Electroporation by using pulsed electric fields with long durations compared with the charging time of the plasma membrane can induce cell fusion or introduce xenomolecules into cells. Nanosecond pulse power technology generates pulses with high-intensity electric fields, but with such short durations that the charging time of the plasma membrane is not reached, but intracellular membranes are affected. To determine more specifically their effects on cell structure and function, human cells were exposed to high intensity (up to 300 kV/cm) nanosecond (10-300 ns) pulsed electric fields (nsPEF) and were analyzed at the cellular and molecular levels. As the pulse duration decreased, plasma membrane electroporation decreased and appearances of apoptosis markers were delayed. NsPEF induced apoptosis within tens of minutes, depending on the pulse duration. Annexin-V binding, caspase activation, decreased forward light scatter, and cytochrome c release into the cytoplasm were coincident. Apoptosis was caspase- and mitochondria-dependent but independent of plasma membrane electroporation and thermal changes. The results suggest that with decreasing pulse durations, nsPEF modulate cell signaling from the plasma membrane to intracellular structures and functions. NsPEF technology provides a unique, high-power, energy-independent tool to recruit plasma membrane and/or intracellular signaling mechanisms that can delete aberrant cells by apoptosis.

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A simple electrical model for living cells predicts an increasing probability for electric field interactions with intracellular substructures of both prokaryotic and eukaryotic cells when the electric pulse duration is reduced into the sub-microsecond range. The validity of this hypothesis was verified experimentally by applying electrical pulses (durations 100 micros-60 ns, electric field intensities 3-150 kV/cm) to Jurkat cells suspended in physiologic buffer containing propidium iodide. Effects on Jurkat cells were assessed by means of temporally resolved fluorescence and light microscopy. For the longest applied pulses, immediate uptake of propidium iodide occurred consistent with electroporation as the cause of increased surface membrane permeability. For nanosecond pulses, more delayed propidium iodide uptake occurred with significantly later uptake of propidium iodide occurring after 60 ns pulses compared to 300 ns pulses. Cellular swelling occurred rapidly following 300 ns pulses, but was minimal following 60 ns pulses. These data indicate that submicrosecond pulses achieve temporally distinct effects on living cells compared to microsecond pulses. The longer pulses result in rapid permeability changes in the surface membrane that are relatively homogeneous across the cell population, consistent with electroporation, while shorter pulses cause surface membrane permeability changes that are temporally delayed and heterogeneous in their magnitude.