The aggregation of proteins into oligomers and amyloid fibrils is characteristic of several neurodegenerative diseases, including Parkinson disease (PD). In PD, the process of aggregation of α-synuclein (α-syn) from monomers, via oligomeric intermediates, into amyloid fibrils is considered the disease-causative toxic mechanism. We developed α-syn mutants that promote oligomer or fibril formation and tested the toxicity of these mutants by using a rat lentivirus system to investigate loss of dopaminergic neurons in the substantia nigra. The most severe dopaminergic loss in the substantia nigra is observed in animals with the α-syn variants that form oligomers (i.e., E57K and E35K), whereas the α-syn variants that form fibrils very quickly are less toxic. We show that α-syn oligomers are toxic in vivo and that α-syn oligomers might interact with and potentially disrupt membranes. P arkinson disease (PD) is the most common movement disorder, currently affecting approximately 2% of the population older than age 60 y. Prominent neuropathological hallmarks of PD are the loss of dopaminergic neurons in the substantia nigra (SN) region of the midbrain (1) and the presence of α-syn-containing intracellular inclusions: Lewy bodies (LBs) and Lewy neurites (2). α-Syn, a 140-aa protein physiologically found in presynaptic terminals of neurons, is the major fibrillar protein in LBs and Lewy neurites in sporadic and inherited PD. Moreover, point mutations (A53T, A30P, E46K) and gene multiplications of human WT (hWT) α-syn are related to rare familial autosomal-dominant forms of early-onset PD (3-6), suggesting that increased gene dosage and aberrant protein structure may accelerate disease onset and progression.Recent reports indicate that the accumulation of α-syn can result in the formation of intermediate-state oligomers, and oligomers of different shapes and sizes have been described (7-10). These oligomers interact with lipids, disrupt membranes (7,8), and cause cell death in vitro (10, 11) and in nonmammalian models, such as Caenorhabditis elegans and Drosophila melanogaster (12). However, we are aware of no previous direct in vivo demonstration of the toxicity of α-syn oligomers in mammals.We aim to establish a model that allows specific testing of the effects of α-syn oligomerization in vitro and in vivo. To elucidate the causal structure-toxicity relationship of these oligomeric protein assemblies in a mammalian system, we designed "conformation-trapped" mutants based on structural modeling of α-syn fibrils (13, 14). Structurally, amyloid fibrils of α-syn are composed of cross-β-sheets (15). Residues from approximately 30 to 110 of α-syn form the core of the fibrils, whereas the approximately 30 N-terminal residues are heterogeneous and the approximately 30 C-terminal residues are flexible (13,14,16,17). Based on our structural model, recently developed from NMR data, the core of α-syn fibrils comprises five β-strands reminiscent of a five-layered "β-sandwich" (14). Several loops adjacent to and between the strands ar...

The cognitive impairment in patients with Alzheimer's disease is closely associated with synaptic loss in the neocortex and limbic system. Although the neurotoxic effects of aggregated amyloid-b oligomers in Alzheimer's disease have been studied extensively in experimental models, less is known about the characteristics of these aggregates across the spectrum of Alzheimer's disease. In this study, postmortem frontal cortex samples from controls and patients with Alzheimer's disease were fractionated and analyzed for levels of oligomers and synaptic proteins. We found that the levels of oligomers correlated with the severity of cognitive impairment (blessed information-memory-concentration score and mini-mental state examination) and with the loss of synaptic markers. Reduced levels of the synaptic vesicle protein, vesicle-associated membrane protein-2, and the postsynaptic protein, postsynaptic density-95, correlated with the levels of oligomers in the various fractions analyzed. The strongest associations were found with amyloid-b dimers and pentamers. Co-immunoprecipitation and doublelabeling experiments supported the possibility that amyloid-b and postsynaptic density-95 interact at synaptic sites. Similarly, in transgenic mice expressing high levels of neuronal amyloid precursor protein, amyloid-b co-immunoprecipitated with postsynaptic density-95. This was accompanied by a decrease in the levels of the postsynaptic proteins Shank1 and Shank3 in patients with Alzheimer's disease and in the brains of amyloid precursor protein transgenic mice. In conclusion, this study suggests that the presence of a subpopulation of amyloid-b oligomers in the brains of patients with Alzheimer's disease might be related to alterations in selected synaptic proteins and cognitive impairment.

During aging and in the progression of Alzheimer's disease (AD), synaptic plasticity and neuronal integrity are disturbed. In addition to the alterations in plasticity in mature neurons, the neurodegenerative process in AD has been shown to be accompanied by alterations in neurogenesis. Members of the bone morphogenetic protein (BMP) family of growth factors have been implicated as important regulators of neurogenesis and neuronal cell fate determination during development; however, their role in adult neurogenesis and in AD is less clear. We show here by qRT-PCR analysis that BMP6 mRNA levels were significantly increased in the hippocampus of human patients with AD and in APP transgenic mice compared to controls. Immunoblot and immunohistochemical analyses confirmed that BMP6 protein levels were increased in human AD brains and APP transgenic mouse brains compared to controls and accumulated around hippocampal plaques. The increased levels of BMP6 were accompanied by defects in hippocampal neurogenesis in AD patients and APP transgenic mice. In support of a role for BMP6 in defective neurogenesis in AD, we show in an in vitro model of adult neurogenesis that treatment with amyloid-␤ 1-42 protein (A␤) resulted in increased expression of BMP6, and that exposure to recombinant BMP6 resulted in reduced proliferation with no toxic effects. Together, these results suggest that A␤-associated increases in BMP6 expression in AD may have deleterious effects on neurogenesis in the hippocampus, and therapeutic approaches could focus on normalization of BMP6 levels to protect against AD-related neurogenic deficits.

Sleep disorders are observed in Parkinson’s disease, Dementia with Lewy Bodies and Alzheimer’s disease, however the underlying mechanisms are unclear. Reduced hypocretin (orexin) levels are reported in Parkinson’s disease and sleep disorders including narcolepsy, however levels in Dementia with Lewy Bodies and Alzheimer’s disease and their relationship to sleep disturbances in these disorders remain undetermined. We examined hypocretin levels in Dementia with Lewy Bodies and Alzheimer’s disease cases and correlated these with sleep habits and clinical characteristics. Whilst limited hypocretin alterations were observed in Alzheimer’s disease, we demonstrate reduced neocortical hypocretin-immunoreactivity in Dementia with Lewy Bodies patients correlating with hypersomnolence and alpha-synuclein levels. These results suggest the involvement of hypocretin in sleep disorders in Dementia with Lewy Bodies.

Neurodegenerative disorders such as Alzheimer’s disease (AD) are characterized by the loss of neurotrophic factors and experimental therapeutical approaches to AD have investigated the efficacy of replacing or augmenting neurotrophic factor activity. Cerebrolysin™, a peptide mixture with neurotrophic-like effects, has been shown to improve cognition in patients with AD and to reduce synaptic and behavioral deficits in transgenic (tg) mice over expressing the amyloid precursor protein (APP). However it is unclear how long lasting the beneficial effects of Cerebrolysin™ are and whether or not behavioral and neuropathological alterations will reappear following treatment interruption. The objective of the present study was to investigate the consequences of interrupting Cerebrolysin™ treatment (“wash-out” effect) 3 and 6 months after the completion of a 3-month treatment period in APP tg mice. We demonstrate that in APP tg mice, Cerebrolysin™-induced amelioration of memory deficits in the water maze and reduction of neurodegenerative pathology persist for 3 months after treatment interruption, however these effects dissipate 6 months following treatment termination. Immunohistochemical analysis demonstrated that the decrease in neocortical and hippocampal amyloid plaque load observed in Cerebrolysin™ treated APP tg mice immediately following treatment was no longer apparent at 3 months after treatment interruption indicating that the beneficial effects of Cerebrolysin™ at this time point were independent of its effect on amyloid-beta deposition. In conclusion the results demonstrate that effects of Cerebrolysin™ persist for a significant period of time following treatment termination and suggest that this prolonged effect may involve the neurotrophic factor-like activity of Cerebrolysin™.