title: Synaptic vesicle Ca2+/H+ antiport: dependence on the proton electrochemical gradient.
abstract: Synaptic vesicles isolated from sheep brain cortex accumulate Ca2+ by a mechanism of secondary active transport associated to the H(+)-pump activity. The process can be visualized either by measuring Ca(2+)-induced H+ release or DeltapH-dependent Ca2+ accumulation. We observed that the amount of Ca2+ taken up by the vesicles increases with the magnitude of the DeltapH across the membrane, particularly at Ca2+ concentrations (approximately 500 microM) found optimal for the antiporter activity. Similarly, H+ release induced by Ca2+ increased with the magnitude of DeltapH. However, above 60% DeltapH (high H(+)-pump activity), the net H+ release from the vesicles decreased as the pump-mediated H+ influx exceeded the Ca(2+)-induced H+ efflux. We also observed that the Ca2+/H+ antiport activity depends, essentially, on the DeltapH component of the electrochemical gradient (approximately 3 nmol Ca2+ taken up/mg protein), although the Deltaphi component may also support some Ca2+ accumulation by the vesicles (approximately 1 nmol/mg protein) in the absence of DeltapH. Both Ca(2+)-induced H+ release and DeltapH-dependent Ca2+ uptake could be driven by an artificially imposed proton motive force. Under normal conditions (H+ pump-induced DeltapH), the electrochemical gradient dependence of Ca2+ uptake by the vesicles was checked by inhibition of the process with specific inhibitors (bafilomycin A(1), ergocryptin, folymicin, DCCD) of the H(+)-pump activity. These results indicate that synaptic vesicles Ca2+/H+ antiport is indirectly linked to ATP hydrolysis and it is essentially dependent on the chemical component (DeltapH) of the electrochemical gradient generated by the H(+)-pump activity.
authors.affiliation: Centro de Biologia Celular, Departamento de Biologia, Universidade de Aveiro, 3810, Aveiro, Portugal. pgoncalves@biio.ua.pt
authors.forename: P P
authors.initials: PP
authors.lastname: Gonçalves
authors.forename: S M
authors.initials: SM
authors.lastname: Meireles
authors.forename: P
authors.initials: P
authors.lastname: Neves
authors.forename: M G
authors.initials: MG
authors.lastname: Vale
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D017920:Antiporters / Q000187:drug effects / Q000378:metabolism*; D001921:Brain / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000187:drug effects / Q000378:metabolism*; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D027682:Cation Transport Proteins; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004563:Electrochemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004873:Ergolines / Q000494:pharmacology; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D018942:Macrolides; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors; D012756:Sheep; D013050:Spectrometry, Fluorescence; D013572:Synaptic Vesicles / Q000187:drug effects / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D017920:Antiporters; D002135:Calcium-Binding Proteins; D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D004873:Ergolines; D018942:Macrolides; C027614:calcium-hydrogen antiporters; C100269:ergocryptine; D004024:Dicyclohexylcarbodiimide; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D006859:Hydrogen; C036978:concanamycin A; C040929:bafilomycin A1; D006180:Proton-Translocating ATPases