Abstract
Synaptic vesicles can be retrieved rapidly or slowly, but the molecular basis of these kinetic differences has not been defined. We now show that substantially different sets of molecules mediate fast and slow endocytosis in the synaptic terminal of retinal bipolar cells. Capacitance measurements of membrane retrieval were made in terminals in which peptides and protein domains were introduced to disrupt known interactions of clathrin, the AP2 adaptor complex, and amphiphysin. All these manipulations caused a selective inhibition of the slow phase of membrane retrieval (time constant approximately 10 s), leaving the fast phase (approximately 1 s) intact. Slow endocytosis after strong stimulation was therefore dependent on the formation of clathrin-coated membrane. Fast endocytosis occurring after weaker stimuli retrieves vesicle membrane in a clathrin-independent manner. All compensatory endocytosis required GTP hydrolysis, but only a subset of released vesicles were primed for fast, clathrin-independent endocytosis.
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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Adaptor Protein Complex 2 / metabolism
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Adenosine Triphosphate / pharmacology
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Animals
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Blotting, Western / methods
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Clathrin / metabolism*
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Dose-Response Relationship, Drug
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Drug Interactions
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Dynamins / metabolism
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Electric Capacitance
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Endocytosis / drug effects
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Endocytosis / physiology*
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Goldfish
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Guanosine 5'-O-(3-Thiotriphosphate) / pharmacology
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Guanosine Triphosphate / metabolism
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Guanosine Triphosphate / pharmacology
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Hydrolysis / drug effects
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Membrane Potentials / drug effects
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Membrane Potentials / physiology
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Membrane Potentials / radiation effects
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Nerve Tissue Proteins / chemistry
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Nerve Tissue Proteins / metabolism
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Neurons / cytology*
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Neurons / metabolism
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Patch-Clamp Techniques / methods
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Peptide Fragments / pharmacology
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Protein Binding / drug effects
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Protein Binding / physiology
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Protein Structure, Tertiary / physiology
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Rats
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Recombinant Proteins / pharmacology
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Retina / cytology*
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Retina / metabolism
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Synaptic Vesicles / classification*
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Synaptic Vesicles / metabolism*
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Time Factors
Substances
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Adaptor Protein Complex 2
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Clathrin
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Nerve Tissue Proteins
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Peptide Fragments
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Recombinant Proteins
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amphiphysin
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Guanosine 5'-O-(3-Thiotriphosphate)
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Guanosine Triphosphate
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Adenosine Triphosphate
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Dynamins