Abstract
The activation of G protein-coupled receptors (GPCRs) can result in an inhibition of Ca(2+)-dependent hormone and neurotransmitter secretion. This has been attributed in part to G protein inhibition of Ca(2+) influx. However, a frequently dominant inhibitory effect, of unknown mechanism, also occurs distal to Ca(2+) entry. Here we characterize direct inhibitory actions of G protein betagamma (Gbetagamma) on Ca(2+)-triggered vesicle exocytosis in permeable PC12 cells. Gbetagamma inhibition was rapid (<1 s) and was attenuated by cleavage of synaptosome-associated protein of 25 kD (SNAP25). Gbetagamma bound soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes, and binding was reduced to SNARE complexes containing cleaved SNAP25 or by Ca(2+)-dependent synaptotagmin binding. Here we show inhibitory coupling between GPCRs and vesicle exocytosis mediated directly by Gbetagamma interactions with the Ca(2+)-dependent fusion machinery.
Publication types
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Comparative Study
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Blotting, Western / methods
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Calcium / pharmacology
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Calcium-Binding Proteins / metabolism
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Cattle
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Dose-Response Relationship, Drug
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Drug Interactions
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Exocytosis / drug effects
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Exocytosis / physiology*
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Eye Proteins / pharmacology
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GTP-Binding Protein Regulators
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GTP-Binding Protein beta Subunits / pharmacology
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GTP-Binding Protein beta Subunits / physiology*
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GTP-Binding Protein gamma Subunits / pharmacology
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GTP-Binding Protein gamma Subunits / physiology*
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Membrane Glycoproteins / metabolism
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Membrane Proteins / pharmacology
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Nerve Tissue Proteins / metabolism
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Nerve Tissue Proteins / pharmacology
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Neural Inhibition / drug effects
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Norepinephrine / metabolism
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PC12 Cells
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Phosphoproteins / pharmacology
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Protein Binding / drug effects
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Rats
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Reaction Time / drug effects
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Recombinant Fusion Proteins / metabolism
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SNARE Proteins
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Secretory Vesicles / drug effects
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Secretory Vesicles / metabolism*
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Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins
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Synaptosomal-Associated Protein 25
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Synaptotagmins
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Time Factors
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Vesicular Transport Proteins / metabolism*
Substances
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Calcium-Binding Proteins
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Eye Proteins
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GTP-Binding Protein Regulators
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GTP-Binding Protein beta Subunits
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GTP-Binding Protein gamma Subunits
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Membrane Glycoproteins
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Membrane Proteins
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Nerve Tissue Proteins
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Phosphoproteins
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Recombinant Fusion Proteins
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SNARE Proteins
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Snap25 protein, rat
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Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins
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Synaptosomal-Associated Protein 25
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Vesicular Transport Proteins
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phosducin
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Synaptotagmins
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Calcium
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Norepinephrine