Aggregation of the amyloid-β-42 (Aβ42) peptide in the brain parenchyma is a pathological hallmark of Alzheimer's disease (AD), and the prevention of Aβ aggregation has been proposed as a therapeutic intervention in AD. However, recent reports indicate that Aβ can form several different prefibrillar and fibrillar aggregates and that each aggregate may confer different pathogenic effects, suggesting that manipulation of Aβ42 aggregation may not only quantitatively but also qualitatively modify brain pathology. Here, we compare the pathogenicity of human Aβ42 mutants with differing tendencies to aggregate. We examined the aggregation-prone, EOFAD-related Arctic mutation (Aβ42Arc) and an artificial mutation (Aβ42art) that is known to suppress aggregation and toxicity of Aβ42 in vitro. In the Drosophila brain, Aβ42Arc formed more oligomers and deposits than did wild type Aβ42, while Aβ42art formed fewer oligomers and deposits. The severity of locomotor dysfunction and premature death positively correlated with the aggregation tendencies of Aβ peptides. Surprisingly, however, Aβ42art caused earlier onset of memory defects than Aβ42. More remarkably, each Aβ induced qualitatively different pathologies. Aβ42Arc caused greater neuron loss than did Aβ42, while Aβ42art flies showed the strongest neurite degeneration. This pattern of degeneration coincides with the distribution of Thioflavin S-stained Aβ aggregates: Aβ42Arc formed large deposits in the cell body, Aβ42art accumulated preferentially in the neurites, while Aβ42 accumulated in both locations. Our results demonstrate that manipulation of the aggregation propensity of Aβ42 does not simply change the level of toxicity, but can also result in qualitative shifts in the pathology induced in vivo.

The amyloid-␤42 (A␤42) peptide has been suggested to play a causative role in Alzheimer disease (AD). Neprilysin (NEP) is one of the rate-limiting A␤-degrading enzymes, and its enhancement ameliorates extracellular amyloid pathology, synaptic dysfunction, and memory defects in mouse models of A␤ amyloidosis. In addition to the extracellular A␤, intraneuronal A␤42 may contribute to AD pathogenesis. However, the protective effects of neuronal NEP expression on intraneuronal A␤42 accumulation and neurodegeneration remain elusive. In contrast, sustained NEP activation may be detrimental because NEP can degrade many physiological peptides, but its consequences in the brain are not fully understood. Using transgenic Drosophila expressing human NEP and A␤42, we demonstrated that NEP efficiently suppressed the formation of intraneuronal A␤42 deposits and A␤42-induced neuron loss. However, neuronal NEP overexpression reduced cAMP-responsive element-binding protein-mediated transcription, caused age-dependent axon degeneration, and shortened the life span of the flies. Interestingly, the mRNA levels of endogenous fly NEP genes and phosphoramidon-sensitive NEP activity declined during aging in fly brains, as observed in mammals. Taken together, these data suggest both the protective and detrimental effects of chronically high NEP activity in the brain. Down-regulation of NEP activity in aging brains may be an evolutionarily conserved phenomenon, which could predispose humans to developing late-onset AD. Alzheimer disease (AD)3 is a progressive neurodegenerative disease defined by two protein deposits in autopsied brains: extracellular amyloid plaques composed of the 40-or 42-amino acid ␤-amyloid peptides (A␤40 or A␤42), and intracellular neurofibrillary tangles composed of abnormally hyperphosphorylated microtubule-associated protein Tau (1). A␤ peptides are physiological metabolites of the amyloid precursor protein (APP) resulting from sequential cleavage by ␤-secretase and ␥-secretase complex, whose catalytic subunits are presenilin 1 (PS1) and presenilin 2 (PS2) (2). Molecular genetic studies of early onset familial AD patients have identified causative mutations in APP, PS1, and PS2 genes, and these mutations promote A␤42 production, aggregation, and stability against clearance (3). Thus, the increased A␤42 levels in the brain are believed to initiate AD pathogenesis.The mechanisms by which A␤42 reaches pathological levels in the brains of late-onset AD (LOAD) patients are not well understood. The steady state level of A␤42 in the brain reflects the balance between production and clearance, and a reduction in clearance activity would raise A␤42 levels. A deficiency in neprilysin (NEP), a major rate-limiting A␤-degrading enzyme (4, 5), accelerates formation of extracellular amyloid deposits (6), amyloid angiopathy (6), synaptic dysfunctions (7), and memory defects (7) caused by human A␤ in transgenic mice. In LOAD patients, NEP mRNA and pro-

phosphorylated tau antibodies tested. Method of fixation did not reveal amyloid plaques in young 3xTg-AD mice. Plaque deposition was sex dependent with females showing more plaques than males beginning at 9 months predominately in the subiculum and to a lesser degree in the hippocampus, cortex and amygdala. Conclusions: These results stress the importance of fixation method for the immunohistochemical visualization of tau epitopes in young 3xTg-AD mice, and that intraneuronal A␤ is not necessarily a precondition for the concurrent staining of tau epitopes in select brain regions in these mutant mice. Background: The loss of the cholinergic neurons of the Nucleus Basalis of Meynert in Alzheimer's disease (AD) patients has received a lot of attention since its discovery in the early 1970s, and most presently available pharmaceutical drugs for AD are thus designed to increase the transmission of the cholinergic system. Despite this intense focus on the cholinergic system in AD the reason for the cholinergic neuron loss is yet unknown. Many mouse models of AD have been generated, but few have shown to develop neuron degeneration. Furthermore, genetically modified mouse models of AD showing loss of cholinergic neurons are scarce. APP/PS1KI mice develop a neuron loss of about 50% in the CA1 of the hippocampal formation, a characteristic also found in AD patients, as well as behavioural deficits and a severe motor pathology at the age of 6 months. This study examined A␤ induced pathology and neuron loss in the cholinergic system of the double transgenic APP/PS1KI mouse model of AD. Methods: Immunohistochemistry, stereology, real time RT-PCR. Results: Swollen ChAT positive dystrophic neurites were found in proximity of plaques especially in the motor nuclei and the striatum, as well as in the cortex, hippocampal formation and thalamus corresponding to regions innervated by the cholinergic basal forebrain and pons complexes. Expression of the APP transgene was found in ChAT positive neurons of motor nuclei accompanied by robust intracellular A␤ accumulation, however, no neurons expressing the APP transgene and thus no neurons accumulating intracellular A␤ aggregates were found in either the forebrain or pons complexes or in the caudate putamen. Stereological quantification showed a 20-30% loss of ChAT positive neurons only in the motor nuclei Mo5 and 7N at 6 and 12 months in APP/PS1KI mice, as compared to PS1KI control mice. No loss of ChAT positive neurons was found in the cholinergic forebrain complex or in the caudate putamen. Conclusions: This study reports cholinergic cell loss correlating with the accumulation of intracellular A␤ in a mouse model of AD and thus supports the hypothesis of intracellular A␤ aggregates as an early pathological alteration triggering neurodegeneration in AD. P1-