Pulsed electric fields create pores in the voltage sensors of voltage-gated ion channels

Abstract

Pulsed electric fields are increasingly used in medicine to transiently increase the cell membrane permeability via electroporation, in order to deliver therapeutic molecules into the cell. One type of events that contributes to this increase in membrane permeability is the formation of pores in the membrane lipid bilayer. However, electrophysiological measurements suggest that membrane proteins are affected as well, particularly voltagegated ion channels (VGICs). The molecular mechanisms by which the electric field could affects these molecules remain unidentified. In this study we used molecular dynamics (MD) simulations to unravel the molecular events that take place in different VGICs when exposing them to electric fields mimicking electroporation conditions. We show that electric fields induce pores in the voltage-sensor domains (VSDs) of different VGICs, and that these pores form more easily in some channels than in others. We demonstrate that poration is more likely in VSDs that are more hydrated and are electrostatically more favorable for the entry of ions. We further show that pores in VSDs can expand into so-called complex pores, which become stabilized by lipid head-groups. Our results suggest that such complex pores are considerably more stable than conventional lipid pores and their formation can lead to severe unfolding of VSDs from the channel. We anticipate that such VSDs become dysfunctional and unable to respond to changes in transmembrane voltage, which is in agreement with previous electrophyiological measurements showing a decrease in the voltage-dependent transmembrane ionic currents following pulse treatment. Finally, we discuss the possibility of activation of VGICs by submicrosecond-duration pulses. Overall our study reveals a new mechanism of electroporation through membranes containing voltage-gated ion channels. Statement of SignificancePulsed electric fields are often used for treatment of excitable cells, e.g., for gene delivery into skeletal muscles, ablation of the heart muscle or brain tumors. Voltage-gated ion channels (VGICs) underlie generation and propagation of action potentials in these cells, and consequently are essential for their proper function. Our study reveals the molecular mechanisms by which pulsed electric fields directly affect VGICs and addresses questions that have been previously opened by electrophysiologists. We analyze VGICs' characteristics, which make them prone for electroporation, including hydration and electrostatic properties. This analysis is easily transferable to other membrane proteins thus opening directions for future investigations. Finally, we propose a mechanism for long-lived membrane permeability following pulse treatment, which to date remains poorly understood.

Paradoxical effects on voltage-gated Na+ conductance in adrenal chromaffin cells by twin vs single high intensity nanosecond electric pulses

“…These results suggest that the inhibition and recovery of I Na by twin pulses reflect different processes as they were completely dissociated by MβCD. At longer intervals, twin pulses led to cumulative functional disruption of Na + channels, perhaps by inflicting damage to the channel protein as suggested by the molecular dynamics study by Rems et al [46]. At shorter intervals, we propose that the rapid timing of delivery of the second pulse stimulated the trafficking of newly synthesized and readily assembled Na V subunits from the Golgi and/or endoplasmic reticulum systems toward the membrane.…”

“…Another viable hypothesis to explain the inhibitory effects of NEPs on I Na is that NEPs may directly influence the Na + channel protein. Recent molecular dynamics simulations by Rems et al [46] showed that pulsed electric fields could create alternate conductive pores near the voltage-sensitive domains (VSDs) of voltage-gated ion channels including Na + channels. These alternate pathways were shown to spontaneously evolve into more complex pores stabilized by lipid head groups and were accompanied by unfolding of the VSD from the channel.…”