Abstract Non-specific disruption of neuronal cell membrane induced by amyloidogenic aggregation of b-amyloid (Ab) peptides is considered a viable mechanism in the pathology of Alzheimer’s disease (AD). Understanding the structural basis of key molecular interactions that are responsible for the membrane disruption remains challenging, mainly due to the heterogeneity nature of Ab-membrane systems and the low abundance of key intermediate states. High-resolution solid-state nuclear magnetic resonance (ssNMR) spectroscopy provides the most powerful method for studying the membrane-disruptive Ab aggregation in its native-state-mimicking environments, i.e., in model liposomes and/or in the presence of neuronal cells. Our long-term goal is to understand the structural basis of key Ab-membrane interactions that are responsible for the non-specific membrane disruption in neurons. During the last funding period, my group has demonstrated the generality of two distinct membrane-disruptive processes in model liposomes: local membrane leaking induced by on-fibrillation-pathway Ab intermediates and membrane fragmentation induced by off- fibrillation-pathway large Ab oligomers. We have also established ssNMR approaches to investigate the structural features and molecular interactions at the intermediate states of individual processes. These ssNMR measurements allow systematic studies of time-dependent structural changes of Ab aggregates and key Ab-lipid interactions in the time frame of membrane disruption process, as well as the modulations of lipid dynamics due to the Ab amyloidosis. In addition, we are exploring the feasibility for extending the studies from membrane- mimicking liposomes to neuronal cells, by using a multipronged strategy combining in-cell ssNMR spectroscopy with cellular imaging/viability assays/quantitative analysis. The current renewal proposal contains three aims. Aim 1 and 2 seek for finalizing the mechanistic studies of membrane-disruptive Ab interactions in liposomes. Specific mechanistic models are proposed and will be demonstrated using extensive ssNMR experiments, including advanced sensitive-enhancing dynamic nuclear polarization (DNP) ssNMR techniques. Key membrane-interruptive intermediate states (including Ab structural features, lipid dynamics and Ab-lipid interactions) will be elucidated. In Aim 3, we strive to developing in-cell ssNMR approaches and quantitative cellular assays/analysis for probing the molecular basis of Ab-induced cellular membrane disruption. This involves identification of Ab-induced membrane-disruptive intermediate states with neuronal cell lines and establishment of in-cell 31P and DNP-assisted ssNMR spectroscopy, including the optimization of sample conditions and spectroscopic methods. Outcomes from the current period, including both the understanding the membrane disruptive effects and the development of ssNMR-based methods, will pave the way for future studies in primary neurons.