Ionizing radiation can be an environmental health risk as well as a cancer therapeutic, but similar roles for non-ionizing radiation are controversial. We have examined effects of wideband, intense non-ionizing radiation applied to cells and tissues as nanosecond pulsed electric fields (nsPEFs). Compared to conventional electroporation pulses, nsPEFs have shorter pulse durations (>= 10ns) and higher electric fields (similar to 300kV/cm). In spite of the high electric field, measured thermal changes are negligible. For these short pulses we have observed a number of nsPEF-induced effects on biological cells and tissues, including apoptosis induction and tumor regression at high electric fields, as well as calcium mobilization, platelet aggregation and enhanced gene transcription at lower, sublethal electric fields. When nsPEFs are below the plasma membrane (PM) charging time constant (similar to 100ns), effects on intracellular membranes are appreciable. Thus, it is possible to observe effects on intracellular structures (membranes) and functions without observing effects on the PM. For Jurkat or HL-60 cells exposed to 10ns or 60ns pulses, mobilization of calcium from intracellular stores is observed at significantly lower electric fields than electric fields needed for PM electroporation or transport. However, as the electric field and/or the pulse duration are increased, it is possible to observe effects on the PM. Thus, as the pulse duration increases there is a cell type-specific increase in effects at the PM, as well as on intracellular structures and functions. To determine nsPEF characteriastics that are responsible for these diverse cell responses, we designed experiments to determine if pulse duration, electric field, or energy density was the stimulus that recruited responses from cells to nsPEFs. The results indicate that nsPEF effects are not due to dose effects, but instead follow a scaling law (E tau) defined by the product of electric field (E) and pulse duration (tau, tau), which can be derived from circuit equations that describe membrane charging for pulses short compared to the membrane charging time constant. For both increases in ethidium homodimer fluorescence, which is used as a marker for plasma membrane electroporation and/or transport, and increases in phosphatidylserine externalization measured by Annexin-V-FITC fluorescence, which is a marker for plasma membrane lipid translocation, a threshold response to increases in both markers exhibited the same E tau value. This law also holds true for intracellular effects that are measured on isolated cells and intact tissues. Thus, the intracellular activation of caspase proteases, which are the executioners for programmed cell death by apoptosis, was independent of the energy density when pulses were applied at 10, 60 and 300ns, but directly proportional to ET. Furthermore, caspase activation in intact fibrosarcoma tumors at 300ns was directly proportional to the electric field. Finally, for apoptosis markers and nsPEF-induced cell death in HCT116 colon carcinoma cells, the E tau scaling law was extended to include pulse number (n) into the equation (E tau n). Thus, nsPEF effects are due to charging of the plasma membrane, as well as effects on intracellular structures and functions in cells and tissues that are defined by the E tau scaling law. It is anticipated that when tau significantly diminishes and charging mechanisms are not likely to predominate, the E tau scaling law will not hold and a new paradigm of ultrashort pulsed electric field effects is expected.
