Non-monotonic response of cell membrane permeabilization under nanosecond pulsed electric field from 1 Hz to 2 MHz: A numerical and experimental study

It is generally regarded that cell membrane permeabilization increases monotonically with the repetition rate of nanosecond pulsed electric field (nsPEF). However, our experiments showed that this was not always the case. In the high-conductivity culture medium (1 S/m), cell permeabilization showed increase (1–100 Hz), plateau (100 Hz–20 kHz), sharp rise (>100 kHz), and saturation (1–2 MHz). In the low-conductivity sucrose buffer (0.1 S/m), cell permeabilization showed increase (1–10 Hz), little change (10 Hz–1 kHz), decrease (10–100 kHz), and then the same sharp rise (100 kHz–2 MHz). An established classical single-cell electroporation model showed that the second-pulse transmembrane-voltage peak was jointly governed by residual-voltage superposition and suppression associated with electroporation-induced membrane-conductance increase; accordingly, the peak changed non-monotonically with increasing repetition rate. Finally, compared with the weak membrane damage induced by 1 Hz nsPEF, 1 MHz nsPEF produced pronounced immediate membrane disruption and sustained post-pulse membrane permeabilization, consistent with the model-predicted MHz-range enhancement. These results indicate that nsPEF repetition rate should not be regarded as a universally monotonic enhancement factor for cell membrane permeabilization.