Internalization of β-amyloid peptide by primary neurons in the absence of apolipoprotein E

Extracellular accumulation of beta-amyloid peptide (A beta) has been linked to the development of Alzheimer disease. The importance of intraneuronal A beta has been recognized more recently. Although considerable evidence indicates that extracellular A beta contributes to the intracellular pool of A beta, the mechanisms involved in A beta uptake by neurons are poorly understood. We examined the molecular mechanisms involved in A beta-(1-42) internalization by primary neurons in the absence of apolipoprotein E. We demonstrated that A beta-(1-42) is more efficiently internalized by axons than by cell bodies of sympathetic neurons, suggesting that A beta-(1 42) uptake might be mediated by proteins enriched in the axons. Although the acetylcholine receptor alpha 7nAChR, previously suggested to be involved in A beta internalization, is enriched in axons, our results indicate that it does not mediate A beta-(1-42) internalization. Moreover, receptors of the low density lipoprotein receptor family are not essential for A beta-(1-42) uptake in the absence of apolipoprotein E because receptor-associated protein had no effect on A beta uptake. By expressing the inactive dynamin mutant dynK44A and the clathrin hub we found that A beta-(1-42) internalization is independent of clathrin but dependent on dynamin, which suggests an endocytic pathway involving caveolae/lipid rafts. Confocal microscopy studies showing that A beta did not co-localize with the early endosome marker EEA1 further support a clathrin-independent mechanism. The lack of co-localization of A beta with caveolin in intracellular vesicles and the normal uptake of A beta by neurons that do not express caveolin indicate that A beta does not require caveolin either. Instead partial co-localization of A beta-(1-42) with cholera toxin subunit B and sensitivity to reduction of cellular cholesterol and sphingolipid levels suggest a caveolae-independent, raft-mediated mechanism. Understanding the molecular events involved in neuronal A beta internalization might identify potential therapeutic targets for Alzheimer disease.