Nanosecond pulsed electric field thresholds for nanopore formation in neural cells

The persistent influx of ions through nanopores created upon cellular exposure to nanosecond pulse electric fields (nsPEF) could be used to modulate neuronal function. One ion, calcium (Ca2+), is important to action potential firing and regulates many ion channels. However, uncontrolled hyper-excitability of neurons leads to Ca2+ overload and neurodegeneration. Thus, to prevent unintended consequences of nsPEF-induced neural stimulation, knowledge of optimum exposure parameters is required. We determined the relationship between nsPEF exposure parameters (pulse width and amplitude) and nanopore formation in two cell types: rodent neuroblastoma (NG108) and mouse primary hippocampal neurons (PHN). We identified thresholds for nanoporation using Annexin V and FM1-43, to detect changes in membrane asymmetry, and through Ca2+ influx using Calcium Green. The ED50 for a single 600 ns pulse, necessary to cause uptake of extracellular Ca2+, was 1.76 kV/cm for NG108 and 0.84 kV/cm for PHN. At 16.2 kV/cm, the ED50 for pulse width was 95 ns for both cell lines. Cadmium, a nonspecific Ca2+ channel blocker, failed to prevent Ca2+ uptake suggesting that observed influx is likely due to nanoporation. These data demonstrate that moderate amplitude single nsPEF exposures result in rapid Ca2+ influx that may be capable of controllably modulating neurological function. (C) The Authors. Published by SPIE under a Creative Commons Attribution 3.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.