Influence of the malate-aspartate shuttle on oxidative metabolism in synaptosomes.
beta-Methyleneaspartate, a specific inhibitor of aspartate aminotransferase (EC 2.6.1.1.), was used to investigate the role of the malate-aspartate shuttle in rat brain synaptosomes. Incubation of rat brain cytosol, "free" mitochondria, synaptosol, and synaptic mitochondria, with 2 mM beta-methyleneaspartate resulted in inhibition of aspartate aminotransferase by 69%, 67%, 49%, and 76%, respectively. The reconstituted malate-aspartate shuttle of "free" brain mitochondria was inhibited by a similar degree (53%). As a consequence of the inhibition of the aspartate aminotransferase, and hence the malate-aspartate shuttle, the following changes were observed in synaptosomes: decreased glucose oxidation via the pyruvate dehydrogenase reaction and the tricarboxylic acid cycle; decreased acetylcholine synthesis; and an increase in the cytosolic redox state, as measured by the lactate/pyruvate ratio. The main reason for these changes can be attributed to decreased carbon flow through the tricarboxylic acid cycle (i.e., decreased formation of oxaloacetate), rather than as a direct consequence of changes in the NAD+/NADH ratio. Malate/glutamate oxidation in "free" mitochondria was also decreased in the presence of 2 mM beta-methyleneaspartate. This is probably a result of decreased glutamate transport into mitochondria as a result of low levels of aspartate, which are needed for the exchange with glutamate by the energy-dependent glutamate-aspartate translocator.
Cheeseman
A J
AJ
Department of Biochemistry, Medical College, St. Bartholomew's Hospital, University of London, England.
Clark
J B
JB
D000109:Acetylcholine / Q000096:biosynthesis; D000818:Animals; D001219:Aspartate Aminotransferases / Q000037:antagonists & inhibitors; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D001921:Brain / Q000201:enzymology*; D003600:Cytosol / Q000201:enzymology; D004734:Energy Metabolism / Q000187:drug effects; D005947:Glucose / Q000378:metabolism; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D008293:Malates / Q000378:metabolism*; D008297:Male; D008928:Mitochondria / Q000201:enzymology; D010084:Oxidation-Reduction; D010101:Oxygen Consumption / Q000187:drug effects; D011773:Pyruvates / Q000378:metabolism; D019289:Pyruvic Acid; D051381:Rats; D011919:Rats, Inbred Strains; D013574:Synaptosomes / Q000201:enzymology*
D007773:Lactates; D008293:Malates; D011773:Pyruvates; D001224:Aspartic Acid; D019344:Lactic Acid; C036683:beta-methyleneaspartate; C030298:malic acid; D019289:Pyruvic Acid; D001219:Aspartate Aminotransferases; D005947:Glucose; D000109:Acetylcholine