title: Localized Ca2+ entry preferentially effects protein dephosphorylation, phosphorylation, and glutamate release.
abstract: The release of neurotransmitter glutamate from isolated nerve terminals (synaptosomes) was found to be tightly coupled to the entry of Ca2+ through voltage-dependent Ca2+ channels, but is relatively unresponsive to "bulk" increases in cytosolic Ca2+ concentrations ([Ca2+]c) effected by Ca2+ ionophore. Under the same conditions, this dependence on Ca2+ influx, specifically through Ca2+ channels, was also seen for the dephosphorylation of a 96-kDa protein, (P96), present in the nerve terminals, as well as the phosphorylation of proteins migrating at 75 kDa (P75), corresponding to the synapsins, a group of well characterized synaptic vesicle-associated proteins. P96 dephosphorylation, following Ca2+ influx, was persistent and insensitive to the phosphatase inhibitor okadaic acid, suggesting a phosphatase other than protein phosphatase 1 and 2A as being responsible. Perhaps through the same phosphatase activity the increase in P75 phosphorylation was rapidly reversed with a time course similar to P96 dephosphorylation. When release, P96 dephosphorylation, and P75 phosphorylation were considered as functions of the [Ca2+]c increases achieved by depolarization and Ca2+ ionophore, there was no correlation of any of these with the overall concentration of Ca2+ in the cytosol. Since the fura-2 method used to measure [Ca2+] gives an averaged [Ca2+]c, these results imply that the release and protein dephosphorylation events are functionally coupled to local [Ca2+]c, in the immediate vicinity of Ca2+ channels. The reported clustering of the latter at the active zone area of the synapse and the parallelism between synaptic vesicle exocytosis and the phosphorylation of synaptic vesicle-associated proteins (p75:synapsins Ia/Ib), suggests that P96 may be similarly localized at the active zone area and, therefore, may be of significance in a modulatory role in glutamate release.
authors.affiliation: Department of Biochemistry, University of Dundee, Scotland.
authors.forename: T S
authors.initials: TS
authors.lastname: Sihra
authors.forename: E
authors.initials: E
authors.lastname: Bogonez
authors.forename: D G
authors.initials: DG
authors.lastname: Nicholls
mesh_terms: D015761:4-Aminopyridine / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism; D002540:Cerebral Cortex / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D004988:Ethers, Cyclic / Q000494:pharmacology; D005971:Glutamates / Q000378:metabolism*; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D009419:Nerve Tissue Proteins / Q000302:isolation & purification / Q000378:metabolism*; D019319:Okadaic Acid; D010449:Peptide Mapping; D010710:Phosphates / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D011189:Potassium Chloride / Q000494:pharmacology; D054645:Protein Phosphatase 1; D051381:Rats; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism*; D013997:Time Factors
chemical_list: D015220:Calcium Channels; D004988:Ethers, Cyclic; D005971:Glutamates; D009419:Nerve Tissue Proteins; D010710:Phosphates; D010750:Phosphoproteins; D019319:Okadaic Acid; D015759:Ionomycin; D011189:Potassium Chloride; D015761:4-Aminopyridine; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D002118:Calcium