Size of the pores created by an electric pulse: Microsecond vs millisecond pulses

Abstract

Here, the sizes of the pores created by square-wave electric pulses with the duration of 100 μs and 2 ms are compared for pulses with the amplitudes close to the threshold of electroporation. Experiments were carried out with three types of cells: mouse hepatoma MH-22A cells, Chinese hamster ovary (CHO) cells, and human erythrocytes. In the case of a short pulse (square-wave with the duration of 100 μs or exponential with the time constant of 22 μs), in the large portion (30-60%) of electroporated (permeable to potassium ions) cells, an electric pulse created only the pores, which were smaller than the molecule of bleomycin (molecular mass of 1450 Da, r≈0.8 nm) or sucrose (molecular mass of 342.3 Da, radius-0.44-0.52 nm). In the case of a long 2-ms duration pulse, in almost all cells, which were electroporated, there were the pores larger than the molecules of bleomycin and/or sucrose. Kinetics of pore resealing depended on the pulse duration and was faster after the shorter pulse. After a short 100-μs duration pulse, the disappearance of the pores permeable to bleomycin was completed after 6-7 min at 24-26°C, while after a long 2-ms duration pulse, this process was slower and lasted 15-20 min. Thus, it can be concluded that a short 100-μs duration pulse created smaller pores than the longer 2-ms duration pulse. This could be attributed to the time inadequacy for pores to grow and expand during the pulse, in the case of short pulses.

Imaging the dynamics of individual electropores

“…This value will be an underestimate as the field experienced in the region of the pore will be reduced upon its formation (32,33); however, it supports previous suggestions that the smallest electrically conductive pores are those able to accommodate at least a single file of ions. This value is also on the order of the smallest pores found in experimental and theoretical studies (19,27,(34)(35)(36).…”

“…4C shows a median-averaged image from such an experiment, overlaid with trajectories of electropores diffusing in the membrane (see also Fig. S6 and (27,38,39), whereas closure is typically in the nanosecondto-microsecond range in simulations (19,33). Within the time resolution of our experiments (16 ms), we observe pores that close immediately upon removal of the applied potential.…”

“…For example, GUVs have been used to show transport across a membrane can be achieved by an electroporation pulse only 10 ns in duration (26). However, many of these systems typically examine small numbers of large, microscopic pores, much larger than the nanoscopic pores observed in cell systems (20-120 nm) (20), predicted by molecular dynamics simulations (17,19) and implied by cellular uptake assays (27).…”

Nanosecond pulsed electric field thresholds for nanopore formation in neural cells

“…Nanopore formation, in contrast to the larger pores formed by longer pulses (electroporation), are less likely to pass large ions such as propidium, but freely allow passage of small ions. 7,8 Various studies using diverse techniques such as electrophysiology, [9][10][11] fluorescence microscopy, 7,11 and direct ion measurement in bulk solution for longer pulse widths 12 have supported the hypothesis that such "nanopores" exist. Interestingly, when exposed cells are under a whole cell patch clamp, nanopore activity appears to have unique electrical characteristics.…”

Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery

“…Thus, in traditional electroporation, both permeabilization and delivery are controlled in large part by the parameters of the applied field, even though different physical mechanisms are involved for both. These electrical parameters, including field strength [37,[43][44][45][46][47][48][49], pulse duration [35,44,47,50], number of pulses [34,51], and pulse shape [52][53][54][55][56], have been studied extensively over the past three decades.…”