title: Glutamate exocytosis from cerebellar granule cells: the mechanism of a transition to an L-type Ca2+ channel coupling.
abstract: When cerebellar granule cells in the presence of 1.3 mM calcium chloride (Ca2+) are depolarized by high potassium chloride (KCl), the release of endogenous glutamate is coupled to a high threshold Ca2+ channel blocked by the spider toxin omega Agatoxin-glutamate-release-inhibitor (Aga-GI) and insensitive to the L-type voltage-dependent Ca2+ channel-inhibitor nifedipine. A prolonged KCl depolarization in the absence of Ca2+ followed by addition of 5 mM Ca2+ results in an enhanced nifedipine-sensitive Ca2+ entry; glutamate exocytosis retains sensitivity to tetanus toxin and bafilomycin A1, is now totally inhibited by nifedipine and shows greatly reduced sensitivity to AGA-GI. Single cell Ca2+ imaging indicates that the L-type channel modulating release is preferentially located at somatic regions rather than neurites. A different pattern of vesicle endocytosis monitored with the fluorescent indicator FM1-43 is seen in response to the two depolarization protocols. Furthermore, vesicles loaded during depolarization with high KCl in the presence of 5 mM Ca2+ extensively exocytose dye in a nifedipine-insensitive manner in response to a second similar stimulation but release little dye in response to stimulus with high KCl in the absence of Ca2+ followed by the addition of 5 mM Ca2+. In contrast, vesicles loaded by stimulating with KCl in the absence of Ca2+ followed by the addition of 5 mM Ca2+ can be released by a second similar stimulus and this release is sensitive to nifedipine. Nifedipine sensitivity is not induced in cerebellar synaptosomes subjected to stimulation with high KCl in the absence of Ca2+ followed by the re-addition of 5 mM Ca2+. The results indicate that different populations of channels and vesicles may be functional during two depolarization protocols.
authors.affiliation: Department of Biochemistry, University of Dundee, Scotland, UK.
authors.forename: J M
authors.initials: JM
authors.lastname: Pocock
authors.forename: M A
authors.initials: MA
authors.lastname: Cousin
authors.forename: J
authors.initials: J
authors.lastname: Parkin
authors.forename: D G
authors.initials: DG
authors.lastname: Nicholls
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology*; D002531:Cerebellum / Q000502:physiology*; D005089:Exocytosis; D018698:Glutamic Acid / Q000378:metabolism*; D008564:Membrane Potentials; D009543:Nifedipine / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors
chemical_list: D015220:Calcium Channels; D018698:Glutamic Acid; D009543:Nifedipine; D011188:Potassium; D002118:Calcium