Recent studies suggest that nanosecond pulsed electric fields (nsPEF) may influence ion channel behavior by inducing conformational changes. While several reports have highlighted the ability of nsPEFs to affect cellular activity the underlying mechanisms, particularly whether they involve direct modulation of channel gating or indirect effects, such as membrane permeabilization, remain under debate. In this context, ion channels are of particular interest due to their central roles in processes, such as pain perception, excitability, and cell signaling. Clarifying how nsPEFs interact with these proteins could open new perspectives for bioelectrical control strategies. Our project aims to experimentally explore the impact of nsPEF on ion channels using genetically encoded bioluminescence resonance energy transfer (BRET)-based biosensors, which enable non-invasive, real-time monitoring of molecular events in living cells. To monitor ion channel behavior in real time, we generated BRET-based biosensors by fusing donor and acceptor pairs (mNeonGreen/nLuc or rGFP/rLuc) to selected ion channels of interest, including members of the TRP, VGIC, and TREK families. These biosensors allow dynamic tracking of conformational changes or calcium fluxes at the plasma membrane or in intracellular compartments. Constructs were transiently transfected in HEK293T cells cultured either as monolayers (2D) or spheroids (3D) and first validated using chemical stimulation. In parallel, a dedicated platform is currently under development to synchronize nsPEF delivery with BRET signal acquisition. The system includes a delivery system and a fiber-coupled spectrometer for live-cell photon detection. With validated biosensors and an operational nsPEF delivery platform, our study provides a solid basis to explore how ion channels respond in live cells. This will allow us to test whether certain pulse parameters can lead to observable changes in their activity.
Ferber et al. (Sun,) studied this question.