Exploring the conformational changes induced by nanosecond pulsed electric fields on the voltage sensing domain of a Ca²⁺ channel

Nanosecond Pulsed Electric Field (nsPEF or Nano Pulsed Stimulation, NPS) is a technology that delivers a series of pulses of high-voltage electric fields during a short period of time, in the order of nanoseconds. The main consequence of nsPEF upon cells is the formation of nanopores, which is followed by the gating of ionic channels. Literature is conclusive in that the physiological mechanisms governing ion channel gating occur in the order of milliseconds. Hence, understanding how these channels can be activated by a nsPEF would be an important step in order to conciliate fundamental biophysical knowledge with improved nsPEF applications. To get insights on both the kinetics and thermodynamics of ion channel gating induced by nsPEF, in this work, we simulated the Voltage Sensing Domain (VSD) of a voltage-gated Ca²⁺ channel, inserted in phospholipidic membranes with different concentrations of cholesterol. We studied the conformational changes of the VSD under a nsPEF mimicked by the application of a continuous electric field lasting 50 ns with different intensities as an approach to reveal novel mechanisms leading to ion channel gating in such short timescales. Our results show that using a membrane with high cholesterol content, under an nsPEF of 50 ns and E^⃗ = 0.2 V/nm, the VSD undergoes major conformational changes. As a whole, our work supports the notion that membrane composition may act as an allosteric regulator, specifically cholesterol content, which is fundamental for the response of the VSD to an external electric field. Moreover, changes on the VSD structure suggest that the gating of voltage-gated Ca²⁺ channels by a nsPEF may be due to major conformational changes elicited in response to the external electric field. Finally, the VSD/cholesterol-bilayer under an nsPEF of 50 ns and ⃗E = 0.2 V/nm elicits a pore formation across the VSD suggesting a new non-reported effect of nsPEF into cells, which can be called a “protein mediated electroporation”.