Abstract
Subplasmalemmal Ca2+, dynamically equilibrated with extracellular Ca2+, affects numerous signaling molecules, effectors, and events within this restricted space. We demonstrated the presence of a novel Ca2+ wave propagating beneath the plasma membrane in response to acute elevation of extracellular [Ca2+], by targeting a Ca2+ sensor, cameleon, to the endothelial plasmalemma. These subcortical waves, spatially distinct from classical cytosolic Ca2+ waves, originated in localized regions and propagated throughout the subplasmalemma. Translocation of an expressed GFP fused with a PH domain of PLC from the plasma membrane to the cytosol accompanied these subcortical waves, and U73122 attenuated not only the GFP-PH translocation, but also the peak amplitude of the subcortical Ca2+ waves; this finding suggests the involvement of local IP3 production through PLC-mediated PIP2 hydrolysis in the initiation of these waves. Changes in NO production as well as PKCbeta-GFP translocation from the cytosol to the plasma membrane, but not of GFP-PLA2 to perinuclear endomembranes, were associated with the subplasmalemmal Ca2+ changes. Thus, extracellular Ca2+ maintains the basal PLC activity of the plasma membrane, is involved in the initiation of compartmentalized subcortical Ca2+ waves, and regulates Ca2+-dependent signaling molecules residing in or translocated to the plasma membrane. The full text of this article is available online at http://circres.ahajournals.org.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Motifs
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Calcium / metabolism
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Calcium Signaling*
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Calcium-Binding Proteins / analysis
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Calcium-Binding Proteins / metabolism
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Cell Compartmentation
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Cell Membrane / physiology*
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Cytosol / metabolism
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Endothelial Cells / physiology*
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Endothelial Cells / ultrastructure
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Endothelium, Vascular / cytology*
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Endothelium, Vascular / physiology
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Estrenes / pharmacology
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Extracellular Fluid / metabolism
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Fluorescence Resonance Energy Transfer
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Fluorescent Dyes / analysis
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GAP-43 Protein / genetics
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Genes, Reporter
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Green Fluorescent Proteins / genetics
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Ion Transport / physiology
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Isoenzymes / genetics
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Membrane Proteins / physiology*
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Microscopy, Confocal
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Nitric Oxide / metabolism
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Phosphatidylinositol 4,5-Diphosphate / metabolism
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Phospholipase C delta
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Phospholipases A / genetics
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Phospholipases A2
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Protein Kinase C / genetics
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Protein Kinase C / metabolism
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Protein Kinase C beta
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Protein Structure, Tertiary
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Protein Transport
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Pyrrolidinones / pharmacology
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Type C Phospholipases / genetics
Substances
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Calcium-Binding Proteins
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Estrenes
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Fluorescent Dyes
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GAP-43 Protein
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Isoenzymes
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Membrane Proteins
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Phosphatidylinositol 4,5-Diphosphate
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Pyrrolidinones
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Recombinant Fusion Proteins
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green fluorescent protein, Aequorea victoria
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yellow cameleon
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1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione
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Green Fluorescent Proteins
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Nitric Oxide
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Protein Kinase C
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Protein Kinase C beta
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Phospholipases A
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Phospholipases A2
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Type C Phospholipases
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Phospholipase C delta
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Calcium