Cholinergic pathways serve important functions in learning and memory processes. The loss of basal forebrain cholinergic neurons and the presence of senile plaques composed by amyloid beta-peptide (A beta) are found in post-mortem brains of Alzheimer's disease (AD) patients. However, the role of A beta in the cholinergic dysfunction observed in AD is not yet clarified. In this study, we observed that the release of [3H]acetylcholine evoked by K(+)-depolarization was significantly lower in cells treated with A beta 25-35 peptide, than in untreated cells or in cells exposed to the reverse sequence peptide A beta 35-25. The levels of pyruvate, the substrate for pyruvate dehydrogenase, the enzyme involved in acetyl coenzyme A synthesis in the brain, which is rate-limiting for the synthesis of acetylcholine, were significantly decreased, about 40%, in A beta treated cells. A beta 25-35 did not affect choline acetyltransferase activity or [3H]choline uptake. 2-[3H]-deoxyglucose uptake was decreased when cells were exposed to A beta 25-35 or to A beta 1-40. Taken together these data suggest that an impairment of glycolysis, and the consequent decrease in pyruvate levels, may be responsible for the decrement of acetylcholine release observed in A beta treated cells, thus sustaining the hypothesis that the cholinergic dysfunction, observed in AD patients, might be associated with extracellular A beta accumulation.