Click
here to close Hello! We notice that
you are using Internet Explorer, which is not supported by Echinobase
and may cause the site to display incorrectly. We suggest using a
current version of Chrome,
FireFox,
or Safari.
Quantification of exocytosis kinetics by DIC image analysis of cortical lawns.
Mooney J
,
Thakur S
,
Kahng P
,
Trapani JG
,
Poccia D
.
???displayArticle.abstract???
Cortical lawns prepared from sea urchin eggs have offered a robust in vitro system for study of regulated exocytosis and membrane fusion events since their introduction by Vacquier almost 40 years ago (Vacquier in Dev Biol 43:62-74, 1975). Lawns have been imaged by phase contrast, darkfield, differential interference contrast, and electron microscopy. Quantification of exocytosis kinetics has been achieved primarily by light scattering assays. We present simple differential interference contrast image analysis procedures for quantifying the kinetics and extent of exocytosis in cortical lawns using an open vessel that allows rapid solvent equilibration and modification. These preparations maintain the architecture of the original cortices, allow for cytological and immunocytochemical analyses, and permit quantification of variation within and between lawns. When combined, these methods can shed light on factors controlling the rate of secretion in a spatially relevant cellular context. We additionally provide a subroutine for IGOR Pro® that converts raw data from line scans of cortical lawns into kinetic profiles of exocytosis. Rapid image acquisition reveals spatial variations in time of initiation of individual granule fusion events with the plasma membrane not previously reported.
Avery,
A cell-free system for regulated exocytosis in PC12 cells.
2000, Pubmed
Avery,
A cell-free system for regulated exocytosis in PC12 cells.
2000,
Pubmed
Blank,
A kinetic analysis of calcium-triggered exocytosis.
2001,
Pubmed
,
Echinobase
Blank,
Submaximal responses in calcium-triggered exocytosis are explained by differences in the calcium sensitivity of individual secretory vesicles.
1998,
Pubmed
,
Echinobase
Chen,
SNARE-mediated lipid mixing depends on the physical state of the vesicles.
2006,
Pubmed
Churchward,
Specific lipids supply critical negative spontaneous curvature--an essential component of native Ca2+-triggered membrane fusion.
2008,
Pubmed
Churchward,
Cholesterol facilitates the native mechanism of Ca2+-triggered membrane fusion.
2005,
Pubmed
,
Echinobase
Coorssen,
Biochemical and functional studies of cortical vesicle fusion: the SNARE complex and Ca2+ sensitivity.
1998,
Pubmed
,
Echinobase
Coorssen,
Regulated secretion: SNARE density, vesicle fusion and calcium dependence.
2003,
Pubmed
,
Echinobase
Decker,
Characterization of cortical secretory vesicles from the sea urchin egg.
1983,
Pubmed
,
Echinobase
Detering,
Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs.
1977,
Pubmed
,
Echinobase
Kaplan,
A rapid-flow perfusion chamber for high-resolution microscopy.
1996,
Pubmed
,
Echinobase
Kinsey,
Isolation and partial characterization of the plasma membrane of the sea urchin egg.
1980,
Pubmed
,
Echinobase
Kopf,
Isolation and characterization of sea urchin egg cortical granules.
1982,
Pubmed
,
Echinobase
Raveh,
Observations of calcium dynamics in cortical secretory vesicles.
2012,
Pubmed
,
Echinobase
Rogasevskaia,
A new approach to the molecular analysis of docking, priming, and regulated membrane fusion.
2011,
Pubmed
,
Echinobase
Steinhardt,
Intracellular calcium release at fertilization in the sea urchin egg.
1977,
Pubmed
,
Echinobase
Sudhof,
The synaptic vesicle cycle.
2004,
Pubmed
Terasaki,
Characterization of sea urchin egg endoplasmic reticulum in cortical preparations.
1991,
Pubmed
,
Echinobase
Terasaki,
Characterization of endoplasmic reticulum by co-localization of BiP and dicarbocyanine dyes.
1992,
Pubmed
Terasaki,
Demonstration of calcium uptake and release by sea urchin egg cortical endoplasmic reticulum.
1991,
Pubmed
,
Echinobase
Terasaki,
Visualization of exocytosis during sea urchin egg fertilization using confocal microscopy.
1995,
Pubmed
,
Echinobase
Trikash,
Fusion of synaptic vesicles and plasma membrane in the presence of synaptosomal soluble proteins.
2006,
Pubmed
Vacquier,
The isolation of intact cortical granules from sea urchin eggs: calcium lons trigger granule discharge.
1975,
Pubmed
,
Echinobase
Vogel,
Poisson-distributed active fusion complexes underlie the control of the rate and extent of exocytosis by calcium.
1996,
Pubmed
,
Echinobase
Vogel,
Proteins on exocytic vesicles mediate calcium-triggered fusion.
1992,
Pubmed
,
Echinobase
Wojcik,
Regulation of membrane fusion in synaptic excitation-secretion coupling: speed and accuracy matter.
2007,
Pubmed
Wong,
FRAP analysis of secretory granule lipids and proteins in the sea urchin egg.
2008,
Pubmed
,
Echinobase
Zimmerberg,
Exocytosis of sea urchin egg cortical vesicles in vitro is retarded by hyperosmotic sucrose: kinetics of fusion monitored by quantitative light-scattering microscopy.
1985,
Pubmed
,
Echinobase
