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Author: U. Valentin Nägerl
Publisher: Humana
ISBN: 9781493949182
Category : Medical
Languages : en
Pages : 0
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Book Description
Synapses underlie rapid and flexible neural communication in the brain and they hold the key to understanding higher brain functions in health and disease. Because they are very small and highly dynamic, it is very difficult to study them with traditional techniques. Fortunately, recent ground-breaking advances in optical microscopy (e.g. STED, PALM, STORM, SIM) have greatly improved our ability to image living synapses at the nanoscale, even down to the level of single molecules. The proposed volume brings together leading researchers to review these exciting new techniques and their application in neurobiological research. It will explain and discuss the basic principles behind the various superresolution modalities, how they are implemented, what their scope and limitations are etc. In addition, several key research discoveries on synapses enabled by these novel approaches will be highlighted.
Author: U. Valentin Nägerl
Publisher: Humana
ISBN: 9781493949182
Category : Medical
Languages : en
Pages : 0
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Book Description
Synapses underlie rapid and flexible neural communication in the brain and they hold the key to understanding higher brain functions in health and disease. Because they are very small and highly dynamic, it is very difficult to study them with traditional techniques. Fortunately, recent ground-breaking advances in optical microscopy (e.g. STED, PALM, STORM, SIM) have greatly improved our ability to image living synapses at the nanoscale, even down to the level of single molecules. The proposed volume brings together leading researchers to review these exciting new techniques and their application in neurobiological research. It will explain and discuss the basic principles behind the various superresolution modalities, how they are implemented, what their scope and limitations are etc. In addition, several key research discoveries on synapses enabled by these novel approaches will be highlighted.
Author: Mirelle Jamilla Tamara Ter Veer
Publisher:
ISBN:
Category :
Languages : en
Pages : 0
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Book Description
The brain is a complex organ consisting of neurons and non-neuronal cells. Communication between neurons takes place via synapses, whose morphological remodeling is thought to be crucial for information processing and storage in the mammalian brain. Recently, this neuro-centric view of synaptic function has evolved, also taking into account the glial processes in close vicinity of the synapse. However, as their structure is well below the spatial resolution of conventional light microscopy, progress in investigating them in a physiological environment, the intact brain, has been impeded. Indeed, little is known on the nanoscale morphological variations of dendritic spines, the interaction with glial processes, and how these affect synaptic transmission in vivo. Here, we aim to visualize the dynamic nano-morphology of dendritic spines in mouse somatosensory cortex in vivo. We implemented super-resolution 2P-STED time-lapse imaging, which allows for high spatial resolution and deep tissue penetration, in anesthetized mice, and show that the nano-morphology of spines is diverse, variable, but on average stable, and that differences in spine morphology can have an effect on spine biochemical compartmentalization in vivo. Moreover, implementation of dual color in vivo super-resolution imaging and a novel astrocytic labeling approach provided the first steps towards nanoscale characterization of neuron-glia interactions in vivo. These findings bring new insights in synapse dynamics at the nanoscale in vivo, and our methodological endeavors help pave the way for a better understanding of how nanoscale aspects of spine morphology and their dynamics might contribute to brain physiology and animal behavior.
Author: Lara Laparra Cuervo
Publisher:
ISBN:
Category :
Languages : en
Pages : 172
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Book Description
The arrival of super-resolution techniques has driven researchers to explore biological areas that were unreachable before. Such techniques not only allowed the improvement of spatial resolution in images but also the possibility to perform quantitative measurements at the single-molecule level. The interest in that particular field has been growing over the years and new and more sophisticated tools have been developed. Neuroscience has been one of the first fields to adapt and benefit from super-resolution microscopy. These techniques opened a new window of opportunity to reveal spatial organization of the neuronal cytoskeleton and the molecular organization and dynamics of the synapse, structures below the spatial resolution limit of conventional light microscopy. Protein organization and stoichiometry is central for synaptic transmission in neurons. Knowing the absolute numbers of proteins playing a key role in diseases can be of extreme interest in order to understand the mechanisms that lead to such neurological disorders. In this thesis we exploited the super-resolution techniques to develop a pioneering method for quantitative single-molecule measurements and to unravel the organization of a synaptic protein complex that was never visualized before with nanoscale spatial resolution. We developed a novel method in order to quantify the photoactivation efficiency of eight different photoswitchable fluorescent proteins commonly used in super-resolution experiments. We used the glycine receptor as a template because of its well-known stoichiometry and tagged eight different photoswitchable fluorescent proteins to the a- and ß-subunits of this receptor and transiently transfected them to Xenopus oocytes. The fact that the fluorescent proteins are genetically encoded make them highly suitable for quantitative single-molecule counting. The photoactivation efficiency, which is the percentage of a fluorescent protein that photoactivates into a fluorescently detectable form, plays a critical role in properly interpreting quantitative measurements. Moreover, we also focused our studies on super-resolution imaging of a synaptic protein complex, called LGI1 complex. This ensemble of proteins is one of the main key players involved in different neurological disorders. Leucine rich glioma activated 1 (LGI1) is a neuronal protein that forms a trans-synaptic bridge linking pre- and postsynaptic transmembrane proteins (ADAM22 and ADAM23) and helps to organize a multimeric complex at the synapse including AMPA receptors and voltage-gated potassium channels (VGKC). LGI1 autoimmune encephalitis is a severe neuropsychiatric disorder related to epilepsy where the patients produce autoantibodies against LGI1, which alter synaptic plasticity. However, the molecular mechanisms that lead to the observed problems in patients still remain largely unknown. Using well-characterized synaptic markers as molecular standards, we determined the positioning of LGI1 and the other related proteins within the synaptic space at nanoscale resolution by means of multi-color STORM. Further, the comparison of this molecular architecture in healthy neurons versus neurons treated with antibodies from patients suffering from LGI1 autoimmune encephalitis showed that these antibodies impact the nanoscale organization of pre-synaptic proteins. These results suggested a loss of LGI1 interaction with pre-synaptic proteins upon antibody binding and gave further insight into early changes in pathology.
Author: Xiaobing Chen
Publisher: Frontiers Media SA
ISBN: 2832500846
Category : Science
Languages : en
Pages : 195
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Book Description
Author: Philip Tinnefeld
Publisher: Springer
ISBN: 3662455471
Category : Science
Languages : en
Pages : 340
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Book Description
This book describes developments in the field of super-resolution fluorescence microscopy or nanoscopy. In 11 chapters, distinguished scientists and leaders in their respective fields describe different nanoscopy approaches, various labeling technologies, and concrete applications. The topics covered include the principles and applications of the most popular nanoscopy techniques STED and (f)PALM/STORM, along with advances brought about by fluorescent proteins and organic dyes optimized for fluorescence nanoscopy. Furthermore, the photophysics of fluorescent labels is addressed, specifically for improving their photoswitching capabilities. Important applications are also discussed, such as the tracking and counting of molecules to determine acting forces in cells, and quantitative cellular imaging, respectively, as well as the mapping of chemical reaction centers at the nano-scale. The 2014 Chemistry Nobel Prize® was awarded for the ground-breaking developments of super-resolved fluorescence microscopy. In this book, which was co-edited by one of the prize winners, readers will find the most recent developments in this field.
Author: Chih-Chieh Jay Yu
Publisher:
ISBN:
Category :
Languages : en
Pages : 146
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Book Description
Expansion microscopy (ExM) enables 3-D, nanoscale-precise imaging of biological specimens by isotropic swelling of hydrogel-embedded, chemically processed tissue. Such capability raises the question of whether nanoscale mapping of biomolecules could be performed in an entire organism, which would allow super-resolution-mediated in situ analyses, such as digital quantification of biomolecules and mapping of synaptic contacts, to be performed within the context of an entire nervous system. The nematode Caenorhabditis elegans could be a suitable model for such organism-wide analyses, due to its tractable physical size, deterministic cell lineage, ease of genetic control, and well-established literature. However, C. elegans is enclosed in a chemically impermeable and mechanically tough cuticle, which could hinder the deployment of ExM. In this thesis, we present a strategy, expansion of C. elegans (ExCel), to expand fixed, cuticle-enclosed intact animals of C. elegans. ExCel enables simultaneous readout of fluorescent proteins, RNAs, DNA locations, and anatomical structures at resolutions of ~65-75 nm (3.3-3.8x linear expansion). We also developed epitope-preserving ExCel, which enables imaging of endogenous proteins stained by antibodies, and iterative ExCel, which enables imaging of fluorescent proteins at a ~25-nm resolution (20x linear expansion). We demonstrate the utility of the ExCel toolbox for multiplexed imaging of multiple molecular types, for mapping synaptic proteins, for identifying previously unreported proteins at cell junctions, and for gene expression analysis in multiple individual neurons of the same animal. In addition to ExCel, we discuss two other ExM-related technologies, including tetragel, which is a highly homogeneous hydrogel network that improves the nanoscale isotropy of biological ultrastructure expanded by ExM, and stochastic arrangement of reporters in clusters (STARC), which is a strategy for recording neuronal activity at a subneurite-level resolution, in densely labeled neuronal populations. Taken together, the work presented in this thesis extends the capabilities of ExM, and lays the foundation for a comprehensive, functionally and structurally informed analysis of an entire organism, which could reveal new insights in neuroscience, organismal development, and systems biology.
Author: Jennifer-Magdalena Masch
Publisher:
ISBN:
Category :
Languages : en
Pages :
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Book Description
Optical nanoscopy has revolutionized far-field microscopy, enabling the observation of subcellular structures and dynamics from completely new perspectives. Among other fields, neuroscience benefits greatly from the remarkable advances in super-resolution microscopy, which provide unprecedented insights into the molecular organization and function of synapses. STED nanoscopy has been one of the most successful methods for live-cell applications and is the only superresolution technique that has been demonstrated for imaging in living mice so far. However, in vivo nanoscale imaging of synapt...
Author: Pasko Rakic
Publisher: Academic Press
ISBN: 0128236736
Category : Psychology
Languages : en
Pages : 560
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Book Description
Synapse Development and Maturation, the latest release in the Comprehensive Developmental Neuroscience series, presents the latest information on the genetic, molecular and cellular mechanisms of neural development. The book provides a much-needed update that underscores the latest research in this rapidly evolving field, with new section editors discussing the technological advances that are enabling the pursuit of new research on brain development. This volume focuses on the synaptogenesis and developmental sequences in the maturation of intrinsic and synapse-driven patterns. Features leading experts in various subfields as section editors and article authors Presents articles that have been peer reviewed to ensure accuracy, thoroughness and scholarship Includes coverage of mechanisms which regulate synapse formation and maintenance during development Covers neural activity, from cell-intrinsic maturation, to early correlated patterns of activity
Author: Nobuhiko Yamamoto
Publisher: Humana
ISBN: 9781071605349
Category : Medical
Languages : en
Pages : 331
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Book Description
This volume looks at the methodology and techniques used by experts to study how certain molecules function in specific locations, and their temporal patterns. Chapters in this book cover topics such as in vivo single-molecule tracking of voltage-gated calcium channels with split-fluorescent proteins in CRISPR-engineering C. elegans; protein-protein interactions in membranes using single particle tracking; neuropathological diseases revealed by quantum-dot single particle tracking; SPoD-OnSPAN; and investigating molecular diffusion inside small neuronal compartments with two-photon fluorescence correlation spectroscopy. In the Neuromethods series style, chapters include the kind of detail and key advice from the specialists needed to get successful results in your laboratory. Cutting-edge and comprehensive, Single Molecule Microscopy is a valuable resource for any researcher interested in learning more about this important field.
Author: Tim Salditt
Publisher: Springer Nature
ISBN: 3030344134
Category : Science
Languages : en
Pages : 634
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Book Description
This open access book, edited and authored by a team of world-leading researchers, provides a broad overview of advanced photonic methods for nanoscale visualization, as well as describing a range of fascinating in-depth studies. Introductory chapters cover the most relevant physics and basic methods that young researchers need to master in order to work effectively in the field of nanoscale photonic imaging, from physical first principles, to instrumentation, to mathematical foundations of imaging and data analysis. Subsequent chapters demonstrate how these cutting edge methods are applied to a variety of systems, including complex fluids and biomolecular systems, for visualizing their structure and dynamics, in space and on timescales extending over many orders of magnitude down to the femtosecond range. Progress in nanoscale photonic imaging in Göttingen has been the sum total of more than a decade of work by a wide range of scientists and mathematicians across disciplines, working together in a vibrant collaboration of a kind rarely matched. This volume presents the highlights of their research achievements and serves as a record of the unique and remarkable constellation of contributors, as well as looking ahead at the future prospects in this field. It will serve not only as a useful reference for experienced researchers but also as a valuable point of entry for newcomers.
