Electric Patch-Clamp Probing and Computational Studies of Lipid Bilayer Structural Fluctuations Induced by Methylamphetamine on a Neuronal Cell Membrane

Investigating how the central nervous system stimulant methylamphetamine (METH) disrupts neuronal cell membranes is crucial, as lack of knowledge of the molecular-level disruptions-such as complex solvation dynamics and alterations in the lipid bilayer structure and texture-caused by the interaction of these lipophilic drugs with neuronal membranes. Even though detection of these drug-induced disruptions of the neuronal membrane is technically challenging, we were able to measure the electrical current fluctuation changes of the disrupted HT22 neuronal cell membrane upon interacting with METH using whole-cell patch-clamp combined differential interference contrast microscopy. Furthermore, we carried out molecular dynamics (MD) simulation studies to investigate the permeation of METH into the lipid bilayer and to study the interaction between lipids and METH. We observed that the fluidity and permeability fluctuations of the HT22 cell membrane have increased in the presence of METH molecules in a concentration-dependent manner resulting in higher electric leaking states for higher METH concentrations used, which have more electric conductance compared to the nonleaking states. Analyzing the autocorrelation function fittings for the leaking vs nonleaking electric current activities, we were able to characterize the conformational dynamics and randomness of the cell membrane permeability fluctuations to study the recovery rate of the bilayer from the temporarily perturbed state. MD simulations with quantitative free energy analysis revealed that the METH molecules easily permeate into the lipid bilayer showing METH-cluster formations and accumulations inside the lipid bilayer close to the lipid headgroup's level region and showing subsequent interactions with lipid head groups and dragging bulk water molecules into the bilayer. Also, we observed that the formed METH clusters for higher METH concentrations do not penetrate the bilayer as a cluster. Instead, interestingly single METH molecule penetration per time into the bilayer from the METH cluster was observed from our MD results, illustrating the leaking vs nonleaking behaviors detected for patch-clamp experiments. Applying our combined new approach, we obtained results with real-time permeability changes and computational observations of the cell membrane due to METH-induced disruptions, which provides comprehensive knowledge important to studying the drug disturbance of crucial cellular functions such as solvation dynamics fluctuations and the molecular behavior of METH in the water lipid interface.