Attenuation of glyceraldehyde-3-phosphate dehydrogenase activity and ATP levels in rat brain synaptosomes by acrylamide

Abstract

The effect of neurotoxin acrylamide (AC) on energy metabolism has been studied in a purified preparation of the synaptosomes. The synaptosomes were prepared by the flotation technique in a discontinuous Ficoll/sucrose gradient. The purity of the synaptosomes was checked by electron microscopy and by assaying the activity of marker enzymes. By these criterias, free mitochondrial contamination in the synaptosomes was found to be > 2%. Incubation of the synaptosomes with different concentrations of AC (2.5, 5.0, and 10mM) produced a concentration-dependent inhibition (15, 35, and 60%, respectively) of glyceraldehyde-3-phosphate dehydrogenase activity. Acrylamide also produced a time-dependent decrease of ATP concentrations in the synaptosomes; about 25% loss of ATP was seen within 1h, while about 60% ATP was lost after 120 min incubation with 10 mM AC. The effect of known inhibitors of glycolysis-iodoacetic acid (IAA), and of oxidative phophorylation-rotenone and antimycin A, was also studied on ATP synthesis by the synaptosomes. IAA was found to be the most potent inhibitor of ATP synthesis, while both rotenone and antimycin A were equally effective in blocking ATP synthesis in the synaptosomes. These studies show that the synaptosome might be used as a suitable in vitro model to study the effect of neurotoxin such as AC on neuronal energy metabolism.