当前位置： SCI文献检索 > Nature Protocols期刊下所有文献 > Analysis of neurotransmitter release mechanisms by photolysis of caged Ca²⁺ in an autaptic neuron culture system.

Analysis of neurotransmitter release mechanisms by photolysis of caged Ca²⁺ in an autaptic neuron culture system.

Abstract：

:Neurotransmitter release is triggered by membrane depolarization, Ca(2+) influx and Ca(2+) sensing by the release machinery, causing synaptic vesicle (SV) fusion with the plasma membrane. Interlinked is a complex membrane cycle in which vesicles are tethered to the release site, primed, fused and recycled. As many of these processes are Ca(2+) dependent and simultaneously occurring, it is difficult to dissect them experimentally. This problem can be partially circumvented by controlling synaptic Ca(2+) concentrations via UV photolysis of caged Ca(2+). We developed a culture protocol for Ca(2+) uncaging in small synapses on the basis of the generation of small glia cell islands with single neurons on top, which are sufficiently small to be covered with a UV-light flash. Neurons are loaded with the photolabile Ca(2+)-chelator nitrophenyl-EGTA and Ca(2+) indicators, and a UV flash is used to trigger Ca(2+)-uncaging and SV fusion. The protocol takes three weeks to complete and provides unprecedented insights into the mechanisms of transmitter release.

journal_name

Nat Protoc

journal_title

Nature protocols

authors

Burgalossi A,Jung S,Man KN,Nair R,Jockusch WJ,Wojcik SM,Brose N,Rhee JS

doi

10.1038/nprot.2012.074

subject

Has Abstract

pub_date

2012-06-21 00:00:00

pages

1351-65

issue

7

eissn

1754-2189

issn

1750-2799

pii

nprot.2012.074

journal_volume

7

pub_type

杂志文章

相关文献

Nature Protocols文献大全
  • A mass spectrometry workflow for measuring protein turnover rates in vivo.

    abstract：:Proteins are continually produced and degraded, to avoid the accumulation of old or damaged molecules and to maintain the efficiency of physiological processes. Despite its importance, protein turnover has been difficult to measure in vivo. Previous approaches to evaluating turnover in vivo have required custom labeli...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-019-0222-y

    authors： Alevra M,Mandad S,Ischebeck T,Urlaub H,Rizzoli SO,Fornasiero EF

    更新日期：2019-12-01 00:00:00

  • Single-molecule mRNA detection and counting in mammalian tissue.

    abstract：:We present a protocol for visualizing and quantifying single mRNA molecules in mammalian (mouse and human) tissues. In the approach described here, sets of about 50 short oligonucleotides, each labeled with a single fluorophore, are hybridized to target mRNAs in tissue sections. Each set binds to a single mRNA molecul...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2013.109

    authors： Lyubimova A,Itzkovitz S,Junker JP,Fan ZP,Wu X,van Oudenaarden A

    更新日期：2013-09-01 00:00:00

  • A whole-tissue RNA-seq toolkit for organism-wide studies of gene expression with PME-seq.

    abstract：:The immune system operates at the scale of the whole organism in mammals. We currently lack experimental approaches to systematically track and study organism-wide molecular processes in mice. Here we describe an integrated toolkit for measuring gene expression in whole tissues, 3-prime mRNA extension sequencing, that...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-019-0291-y

    authors： Pandey S,Takahama M,Gruenbaum A,Zewde M,Cheronis K,Chevrier N

    更新日期：2020-04-01 00:00:00

  • Mapping in vivo target interaction profiles of covalent inhibitors using chemical proteomics with label-free quantification.

    abstract：:Activity-based protein profiling (ABPP) has emerged as a valuable chemical proteomics method to guide the therapeutic development of covalent drugs by assessing their on-target engagement and off-target activity. We recently used ABPP to determine the serine hydrolase interaction landscape of the experimental drug BIA...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2017.159

    authors： van Rooden EJ,Florea BI,Deng H,Baggelaar MP,van Esbroeck ACM,Zhou J,Overkleeft HS,van der Stelt M

    更新日期：2018-04-01 00:00:00

  • A protocol for isolation and culture of mesenchymal stem cells from mouse compact bone.

    abstract：:Unlike humans, mouse bone marrow-derived mesenchymal stem cells (MSCs) cannot be easily harvested by adherence to plastic owing to the contamination of cultures by hematopoietic cells. The design of the protocol described here is based on the phenomenon that compact bones abound in MSCs and hematopoietic cells exist i...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2009.238

    authors： Zhu H,Guo ZK,Jiang XX,Li H,Wang XY,Yao HY,Zhang Y,Mao N

    更新日期：2010-03-01 00:00:00

  • τ-SGA: synthetic genetic array analysis for systematically screening and quantifying trigenic interactions in yeast.

    abstract：:Systematic complex genetic interaction studies have provided insight into high-order functional redundancies and genetic network wiring of the cell. Here, we describe a method for screening and quantifying trigenic interactions from ordered arrays of yeast strains grown on agar plates as individual colonies. The proto...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-020-00456-3

    authors： Kuzmin E,Rahman M,VanderSluis B,Costanzo M,Myers CL,Andrews BJ,Boone C

    更新日期：2021-01-18 00:00:00

  • Fabrication and practical applications of molybdenum disulfide nanopores.

    abstract：:Among the different developed solid-state nanopores, nanopores constructed in a monolayer of molybdenum disulfide (MoS2) stand out as powerful devices for single-molecule analysis or osmotic power generation. Because the ionic current through a nanopore is inversely proportional to the thickness of the pore, ultrathin...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-019-0131-0

    authors： Graf M,Lihter M,Thakur M,Georgiou V,Topolancik J,Ilic BR,Liu K,Feng J,Astier Y,Radenovic A

    更新日期：2019-04-01 00:00:00

  • Microwave-assisted preparation of the quorum-sensing molecule 2-heptyl-3-hydroxy-4(1H)-quinolone and structurally related analogs.

    abstract：:An optimized procedure for the efficient preparation of 2-heptyl-3-hydroxy-4(1H)-quinolone (Pseudomonas quinolone signal or PQS) and a diverse range of structurally related 2-alkyl-4-quinolones with biological activity is presented. The two-step synthesis begins with the formation of α-chloro ketones by the coupling o...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2012.054

    authors： Hodgkinson JT,Galloway WR,Welch M,Spring DR

    更新日期：2012-05-24 00:00:00

  • A protocol for isolation and culture of human umbilical vein endothelial cells.

    abstract：:We describe a protocol for easy isolation and culture of human umbilical vein endothelial cells (HUVECs) to supply every researcher with a method that can be applied in cell biology laboratories with minimum equipment. Endothelial cells (ECs) are isolated from umbilical vein vascular wall by a collagenase treatment, t...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2007.54

    authors： Baudin B,Bruneel A,Bosselut N,Vaubourdolle M

    更新日期：2007-01-01 00:00:00

  • Six alternative proteases for mass spectrometry-based proteomics beyond trypsin.

    abstract：:Protein digestion using a dedicated protease represents a key element in a typical mass spectrometry (MS)-based shotgun proteomics experiment. Up to now, digestion has been predominantly performed with trypsin, mainly because of its high specificity, widespread availability and ease of use. Lately, it has become appar...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2016.057

    authors： Giansanti P,Tsiatsiani L,Low TY,Heck AJ

    更新日期：2016-05-01 00:00:00

  • Identifying and quantifying proteolytic events and the natural N terminome by terminal amine isotopic labeling of substrates.

    abstract：:Analysis of the sequence and nature of protein N termini has many applications. Defining the termini of proteins for proteome annotation in the Human Proteome Project is of increasing importance. Terminomics analysis of protease cleavage sites in degradomics for substrate discovery is a key new application. Here we de...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2011.382

    authors： Kleifeld O,Doucet A,Prudova A,auf dem Keller U,Gioia M,Kizhakkedathu JN,Overall CM

    更新日期：2011-09-22 00:00:00

  • Direct metabolomics for plant cells by live single-cell mass spectrometry.

    abstract：:Live single-cell mass spectrometry (live MS) provides a mass spectrum that shows thousands of metabolite peaks from a single live plant cell within minutes. By using an optical microscope, a cell is chosen for analysis and a metal-coated nanospray microcapillary tip is used to remove the cell's contents. After adding ...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2015.084

    authors： Fujii T,Matsuda S,Tejedor ML,Esaki T,Sakane I,Mizuno H,Tsuyama N,Masujima T

    更新日期：2015-09-01 00:00:00

  • Vibrational spectroscopic image analysis of biological material using multivariate curve resolution-alternating least squares (MCR-ALS).

    abstract：:Raman and Fourier transform IR (FTIR) microspectroscopic images of biological material (tissue sections) contain detailed information about their chemical composition. The challenge lies in identifying changes in chemical composition, as well as locating and assigning these changes to different conditions (pathology, ...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2015.008

    authors： Felten J,Hall H,Jaumot J,Tauler R,de Juan A,Gorzsás A

    更新日期：2015-02-01 00:00:00

  • High-throughput ballistic injection nanorheology to measure cell mechanics.

    abstract：:High-throughput ballistic injection nanorheology is a method for the quantitative study of cell mechanics. Cell mechanics are measured by ballistic injection of submicron particles into the cytoplasm of living cells and tracking the spontaneous displacement of the particles at high spatial resolution. The trajectories...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2011.436

    authors： Wu PH,Hale CM,Chen WC,Lee JS,Tseng Y,Wirtz D

    更新日期：2012-01-05 00:00:00

  • Cryopreservation and banking of mammalian cell lines.

    abstract：:This protocol describes the principles and methods used for the preparation of cryopreserved cell stocks. Following these procedures will ensure the availability of reproducible cultures for use within a single laboratory at different times and for different collaborating laboratories. Although the basic principle is ...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2008.190

    authors： Stacey GN,Masters JR

    更新日期：2008-01-01 00:00:00

  • Accessing crystal-crystal interaction forces with oriented nanocrystal atomic force microscopy probes.

    abstract：:Biominerals serve as critical structures of living systems and play important roles in biochemical processes. Understanding their crystallization mechanisms is therefore central to many areas of biology, biogeoscience, and biochemistry. Some biominerals, such as bone and dentin, are hierarchical nanocomposite structur...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-018-0027-4

    authors： Zhang X,He Y,Liu J,Bowden ME,Kovarik L,Mao SX,Wang C,De Yoreo JJ,Rosso KM

    更新日期：2018-09-01 00:00:00

  • Development of a highly reproducible three-dimensional organotypic model of the oral mucosa.

    abstract：:In this report we describe the development of a standardized three-dimensional (3D) system of the human oral mucosa based on an immortalized human oral keratinocyte cell line (OKF6/TERT-2). The procedure takes approximately 2-3 weeks to complete and includes three main stages: preparation of collagen-embedded fibrobla...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2006.323

    authors： Dongari-Bagtzoglou A,Kashleva H

    更新日期：2006-01-01 00:00:00

  • 3D computational reconstruction of tissues with hollow spherical morphologies using single-cell gene expression data.

    abstract：:Single-cell gene expression analysis has contributed to a better understanding of the transcriptional heterogeneity in a variety of model systems, including those used in research in developmental, cancer and stem cell biology. Nowadays, technological advances facilitate the generation of large gene expression data se...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2015.022

    authors： Durruthy-Durruthy R,Gottlieb A,Heller S

    更新日期：2015-03-01 00:00:00

  • Synthesis of complex head-to-side-chain cyclodepsipeptides.

    abstract：:Cyclodepsipeptides are cyclic peptides in which at least one amide link on the backbone is replaced with an ester link. These natural products present a high structural diversity that corresponds to a broad range of biological activities. Therefore, they are very promising pharmaceutical candidates. Most of the cyclod...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2016.116

    authors： Pelay-Gimeno M,Albericio F,Tulla-Puche J

    更新日期：2016-10-01 00:00:00

  • Real-time in vivo imaging of transgenic bioluminescent blood stages of rodent malaria parasites in mice.

    abstract：:This protocol describes a methodology for imaging the sequestration of infected erythrocytes of the rodent malaria parasite Plasmodium berghei in the bodies of live mice or in dissected organs, using a transgenic parasite that expresses luciferase. Real-time imaging of infected erythrocytes is performed by measuring b...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2006.69

    authors： Franke-Fayard B,Waters AP,Janse CJ

    更新日期：2006-01-01 00:00:00

  • Isolation of plasma membrane-associated membranes from rat liver.

    abstract：:Dynamic interplay between intracellular organelles requires a particular functional apposition of membrane structures. The organelles involved come into close contact, but do not fuse, thereby giving rise to notable microdomains; these microdomains allow rapid communication between the organelles. Plasma membrane-asso...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2014.016

    authors： Suski JM,Lebiedzinska M,Wojtala A,Duszynski J,Giorgi C,Pinton P,Wieckowski MR

    更新日期：2014-02-01 00:00:00

  • Genome-wide measurement of protein-DNA binding dynamics using competition ChIP.

    abstract：:Competition chromatin immunoprecipitation (competition ChIP) enables experimenters to measure protein-DNA dynamics at a single locus or across the entire genome, depending on the detection method. Competition ChIP relies on a cell containing two copies of a single DNA-associated factor, with each copy of the factor di...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2013.077

    authors： Lickwar CR,Mueller F,Lieb JD

    更新日期：2013-01-01 00:00:00

  • Count-based differential expression analysis of RNA sequencing data using R and Bioconductor.

    abstract：:RNA sequencing (RNA-seq) has been rapidly adopted for the profiling of transcriptomes in many areas of biology, including studies into gene regulation, development and disease. Of particular interest is the discovery of differentially expressed genes across different conditions (e.g., tissues, perturbations) while opt...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2013.099

    authors： Anders S,McCarthy DJ,Chen Y,Okoniewski M,Smyth GK,Huber W,Robinson MD

    更新日期：2013-09-01 00:00:00

  • Standardization of complex biologically derived spectrochemical datasets.

    abstract：:Spectroscopic techniques such as Fourier-transform infrared (FTIR) spectroscopy are used to study interactions of light with biological materials. This interaction forms the basis of many analytical assays used in disease screening/diagnosis, microbiological studies, and forensic/environmental investigations. Advantag...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-019-0150-x

    authors： Morais CLM,Paraskevaidi M,Cui L,Fullwood NJ,Isabelle M,Lima KMG,Martin-Hirsch PL,Sreedhar H,Trevisan J,Walsh MJ,Zhang D,Zhu YG,Martin FL

    更新日期：2019-05-01 00:00:00

  • Multiplexed array-based and in-solution genomic enrichment for flexible and cost-effective targeted next-generation sequencing.

    abstract：:The unprecedented increase in the throughput of DNA sequencing driven by next-generation technologies now allows efficient analysis of the complete protein-coding regions of genomes (exomes) for multiple samples in a single sequencing run. However, sample preparation and targeted enrichment of multiple samples has bec...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2011.396

    authors： Harakalova M,Mokry M,Hrdlickova B,Renkens I,Duran K,van Roekel H,Lansu N,van Roosmalen M,de Bruijn E,Nijman IJ,Kloosterman WP,Cuppen E

    更新日期：2011-11-03 00:00:00

  • Transformation of the rodent malaria parasite Plasmodium chabaudi.

    abstract：:The rodent malaria parasite Plasmodium chabaudi chabaudi shares many features with human malaria species, including P. falciparum, and is the in vivo model of choice for many aspects of malaria research in the mammalian host, from sequestration of parasitized erythrocytes, to antigenic variation and host immunity and ...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2011.313

    authors： Spence PJ,Cunningham D,Jarra W,Lawton J,Langhorne J,Thompson J

    更新日期：2011-04-01 00:00:00

  • Generating high-purity cardiac and endothelial derivatives from patterned mesoderm using human pluripotent stem cells.

    abstract：:Human pluripotent stem cells (hPSCs) provide a valuable model for the study of human development and a means to generate a scalable source of cells for therapeutic applications. This protocol specifies cell fate efficiently into cardiac and endothelial lineages from hPSCs. The protocol takes 2 weeks to complete and re...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2016.153

    authors： Palpant NJ,Pabon L,Friedman CE,Roberts M,Hadland B,Zaunbrecher RJ,Bernstein I,Zheng Y,Murry CE

    更新日期：2017-01-01 00:00:00

  • Rapid bacterial artificial chromosome modification for large-scale mouse transgenesis.

    abstract：:We report here a high-throughput method for the modification of bacterial artificial chromosomes (BACs) that uses a novel two-plasmid approach. In this protocol, a vector modified in our laboratory to hold an R6Kγ origin of replication and a marker recombination cassette is inserted into a BAC in a single recombinatio...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2010.131

    authors： Gong S,Kus L,Heintz N

    更新日期：2010-09-01 00:00:00

  • Cloning of ground-state intestinal stem cells from endoscopic biopsy samples.

    abstract：:'Adult' or 'somatic' stem cells harbor an intrinsic ability to regenerate tissues. Heterogeneity of such stem cells along the gastrointestinal tract yields the known segmental specificity of this organ and may contribute to the pathology of certain enteric conditions. Here we detail technology for the generation of 'l...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/s41596-020-0298-4

    authors： Duleba M,Yamamoto Y,Neupane R,Rao W,Xie J,Qi Y,Liew AA,Niroula S,Zhang Y,Mahalingam R,Wang S,Goller K,Ajani JA,Vincent M,Ho KY,Hou JK,Hyams JS,Sylvester FA,Crum CP,McKeon F,Xian W

    更新日期：2020-05-01 00:00:00

  • Tracking recent adaptive evolution in microbial species using TimeZone.

    abstract：:An important goal of the analysis of sequenced genomes of microbial pathogens is to improve the therapy of infectious diseases. In this context, a major challenge is to detect genomic-level evolutionary changes that increase microbial virulence. TimeZone, a genome analysis software package, is designed to detect footp...

    journal_title：Nature protocols

    pub_type： 杂志文章

    doi：10.1038/nprot.2013.031

    authors： Chattopadhyay S,Paul S,Dykhuizen DE,Sokurenko EV

    更新日期：2013-04-01 00:00:00