Quaternary Structure Defines a Large Class of Amyloid-ß Oligomers Neutralized by Sequestration.

The accumulation of amyloid-ß (Aß) as amyloid fibrils and toxic oligomers is an important step in the development of Alzheimer's disease (AD). However, there are numerous potentially toxic oligomers and little is known about their neurological effects when generated in the living brain. Here we show that Aß oligomers can be assigned to one of at least two classes (type 1 and type 2) based on their temporal, spatial, and structural relationships to amyloid fibrils. The type 2 oligomers are related to amyloid fibrils and represent the majority of oligomers generated in vivo, but they remain confined to the vicinity of amyloid plaques and do not impair cognition at levels relevant to AD. Type 1 oligomers are unrelated to amyloid fibrils and may have greater potential to cause global neural dysfunction in AD because they are dispersed. These results refine our understanding of the pathogenicity of Aß oligomers in vivo.

A monoclonal antibody against synthetic Aß dimer assemblies neutralizes brain-derived synaptic plasticity-disrupting Aß.

Diverse lines of evidence indicate that pre-fibrillar, diffusible assemblies of the amyloid ß-protein (Aß) play an important role in Alzheimer's disease pathogenesis. Although the precise molecular identity of these soluble toxins remains unsettled, recent experiments suggest that sodium dodecyl sulfate (SDS)-stable Aß dimers may be the basic building blocks of Alzheimer's disease-associated synaptotoxic assemblies and as such present an attractive target for therapeutic intervention. In the absence of sufficient amounts of highly pure cerebral Aß dimers, we have used synthetic disulfide cross-linked dimers (free of Aß monomer or fibrils) to generate conformation-specific monoclonal antibodies. These dimers aggregate to form kinetically trapped protofibrils, but do not readily form fibrils. We identified two antibodies, 3C6 and 4B5, which preferentially bind assemblies formed from covalent Aß dimers, but do not bind to Aß monomer, amyloid precursor protein, or aggregates formed by other amyloidogenic proteins. Monoclonal antibody 3C6, but not an IgM isotype-matched control antibody, ameliorated the plasticity-disrupting effects of Aß extracted from the aqueous phase of Alzheimer's disease brain, thus suggesting that 3C6 targets pathogenically relevant Aß assemblies. These data prove the usefulness of covalent dimers and their assemblies as immunogens and recommend further investigation of the therapeutic and diagnostic utility of monoclonal antibodies raised to such assemblies.

Protection against Aß-mediated rapid disruption of synaptic plasticity and memory by memantine.

Soluble amyloid-ß protein (Aß) may cause cognitive impairment in Alzheimer's disease in the absence of significant neurodegeneration. Here, the ability of the NMDA receptor (NMDAR) antagonist memantine to prevent synthetic Aß-mediated rapid functional deficits in learned behavior and synaptic plasticity was assessed in the rat. In vitro, pretreatment with a clinically relevant, NMDAR blocking concentration of memantine partially inhibited the induction of long-term potentiation (LTP) in the dentate gyrus and prevented further inhibition caused by exposure to Aß(1-42). Whereas systemic injection with memantine alone inhibited LTP in the CA1 area in vivo, a subthreshold dose partially abrogated the inhibition of LTP by intracerebroventricular soluble Aß(1-42). Similarly, systemic treatment with memantine alone impaired performance of an operant learning task and a subthreshold dose prevented the Aß(1-42)-mediated increase in perseveration errors. The acute protection afforded by memantine, albeit in a narrow dose range, against the rapid disruptive effects of soluble Aß(1-42) on synaptic plasticity and learned behavior strongly implicate NMDAR-dependent reversible dysfunction of synaptic mechanisms in Aß-mediated cognitive impairment.

Cognitive effects of cell-derived and synthetically derived Aß oligomers.

Soluble forms of amyloid-ß peptide (Aß) are a molecular focus in Alzheimer's disease research. Soluble Aß dimers (≈8 kDa), trimers (≈12 kDa), tetramers (≈16 kDa) and Aß*56 (≈56 kDa) have shown biological activity. These Aß molecules have been derived from diverse sources, including chemical synthesis, transfected cells, and mouse and human brain, leading to uncertainty about toxicity and potency. Herein, synthetic Aß peptide-derived oligomers, cell- and brain-derived low-n oligomers, and Aß*56, were injected intracerebroventricularly (icv) into rats assayed under the Alternating Lever Cyclic Ratio (ALCR) cognitive assay. Cognitive deficits were detected at 1.3 µM of synthetic Aß oligomers and at low nanomolar concentrations of cell-secreted Aß oligomers. Trimers, from transgenic mouse brain (Tg2576), did not cause cognitive impairment at any dose tested, whereas Aß*56 induced concentration-dependent cognitive impairment at 0.9 and 1.3µM. Thus, while multiple forms of Aß have cognition impairing activity, there are significant differences in effective concentration and potency.

Oligomers of the amyloid-beta protein disrupt working memory: confirmation with two behavioral procedures.

Converging lines of evidence suggest that oligomers of amyloid-beta play a role in the cognitive impairment characteristic of Alzheimer's disease, but only three studies have provided experimental evidence of such impairment. To provide additional information about the effects of these oligomers on memory, the present study examined the memory of groups of rats exposed to ICV injections of the culture media (CM) of Chinese Hamster Ovary cells that were (7PA2) and were not (CHO-) transfected with a human mutation of amyloid precursor protein that appears to cause early-onset Alzheimer's disease. The 7PA2 CM, which contained concentrations of soluble amyloid-beta oligomers physiologically relevant to those found in human brain, significantly disrupted working memory in rats tested in a radial-arm maze. In contrast, CHO- CM, which did not contain such oligomers, had no effect on memory. The disruptive effects of 7PA2-derived amyloid-beta oligomers, evident 2h after exposure, disappeared within a day. These findings are compared to results from 7PA2 CM tested under a complex procedure thought to measure aspects of executive function. The results confirm the disruptive effects of low-n amyloid-beta oligomers and extend them to a well-established rat model of memory.

Orally available compound prevents deficits in memory caused by the Alzheimer amyloid-beta oligomers.

OBJECTIVE: Despite progress in defining a pathogenic role for amyloid beta protein (Abeta) in Alzheimer's disease, orally bioavailable compounds that prevent its effects on hippocampal synaptic plasticity and cognitive function have not yet emerged. A particularly attractive therapeutic strategy is to selectively neutralize small, soluble Abeta oligomers that have recently been shown to mediate synaptic dysfunction. METHODS: Using electrophysiological, biochemical, and behavioral assays, we studied how scyllo-inositol (AZD-103; molecular weight, 180) neutralizes the acutely toxic effects of Abeta on synaptic function and memory recall. RESULTS: Scyllo-inositol, but not its stereoisomer, chiro-inositol, dose-dependently rescued long-term potentiation in mouse hippocampus from the inhibitory effects of soluble oligomers of cell-derived human Abeta. Cerebroventricular injection into rats of the soluble Abeta oligomers interfered with learned performance on a complex lever-pressing task, but administration of scyllo-inositol via the drinking water fully prevented oligomer-induced errors. INTERPRETATION: A small, orally available natural product penetrates into the brain in vivo to rescue the memory impairment produced by soluble Abeta oligomers through a mechanism that restores hippocampal synaptic plasticity.

Natural oligomers of the amyloid-beta protein specifically disrupt cognitive function.

A central unresolved problem in research on Alzheimer disease is the nature of the molecular entity causing dementia. Here we provide the first direct experimental evidence that a defined molecular species of the amyloid-beta protein interferes with cognitive function. Soluble oligomeric forms of amyloid-beta, including trimers and dimers, were both necessary and sufficient to disrupt learned behavior in a manner that was rapid, potent and transient; they produced impaired cognitive function without inducing permanent neurological deficits. Although beta-amyloidosis has long been hypothesized to affect cognition, the abnormally folded protein species associated with this or any other neurodegenerative disease has not previously been isolated, defined biochemically and then specifically characterized with regard to its effects on cognitive function. The biochemical isolation of discrete amyloid-beta moieties with pathophysiological properties sets the stage for a new approach to studying the molecular mechanisms of cognitive impairment in Alzheimer disease and related neurodegenerative disorders.

Our journey with neuropeptide Y: effects on ingestive behaviors and energy expenditure.

Clark and colleagues first described the robust orexigenic effects of neuropeptide Y (NPY) in 1984. Our group as well as Stanley et al. confirmed these effects in the same year. During the next 20 years, we investigated the effects of NPY on diet preferences, opioid-related feeding, distributed neural feeding networks, energy metabolism, motivation and discriminative stimulus effects. These data together with data from other laboratories indicate that NPY increases feeding, even when rats work for food; that NPY decreases energy expenditure, particularly by altering thermogenesis; and that NPY's effects on energy metabolism are mediated by a widely distributed neural network involving other neuroregulators known to be involved in energy regulation.

Naltrexone infusion inhibits the development of preference for a high-sucrose diet.

We hypothesized that the opioid antagonist naltrexone would inhibit the redevelopment of a preference for a high-sucrose diet after an abstention period from this diet. Rats that chose between a starch or sucrose diet for 10 days preferred the sucrose diet. Rats were then given access to the starch diet alone for another 10-day period. A miniosmotic pump containing saline or naltrexone was then implanted (70 microg/h; 1.7 mg/day) for approximately 10 days. During the saline infusion, 77% of the total energy came from the sucrose diet, whereas during the naltrexone infusion, 33% of the total energy came from the sucrose diet. We repeated this study in another group of rats but did not restrict the sucrose diet. In this case naltrexone failed to decrease preference for the sucrose diet. Thus naltrexone infusion inhibited redevelopment of a preference for a sucrose diet after a period of restriction to a starch diet for 10 days but had no effect on preference if both diets were present throughout the study.