Asymmetrical bipolar nanosecond electric pulse widths modify bipolar cancellation

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.

The interphase interval within a bipolar nanosecond electric pulse modulates bipolar cancellation

“…To test if this reversal is partially due to symmetry among the BP nsEPs, we exposed CHO‐K1 cells to a ↑900↓300 nsEP with a lengthened interphase interval. In comparison to a ↑600 UP nsEP exposure, this BP nsEP eliminates pulse symmetry, maintains a doubling of total pulse duration, and was previously described to yield BPC of YP‐1 uptake by 55% [Valdez et al, ]. When we separated the ↑900↓300 nsEP pulse by 10 μs and 10 ms, we observed YP‐1 uptake was differentially reduced by 41% and 14%, respectively (Fig.…”

“…We previously observed BPC of YP‐1 uptake from the delivery of ↑300↓300, ↑600↓600, and ↑900↓900 nsEP exposures (Fig. ) [Valdez et al, ]. In this report, we increased the interphase interval between these symmetrical BP nsEP waveforms, and were able to alter their BPC effect on YP‐1 uptake (Fig.…”

“…From the ↑900↓300 nsEP exposure with a 10 ms and 10 μs interphase interval, the trajectory of YP‐1 deviated at 3 s post exposure in comparison to a ↑600 UP nsEP. From our previous work, we demonstrated that YP‐1 uptake generated from a ↑900 UP nsEP can be cancelled by an opposite polarity ↓300 nsEP (↑900 vs. ↑900↓300) [Valdez et al, ]. To extend on this finding we applied a 10 μs, 10 ms, or 1 s interphase interval between the ↑900↓300 asymmetrical BP nsEP, and evaluated this effect on YP‐1 uptake (Fig.…”

“…A region of interest (ROI) was coded from an ImageJ macro tool that encircled the exposed cell. Data archiving procedures were previously described [Valdez et al, ]. In GraphPad (Prism, La Jolla, CA), YP‐1 uptake from each cell was calculated by subtracting the average baseline YP‐1 fluorescence value from each pre‐ and post‐exposure data point.…”

“…On the cellular level, a BP nsEP exposure generates less Propidium Iodide (PI) uptake compared to a UP nsEP of the same duration [Ibey et al, ]. Furthermore, the extent of Ca 2+ and YO‐PRO‐1 (YP‐1) uptake from a UP nsEP can be cancelled by the immediate delivery of a second pulse, opposite in polarity and equivalent in duration [Pakhomov et al, ; Valdez et al, ; Gianulis et al, ].…”