title: Internalization of beta-amyloid peptide by primary neurons in the absence of apolipoprotein E.
abstract: Extracellular accumulation of beta-amyloid peptide (Abeta) has been linked to the development of Alzheimer disease. The importance of intraneuronal Abeta has been recognized more recently. Although considerable evidence indicates that extracellular Abeta contributes to the intracellular pool of Abeta, the mechanisms involved in Abeta uptake by neurons are poorly understood. We examined the molecular mechanisms involved in Abeta-(1-42) internalization by primary neurons in the absence of apolipoprotein E. We demonstrated that Abeta-(1-42) is more efficiently internalized by axons than by cell bodies of sympathetic neurons, suggesting that Abeta-(1-42) uptake might be mediated by proteins enriched in the axons. Although the acetylcholine receptor alpha7nAChR, previously suggested to be involved in Abeta internalization, is enriched in axons, our results indicate that it does not mediate Abeta-(1-42) internalization. Moreover, receptors of the low density lipoprotein receptor family are not essential for Abeta-(1-42) uptake in the absence of apolipoprotein E because receptor-associated protein had no effect on Abeta uptake. By expressing the inactive dynamin mutant dynK44A and the clathrin hub we found that Abeta-(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 Abeta did not co-localize with the early endosome marker EEA1 further support a clathrin-independent mechanism. The lack of co-localization of Abeta with caveolin in intracellular vesicles and the normal uptake of Abeta by neurons that do not express caveolin indicate that Abeta does not require caveolin either. Instead partial co-localization of Abeta-(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 Abeta internalization might identify potential therapeutic targets for Alzheimer disease.
authors.affiliation: Department of Pharmacology, University of Alberta, Edmonton, Canada.
authors.forename: Lucila
authors.initials: L
authors.lastname: Saavedra
authors.forename: Amany
authors.initials: A
authors.lastname: Mohamed
authors.forename: Victoria
authors.initials: V
authors.lastname: Ma
authors.forename: Satyabrata
authors.initials: S
authors.lastname: Kar
authors.forename: Elena Posse
authors.initials: EP
authors.lastname: de Chaves
mesh_terms: D000320:Adrenergic Fibers / Q000378:metabolism; D000544:Alzheimer Disease / Q000235:genetics / Q000378:metabolism* / Q000628:therapy; D016229:Amyloid beta-Peptides / Q000378:metabolism*; D000818:Animals; D001057:Apolipoproteins E; D001369:Axons / Q000378:metabolism*; D021941:Caveolae / Q000378:metabolism*; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D002966:Clathrin / Q000235:genetics / Q000378:metabolism; D034281:Dynamins / Q000235:genetics / Q000378:metabolism; D004705:Endocytosis / Q000235:genetics; D010455:Peptides / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011978:Receptors, Nicotinic / Q000235:genetics / Q000378:metabolism; D013107:Sphingolipids / Q000378:metabolism; D064569:alpha7 Nicotinic Acetylcholine Receptor
chemical_list: D016229:Amyloid beta-Peptides; D001057:Apolipoproteins E; C579165:Chrna7 protein, rat; D002966:Clathrin; D010455:Peptides; D011978:Receptors, Nicotinic; D013107:Sphingolipids; D064569:alpha7 Nicotinic Acetylcholine Receptor; D002772:Cholera Toxin; D002784:Cholesterol; D034281:Dynamins