Analysis of microdissected human neurons by a sensitive ELISA reveals a correlation between elevated intracellular concentrations of Abeta42 and Alzheimer's disease neuropathology.
ABSTRACT In Alzheimer's disease (AD), Purkinje neurons in the cerebellum are spared, while, for instance, pyramidal neurons in the hippocampus are neuropathologically affected. Several lines of evidence suggest that the pathogenesis could be induced by the concentration-dependent polymerization of the amyloid beta-peptide (Abeta) into extracellular oligomers. The role of intracellular Abeta is not fully investigated, but recent data indicate that also this pool could be of importance. Here, we use laser capture microdissection microscopy for isolation of Purkinje neurons from AD cases and controls, and quantify the low levels of intracellular Abeta using a novel and highly sensitive ELISA. Similar to Cornu Ammonis 1 pyramidal neurons, the intracellular levels of the most toxic variant, Abeta42, as well as the Abeta42/Abeta40 ratio, were increased in Purkinje neurons from sporadic AD cases as compared to controls. However, the levels of Abeta42 as well as Abeta40 were clearly lower in Purkinje neurons than in pyramidal neurons. Based on the volume of the captured Purkinje neurons, the intraneuronal concentrations of Abeta42 were calculated to be 200 nM in sporadic AD cases and 90 nM in controls. The corresponding concentrations in pyramidal neurons from hippocampus were 3 muM and 660 nM, respectively. The Abeta40 concentration was not significantly altered in AD cases compared to controls. However, we found ten times higher concentration of Abeta40 in pyramidal neurons (10 muM) compared to Purkinje neurons (1 muM). Finally, we suggest that high concentration of intracellular Abeta42 correlates with vulnerability to AD neuropathology.
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ABSTRACT: Synaptic degeneration is one of the earliest hallmarks of Alzheimer disease. The molecular mechanism underlying this degeneration is not fully elucidated but one key player appears to be the synaptotoxic amyloid β-peptide (Aβ). The exact localization of the production of Aβ and the mechanisms whereby Aβ is released remain elusive. We have earlier shown that Aβ can be produced in crude synaptic vesicles and it has been reported that increased synaptic activity results in increased secreted but decreased intracellular Aβ levels. Therefore, we considered whether Aβ could be produced in synaptic vesicles and/or released through the same mechanisms as neurotransmitters in synaptic vesicle exocytosis. Small amounts of Aβ were found to be produced in pure synaptic vesicle preparations. We also studied the release of glutamate and Aβ from rat cortical nerve terminals (synaptosomes). We found that large amounts of Aβ were secreted from non-stimulated synaptosomes, from which glutamate was not released. On the contrary, we could not detect any differences in Aβ release between non-stimulated synaptosomes and synaptosomes stimulated with KCl or 4-aminopyridine, whereas glutamate release was readily inducible in this system. To conclude, our results indicate that the major release mechanism of Aβ from isolated nerve terminals differs from the synaptic release of glutamate and that the activity-dependent increase of secreted Aβ, reported by several groups using intact cells, is likely dependent on post-synaptic events, trafficking and/or protein synthesis mechanisms.Neuroscience Letters 03/2014; 566. DOI:10.1016/j.neulet.2014.02.050
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ABSTRACT: The aberrant accumulation of aggregated beta-amyloid peptides (Abeta) as plaques is a hallmark of Alzheimer's disease (AD) neuropathology and reduction of Abeta has become a leading direction of emerging experimental therapies for the disease. The mechanism(s) whereby Abeta is involved in the pathophysiology of the disease remain(s) poorly understood. Initially fibrils, and subsequently oligomers of extracellular Abeta have been viewed as the most important pathogenic form of Abeta in AD. More recently, the intraneuronal accumulation of Abeta has been described in the brain, although technical considerations and its relevance in AD have made this a controversial topic. Here, we review the emerging evidence linking intraneuronal Abeta accumulation to the development of synaptic pathology and plaques in AD, and discuss the implications of intraneuronal beta-amyloid for AD pathology, biology, diagnosis and therapy.Acta Neuropathologica 03/2010; 119(5):523-41. DOI:10.1007/s00401-010-0679-9
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ABSTRACT: Alzheimer's disease is the most common cause of dementia in elderly people. Research into Alzheimer's disease therapy has been at least partly successful in terms of developing symptomatic treatments, but has also had several failures in terms of developing disease-modifying therapies. These successes and failures have led to debate about the potential deficiencies in our understanding of the pathogenesis of Alzheimer's disease and potential pitfalls in diagnosis, choice of therapeutic targets, development of drug candidates, and design of clinical trials. Many clinical and experimental studies are ongoing, but we need to acknowledge that a single cure for Alzheimer's disease is unlikely to be found and that the approach to drug development for this disorder needs to be reconsidered. Preclinical research is constantly providing us with new information on pieces of the complex Alzheimer's disease puzzle, and an analysis of this information might reveal patterns of pharmacological interactions instead of single potential drug targets. Several promising randomised controlled trials are ongoing, and the increased collaboration between pharmaceutical companies, basic researchers, and clinical researchers has the potential to bring us closer to developing an optimum pharmaceutical approach for the treatment of Alzheimer's disease.The Lancet Neurology 07/2010; 9(7):702-16. DOI:10.1016/S1474-4422(10)70119-8