
authors.identifier: 0000-0001-8708-1015
authors.initials: T
authors.lastname: Nieus
authors.affiliation: Neuroscience and Brain Technologies Department, and.
authors.forename: Christian
authors.identifier: 0000-0002-2841-0445
authors.initials: C
authors.lastname: Rosillo
authors.affiliation: Neuroscience and Brain Technologies Department, and.
authors.forename: Sabyasachi
authors.initials: S
authors.lastname: Das
authors.affiliation: Institute of Pharmacology and Toxicology, University of Zurich, 8057 Zurich, Switzerland, and.
authors.forename: Shiva K
authors.identifier: 0000-0003-0074-1805
authors.initials: SK
authors.lastname: Tyagarajan
authors.affiliation: Nanoscopy Department.|Nikon Imaging Center, Fondazione Istituto Italiano di Tecnologia, 16163 Genoa, Italy.|Università di Genova, Department of Physics, 16146 Genoa, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Diaspro
authors.affiliation: Pattern Analysis and Computer Vision Department.
authors.forename: Alessio
authors.identifier: 0000-0002-2262-4872
authors.initials: A
authors.lastname: Del Bue
authors.affiliation: Neuroscience and Brain Technologies Department, and.
authors.forename: Enrica Maria
authors.identifier: 0000-0001-5168-4148
authors.initials: EM
authors.lastname: Petrini
authors.affiliation: Neuroscience and Brain Technologies Department, and andrea.barberis@iit.it francesca.cella@iit.it.
authors.forename: Andrea
authors.identifier: 0000-0002-5054-6517
authors.initials: A
authors.lastname: Barberis
authors.affiliation: Nanoscopy Department, andrea.barberis@iit.it francesca.cella@iit.it.|Nikon Imaging Center, Fondazione Istituto Italiano di Tecnologia, 16163 Genoa, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Cella Zanacchi
mesh_terms: D018750:6-Cyano-7-nitroquinoxaline-2,3-dione / Q000494:pharmacology; D000465:Algorithms; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D003198:Computer Simulation; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005260:Female; D059330:GABAergic Neurons / Q000166:cytology* / Q000187:drug effects; D006624:Hippocampus / Q000166:cytology; D036881:Long-Term Synaptic Depression / Q000187:drug effects / Q000502:physiology*; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008959:Models, Neurological; D016202:N-Methylaspartate / Q000494:pharmacology; D010455:Peptides / Q000378:metabolism; D011108:Polymers; D057907:Post-Synaptic Density / Q000187:drug effects / Q000378:metabolism*; D011963:Receptors, GABA-A / Q000378:metabolism; D014633:Valine / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D002352:Carrier Proteins; D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D008565:Membrane Proteins; D010455:Peptides; D011108:Polymers; D011963:Receptors, GABA-A; C073441:gephyrin; C020749:GAT; D016202:N-Methylaspartate; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; C056639:2-amino-5-phosphopentanoic acid; D014633:Valine
keywords: GABAA receptors; PALM; STORM; clustering; gephyrin; model simulations; plasticity