P. Thomas Vernier

Nanosecond Electric Pulses Cause Mitochondrial Membrane Permeabilization in Jurkat Cells

Nanosecond, high-voltage electric pulses (nsEP) induce permeabilization of the plasma membrane and the membranes of cell organelles, leading to various responses in cells including cytochrome c release from mitochondria and caspase activation associated with apoptosis. We report here evidence for nsEP-induced permeabilization of mitochondrial membranes in living cells. Using three different methods with fluorescence indicators-rhodamine 123 (R123), tetramethyl rhodamine ethyl ester (TMRE), and cobalt-quenched calcein-we have shown that multiple nsEP (five pulses or more, 4u2009ns duration, 10u2009MV/m, 1u2009kHz repetition rate) cause an increase of the inner mitochondrial membrane permeability and an associated loss of mitochondrial membrane potential. These effects could be a consequence of nsEP permeabilization of the inner mitochondrial membrane or the activation of mitochondrial membrane permeability transition pores. Plasma membrane permeabilization (YO-PRO-1 influx) was detected in addition to mitochondrial membrane permeabilization.

Effects of high voltage nanosecond electric pulses on eukaryotic cells (in vitro): A systematic review

For this systematic review, 203 published reports on effects of electroporation using nanosecond high-voltage electric pulses (nsEP) on eukaryotic cells (human, animal, plant) in vitro were analyzed. A field synopsis summarizes current published data in the field with respect to publication year, cell types, exposure configuration, and pulse duration. Published data were analyzed for effects observed in eight main target areas (plasma membrane, intracellular, apoptosis, calcium level and distribution, survival, nucleus, mitochondria, stress) and an additional 107 detailed outcomes. We statistically analyzed effects of nsEP with respect to three pulse duration groups: A: 1-10ns, B: 11-100ns and C: 101-999ns. The analysis confirmed that the plasma membrane is more affected with longer pulses than with short pulses, seen best in uptake of dye molecules after applying single pulses. Additionally, we have reviewed measurements of nsEP and evaluations of the electric fields to which cells were exposed in these reports, and we provide recommendations for assessing nanosecond pulsed electric field effects in electroporation studies.

Multiple nanosecond electric pulses increase the number but not the size of long-lived nanopores in the cell membrane

Exposure to intense, nanosecond-duration electric pulses (nsEP) opens small but long-lived pores in the plasma membrane. We quantified the cell uptake of two membrane integrity marker dyes, YO-PRO-1 (YP) and propidium (Pr) in order to test whether the pore size is affected by the number of nsEP. The fluorescence of the dyes was calibrated against their concentrations by confocal imaging of stained homogenates of the cells. The calibrations revealed a two-phase dependence of Pr emission on the concentration (with a slower rise at<4μM) and a linear dependence for YP. CHO cells were exposed to nsEP trains (1 to 100 pulses, 60ns, 13.2kV/cm, 10Hz) with Pr and YP in the medium, and the uptake of the dyes was monitored by time-lapse imaging for 3min. Even a single nsEP triggered a modest but detectable entry of both dyes, which increased linearly when more pulses were applied. The influx of Pr per pulse was constant and independent of the pulse number. The influx of YP per pulse was highest with 1- and 2-pulse exposures, decreasing to about twice the Pr level for trains from 5 to 100 pulses. The constant YP/Pr influx ratio for trains of 5 to 100 pulses suggests that increasing the number of pulses permeabilizes cells to a greater extent by increasing the pore number and not the pore diameter.

Basic Features of a Cell Electroporation Model: Illustrative Behavior for Two Very Different Pulses

Science increasingly involves complex modeling. Here we describe a model for cell electroporation in which membrane properties are dynamically modified by poration. Spatial scales range from cell membrane thickness (5xa0nm) to a typical mammalian cell radius (10xa0

Electric Field-Driven Water Dipoles: Nanoscale Architecture of Electroporation

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Picosecond and Terahertz Perturbation of Interfacial Water and Electropermeabilization of Biological Membranes

μm), and can be used with idealized and experimental pulse waveforms. The model consists of traditional passive components and additional active components representing nonequilibrium processes. Model responses include measurable quantities: transmembrane voltage, membrane electrical conductance, and solute transport rates and amounts for the representative “long” and “short” pulses. The long pulse—1.5 kV/cm, 100xa0

Frequency spectrum of induced transmembrane potential and permeabilization efficacy of bipolar electric pulses

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