Abstract
In order to elucidate the molecular mechanism of phorbol ester-induced potentiation of neurotransmitter release, changes in the subcellular distribution of secretory vesicles were studied in PC12 cells. Dopamine (DA) and acetylcholine containing vesicles were selectively labelled by expressing green fluorescent protein-conjugated vesicular monoamine transporter and vesicular acetylcholine transporter, respectively. In the resting state, these vesicles were distributed throughout the cytoplasm. Phorbol-12-myristate-13-acetate (PMA), but not the inactive analogue 4 alpha-PMA, induced a redistribution of both types of secretory vesicles near the plasma membrane, and this change was abolished by a protein kinase C (PKC) inhibitor, bisindolylmaleimide I (BIS). PMA also induced a marked enhancement of depolarization-induced DA release and phosphorylation of SNAP-25 at Ser187. BIS completely inhibited PMA-induced SNAP-25 phosphorylation but suppressed PMA-induced enhancement of DA release only partially. These results suggest that PMA enhances neurotransmitter release from PC12 cells by both PKC-dependent and PKC-independent mechanisms, and PKC enhances neurotransmitter release by recruiting secretory vesicles to the plasma membrane.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Acetylcholine / metabolism
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Animals
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Carcinogens / pharmacology
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Carrier Proteins / drug effects
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Carrier Proteins / metabolism
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Cell Membrane / drug effects
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Cell Membrane / enzymology*
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Central Nervous System / cytology
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Central Nervous System / enzymology*
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Central Nervous System / metabolism
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Dopamine / metabolism
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Enzyme Inhibitors / pharmacology
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Green Fluorescent Proteins
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Indicators and Reagents / metabolism
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Luminescent Proteins / metabolism
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Membrane Glycoproteins / drug effects
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Membrane Glycoproteins / metabolism
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Membrane Proteins / drug effects
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Membrane Proteins / metabolism
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Membrane Transport Proteins*
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Nerve Tissue Proteins / drug effects
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Nerve Tissue Proteins / metabolism
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Neuropeptides*
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Neurotransmitter Agents / metabolism*
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PC12 Cells
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Presynaptic Terminals / drug effects
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Presynaptic Terminals / enzymology*
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Presynaptic Terminals / metabolism
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Protein Kinase C / drug effects
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Protein Kinase C / metabolism*
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Protein Transport / drug effects
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Protein Transport / physiology*
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Rats
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Red Fluorescent Protein
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Secretory Vesicles / drug effects
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Secretory Vesicles / enzymology*
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Secretory Vesicles / metabolism
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Synaptic Vesicles / drug effects
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Synaptic Vesicles / enzymology
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Synaptosomal-Associated Protein 25
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Tetradecanoylphorbol Acetate / pharmacology
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Vesicular Acetylcholine Transport Proteins
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Vesicular Biogenic Amine Transport Proteins
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Vesicular Monoamine Transport Proteins
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Vesicular Transport Proteins*
Substances
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Carcinogens
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Carrier Proteins
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Enzyme Inhibitors
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Indicators and Reagents
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Luminescent Proteins
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Membrane Glycoproteins
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Membrane Proteins
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Membrane Transport Proteins
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Nerve Tissue Proteins
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Neuropeptides
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Neurotransmitter Agents
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Slc18a3 protein, rat
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Snap25 protein, rat
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Synaptosomal-Associated Protein 25
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Vesicular Acetylcholine Transport Proteins
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Vesicular Biogenic Amine Transport Proteins
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Vesicular Monoamine Transport Proteins
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Vesicular Transport Proteins
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Green Fluorescent Proteins
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Protein Kinase C
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Acetylcholine
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Tetradecanoylphorbol Acetate
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Dopamine