Abstract:The toxicity of the nonaggregated amyloid ␤-peptide (1-40) [A␤(1-40)] on the viability of rat cortical neurons in primary culture was investigated. We demonstrated that low concentrations of A␤ peptide, in a nonfibrillar form, induced a time-and dose-dependent apoptotic cell death, including DNA condensation and fragmentation. We compared the neurotoxicity of the A␤(1-40) peptide with those of several A␤-peptide domains, comprising the membrane-destabilizing C-terminal domain of A␤ peptide (e.g., amino acids 29 -40 and 29 -42). These peptides reproduced the effects of the (1-40) peptide, whereas mutant nonfusogenic A␤ peptides and the central region of the A␤ peptide (e.g., amino acids 13-28) had no effect on cell viability. We further demonstrated that the neurotoxicity of the nonaggregated A␤ peptide paralleled a rapid and stable interaction between the A␤ peptide and the plasma membrane of neurons, preceding apoptosis and DNA fragmentation. By contrast, the peptide in a fibrillar form induced a rapid and dramatic neuronal death mainly through a necrotic pathway, under our conditions. Taken together, our results suggest that A␤ induces neuronal cell death by either apoptosis and necrosis and that an interaction between the nonfibrillar C-terminal domain of the A␤ peptide and the plasma membrane of cortical neurons might represent an early event in a cascade leading to neurodegeneration. Key Words: Alzheimer's disease -Amyloid ␤-peptideApoptosis-Fusogenic peptides-Neurotoxicity-Cortical primary neurons.

In the present study, we have determined the nature and the kinetics of the cellular events triggered by the exposure of cells to non-fibrillar amyloid-␤ peptide (A␤). When cortical neurons were treated with low concentrations of soluble A␤ (1-40), an early reactive oxygen species (ROS)-dependent cytoskeleton disruption precedes caspase activation. Indeed, caspase activation and neuronal cell death were prevented by the microtubulestabilizing drug taxol. A perturbation of the microtubule network was noticeable after being exposed to A␤ for 1 h, as revealed by electron microscopy and immunocytochemistry. Microtubule disruption and neuronal cell death induced by A␤ were inhibited in the presence of antioxidant molecules, such as probucol. These data highlight the critical role of ROS production in A␤-mediated cytoskeleton disruption and neuronal cell death. Finally, using FRAP (fluorescence recovery after photo bleaching) analysis, we observed a time-dependent biphasic modification of plasma membrane fluidity, as early as microtubule disorganization. Interestingly, molecules that inhibited neurotubule perturbation and cell death did not affect the membrane destabilizing properties of A␤, suggesting that the lipid phase of the plasma membrane might represent the earliest target for A␤. Altogether our results convey the idea that upon interaction with the plasma membrane, the non-fibrillar A␤ induces a rapid ROS-dependent disorganization of the cytoskeleton, which results in apoptosis.A common feature of Alzheimer's disease (AD), 1 the most common form of dementia, is the accumulation and the aggregation of the amyloid-␤ peptide (A␤), a 39-to 43-amino acid peptide derived from the proteolytic cleavage of the amyloid precursor protein (1, 2). Although A␤ represents a key factor in AD (3), the nature of the toxic form of A␤ early involved in AD pathology remains unclear. Whether it is the fibrillar or the non-fibrillar peptides that are the more deleterious remains a controversial issue (4). The amyloid cascade hypothesis causally links AD clinico-pathological process and neuronal cell death to the aggregation and deposition of A␤ (5-7). However, this hypothesis has been challenged by recent evidences indicating that the non-fibrillar A␤ also plays a major role in AD (8, 9). A recent elegant study has demonstrated that the fibrils from AD brain are composed of amyloid peptide moieties arranged at right angles to the backbone of the amyloid P protein wrapped in glycosaminoglycans (10). Thus, the fibrils are not simply made of chains of self-aggregated A␤ and do not comprise long chains of multimeric A␤, similar to those used to evaluate the neurotoxicity of the fibrillar A␤ in vitro and in vivo. Moreover, the synaptic loss in AD brain has been correlated with the soluble pool of A␤ peptides rather than the fibrillar one, implying that the non-fibrillar A␤ may be a crucial pathological factor in AD (11-13). Several studies, based on the use of transgenic mice, have demonstrated that neurodegeneration and specific spatia...

Despite an exponential production of data, Alzheimer's disease (AD) remains an enigma. Unresolved questions persist in the face of the heterogeneity of this neuropathology. Recent progress in understanding mechanisms for AD results from the study of amyloid precursor protein (APP) metabolism and the involvement of senile plaque-associated proteins. In addition to the amyloid cascade hypothesis, alternative schemes emerge, in which the amyloid peptide is not the primary effector of the disease. Perturbations of vesicular trafficking, the cytoskeletal network, and membrane cholesterol distribution could be central events. Furthermore, since the physiological role of APP, presenilins, and apolipoprotein E in the central nervous system are not completely understood, their involvement in AD etiology remains speculative. New actors have to be found to try to explain sporadic cases and non-elucidated familial cases.

Although the genetic link between the 14 allele of apolipoprotein E (apoE) and Alzheimer's disease (AD) is well established, the apoE isoform-speci®c activity underlying this correlation remains unclear. We have recently characterized the interaction of the soluble the amyloid-b peptide (Ab) with model membrane and demonstrated that non-®brillar Ab peptide, including N-terminal truncated forms of Ab, induced apoptotic cell death in primary rat cortical neurones in vitro. To further investigate the potential interaction between apoE and Ab in the pathogenesis of AD, we have determined the effect of apoE isoforms on the neurotoxicity of non-®brillar Ab peptides. We demonstrate here that the apoE2 and E3 isoforms protect cortical neurones against apoptotic cell death induced by a non-®brillar form of the Ab 1240 , Ab 12242 , Ab 29240 and Ab 29242 peptides, whereas apoE4 had no effect. This effect involves the formation of stable complexes between apoE and the C-terminal domain (e.g. amino acids 29±40) of Ab 1240 . Interestingly, apoE had no effect on the toxicity induced by aggregated Ab peptides, suggesting a lack of interaction between apoE and amyloid ®brils. Our results provide evidence that interaction with the C-terminal domain of Ab, apoE2 and E3, but not apoE4, inhibits the interactions of the non-®brillar Ab peptide with the plasma membrane of neurones, Ab peptide aggregation and subsequent neurotoxicity.