Asymmetrical bipolar nanosecond electric pulse widths modify bipolar cancellation

Chris M Valdez,Caleb C Roth,Bennett L Ibey,Graham A Throckmorton,Erick K Moen,Ronald A Barnes

doi:10.1038/s41598-017-16142-6

Abstract

A bipolar (BP) nanosecond electric pulse (nsEP) exposure generates reduced calcium influx compared to a unipolar (UP) nsEP. This attenuated physiological response from a BP nsEP exposure is termed “bipolar cancellation” (BPC). The predominant BP nsEP parameters that induce BPC consist of a positive polarity (↑) front pulse followed by the delivery of a negative polarity (↓) back pulse of equal voltage and width; thereby the duration is twice a UP nsEP exposure. We tested these BPC parameters, and discovered that a BP nsEP with symmetrical pulse widths is not required to generate BPC. For example, our data revealed the physiological response initiated by a ↑900 nsEP exposure can be cancelled by a second pulse that is a third of its duration. However, we observed a complete loss of BPC from a ↑300 nsEP followed by a ↓900 nsEP exposure. Spatiotemporal analysis revealed these asymmetrical BP nsEP exposures generate distinct local YO-PRO®-1 uptake patterns across the plasma membrane. From these findings, we generated a conceptual model that suggests BPC is a phenomenon balanced by localized charging and discharging events across the membrane.

Highlights

® exposures generate distinct localYO-PRO -1 uptake patterns across the plasma membrane
We evaluated if the physiological response initiated by a BP nanosecond electric pulse (nsEP) that exhibits asymmetry among individual pulse widths would cancel the physiological effect initiated by the first pulse
We demonstrated that the negative impact on membrane integrity triggered by a ↑300 nsEP exposure can be cancelled by a ↑300↓300 nsEP

Summary

Introduction

® exposures generate distinct localYO-PRO -1 uptake patterns across the plasma membrane. A nsEP exposure can generate membrane perturbations that facilitate entry of small ions into the cell causing membrane depolarization, intracellular second messenger signaling, and apoptotic pathway activation2–4 These various cellular effects can be tuned by modulating nsEP intensity (duration, amplitude, and number of pulses) highlighting the mechanistic depth and breadth of its impact on biological systems. Cells exposed to a BP nsEP exhibit reduced calcium and propidium uptake as compared to the same total duration UP exposures, suggesting the BP nsEP generates less membrane perturbation. The delivery of asymmetrical BP pulses is specific to microsecond pulse exposures; the data demonstrates that asymmetry across the individual BP pulse widths can generate lethal thresholds from irreversible electroporation that are similar to high frequency unipolar applications8 These observations extend the possibility that asymmetry across BP nsEP widths may modify our current understanding of BPC.

Methods

Results

Conclusion