title: Two mechanisms of synaptic vesicle recycling in rat brain nerve terminals.
abstract: KCl and 4-aminopyridine (4-AP) evoke glutamate release from rat brain cortical nerve terminals by voltage clamping or by Na(+) channel-generated repetitive action potentials, respectively. Stimulation by 4-AP but not KCl is largely mediated by protein kinase C (PKC). To determine whether KCl and 4-AP utilise the same mechanism to release glutamate, we correlated glutamate release with release of the hydrophobic synaptic vesicle (SV) marker FM2-10. A strong correlation was observed for increasing concentrations of KCl and after application of phorbol 12-myristate 13-acetate (PMA) or staurosporine. The parallel increase in exocytosis measured by two approaches suggested it occurred by a PKC-independent mechanism involving complete fusion of SVs with the plasma membrane. At low concentrations of 4-AP, alone or with staurosporine, glutamate and FM2-10 release also correlated. However, higher concentrations of 4-AP or of 4-AP plus PMA greatly increased glutamate release but did not further increase FM2-10 release. This divergence suggests that 4-AP recruits an additional mechanism of release during strong stimulation that is PKC dependent and is superimposed upon the first mechanism. This second mechanism is characteristic of kiss-and-run, which is not detectable by styryl dyes. Our data suggest that glutamate release in nerve terminals occurs via two mechanisms: (1) complete SV fusion, which is PKC independent; and (2) a kiss-and-run-like mechanism, which is PKC dependent. Recruitment of a second release mechanism may be a widespread means to facilitate neurotransmitter release in central neurons.
authors.affiliation: Cell Signalling Unit, Children's Medical Research Institute, Sydney, New South Wales, Australia. m.cousin@ed.ac.uk
authors.forename: M A
authors.initials: MA
authors.lastname: Cousin
authors.forename: P J
authors.initials: PJ
authors.lastname: Robinson
mesh_terms: D015761:4-Aminopyridine / Q000494:pharmacology; D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001921:Brain / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005456:Fluorescent Dyes; D018698:Glutamic Acid / Q000378:metabolism; D018942:Macrolides; D026902:Potassium Channel Blockers; D011189:Potassium Chloride / Q000494:pharmacology; D017729:Presynaptic Terminals / Q000187:drug effects / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D051381:Rats; D019311:Staurosporine / Q000494:pharmacology; D013572:Synaptic Vesicles / Q000187:drug effects / Q000378:metabolism*; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D025262:Vacuolar Proton-Translocating ATPases
chemical_list: D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; C073804:FM1 43; C086686:FM2 10; D005456:Fluorescent Dyes; D018942:Macrolides; D026902:Potassium Channel Blockers; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D018698:Glutamic Acid; D011189:Potassium Chloride; C040929:bafilomycin A1; D015761:4-Aminopyridine; D011493:Protein Kinase C; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate