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Experimental results on the effect of a single 5-6 ns, 5-7 MV/m electric pulse on electrically excitable bovine chromaffin cells are presented. Effects on intracellular calcium level were assessed by loading the cells with the calcium-sensitive fluorescence indicator Calcium Green and imaging the cells during nanosecond field exposure in microelectrode chambers that were fabricated on a glass microscope slide with gold electrodes. Consistent with earlier findings that utilized different microelectrode chambers for pulse exposure, a single pulse elicited a rapid and transient rise in intracellular calcium by a mechanism that depends on extracellular calcium, which appears to enter the cells largely through voltage-gated calcium channels. In parallel experiments to assess catecholamine release, chromaffin cells were placed into electroporation cuvettes for nanosecond pulse exposure. As measured by high performance liquid chromatography coupled with electrochemical detection, a single pulse elicited an increase in both norepinephrine and epinephrine release that was also dependent on extracellular calcium and involved influx of calcium through voltage gated-calcium channels. Taken together these results indicate that a single nanosecond pulse can act as a stimulus to trigger calcium-dependent catecholamine release from chromaffin cells. [ABSTRACT FROM AUTHOR]

A compact pulse generator capable of producing high voltage pulses with half-maximum widths as short as 2.5 ns and amplitudes as high as 5 kV has been developed to enable current and future in vivo and in vitro research into the effects of ultra-short, intense electric fields on biological matter. This pulse generator is small, simple, and free of saturable magnetic cores, which frequently introduce amplitude jitter and an undesirable correlation between amplitude and pulse width. In place of a non-linear pulse-forming network is a single-stage resonant network that drives a bank of junction recovery diodes. The diodes function as an opening switch that commutes current from an inductor to a resistive load. The use of air-core inductors in the resonant network results in a stable output pulse with an amplitude that scales linearly with input voltage and a pulse width that is independent of amplitude. The ability to scale the output amplitude independently of the pulse width simplifies the setup for experiments that require pulses with different electric field strengths but the same rise time and duration. Jurkat T lymphoblast cells exposed to 2.5 ns fields produced by this pulse generator showed an increasing degree of electropermeabilization with increasing pulse dosage and electric field intensity. [ABSTRACT FROM AUTHOR]

Research on the electroperturbation effects of ultrashort high field pulses in cancer cells requires subnanosecond rise time, high voltage pulses delivered to low impedance biological loads. Here we present a compact solid-state pulse generator developed for this application. The pulse is generated by switching a chain of avalanche transistors configured as a tapered transmission line from high voltage to ground. The system features a built in 1400:1 capacitively compensated resistive voltage divider. The divider, with a 3 dB point at 910 MHz, overcomes challenges in the direct measurement of the high frequency components of the output pulse. The generator is capable of producing a 0.8 ns rise time, 1.3 ns wide, 1.1 kV pulse into a 50 Ω load at a maximum repetition rate of 200 kHz. Techniques to implement physical layouting strategies to achieve subnanosecond rise times are outlined. Problems faced in integrating the subnanosecond pulse generator with a biological load are discussed. This pulse generator will be used in experiments aimed at electromanipulation of intracellular biomolecular structures. [ABSTRACT FROM AUTHOR]

Megavolt-per-meter electric pulses with durations shorter than charging time constants associated with external cell membrane dielectric properties can generate significant voltages on the membranes of intracellular structures. Nanosecond-duration, high-field (2-4 MV/m) pulses are not immediately lethal to cells and do not produce the conductive openings in the cytoplasmic membrane associated with long-pulse, low-field electroporation, but can induce profound physiological changes, including apoptosis (programmed cell death). We demonstrate rapid, non-destructive, field-dependent translocation of the plasma membrane inner leaflet phospholipid phosphatidylserine in Jurkat T lymphocytes, and we show that cells which exhibit a similar geometry in suspension, rat glioma C6 cells, are highly resistant to these pulses and respond differently even to much higher doses. [ABSTRACT FROM AUTHOR]

This paper describes three pulse generators: a spark gap switched coaxial cable, a spark gap switched Blumlein, and a solid state modulator, developed for applying ultrashort electrical pulses to biological materials in culture. Recent research has shown that ultrashort pulsed electric fields can induce apoptosis in biological cells, and that pulses as short as 10 ns with field amplitude greater than 1 MV/m cause membrane phospholipid rearrangement and activation of the effector enzymes of apoptosis. Pulses of very short duration use only tens of mJ per mL per pulse to induce apoptosis and other intracellular effects without causing thermal trauma. The pulse generators discussed here, each of a different topology, deliver ns pulsed electric fields (nsPEF) to cells in liquid suspension, and can be modified to drive electrodes for external, surgical, or endoscopic treatment of tissues in situ. [ABSTRACT FROM AUTHOR]