Aspartate release from rat hippocampal synaptosomes.
Certain excitatory pathways in the rat hippocampus can release aspartate along with glutamate. This study utilized rat hippocampal synaptosomes to characterize the mechanism of aspartate release and to compare it with glutamate release. Releases of aspartate and glutamate from the same tissue samples were quantitated simultaneously. Both amino acids were released by 25 mM K(+), 300 microM 4-aminopyridine (4-AP) and 0.5 and 1 microM ionomycin in a predominantly Ca(2+)-dependent manner. For a roughly equivalent quantity of glutamate released, aspartate release was significantly greater during exposure to elevated [K(+)] than to 4-AP and during exposure to 0.5 than to 1 microM ionomycin. Aspartate release was inefficiently coupled to P/Q-type voltage-dependent Ca(2+) channels and was reduced by KB-R7943, an inhibitor of reversed Na(+)/Ca(2+) exchange. In contrast, glutamate release depended primarily on Ca(2+) influx through P/Q-type channels and was not significantly affected by KB-R7943. Pretreatment of the synaptosomes with tetanus toxin and botulinum neurotoxins C and F reduced glutamate release, but not aspartate release. Aspartate release was also resistant to bafilomycin A(1), an inhibitor of vacuolar H(+)-ATPase, whereas glutamate release was markedly reduced. (+/-) -Threo-3-methylglutamate, a non-transportable competitive inhibitor of excitatory amino acid transport, did not reduce aspartate release. Niflumic acid, a blocker of Ca(2+)-dependent anion channels, did not alter the release of either amino acid. Exogenous aspartate and aspartate recently synthesized from glutamate accessed the releasable pool of aspartate as readily as exogenous glutamate and glutamate recently synthesized from aspartate accessed the releasable glutamate pool. These results are compatible with release of aspartate from either a vesicular pool by a "non-classical" form of exocytosis or directly from the cytoplasm by an as-yet-undescribed Ca(2+)-dependent mechanism. In either case, they suggest aspartate is released mainly outside the presynaptic active zones and may therefore serve as the predominant agonist for extrasynaptic N-methyl-D-aspartate receptors.
Department of Pharmacology and Cancer Biology, Box 3813, 100B Research Park 2, Research Drive, Duke University Medical Center, Durham, NC 27710, USA.
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Bradford
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Nadler
D015761:4-Aminopyridine / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism*; D015153:Blotting, Western / Q000379:methods; D001905:Botulinum Toxins / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D005260:Female; D018698:Glutamic Acid / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology*; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000494:pharmacology; D009544:Niflumic Acid / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D026902:Potassium Channel Blockers / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013111:Spider Venoms / Q000494:pharmacology; D013574:Synaptosomes / Q000378:metabolism*; D013744:Tetanus Toxin / Q000494:pharmacology; D013890:Thiourea / Q000031:analogs & derivatives* / Q000494:pharmacology; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000494:pharmacology
C101670:2-(2-(4-(4-nitrobenzyloxy)phenyl)ethyl)isothiourea methanesulfonate; D002121:Calcium Channel Blockers; D016861:Cyclooxygenase Inhibitors; D018690:Excitatory Amino Acid Agonists; D007476:Ionophores; D026902:Potassium Channel Blockers; C115521:SNX 482; D013111:Spider Venoms; D013744:Tetanus Toxin; C035979:tetanospasmin; D001224:Aspartic Acid; D018698:Glutamic Acid; D009544:Niflumic Acid; D015759:Ionomycin; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; D015761:4-Aminopyridine; D008666:Metalloendopeptidases; D001905:Botulinum Toxins; D013890:Thiourea; D011188:Potassium; D002118:Calcium