[Effects of energy controllable steep pulses on intracellular calcium concentration and cell membrane potential].

Background and objective: Our previous experiments showed that steep pulses could kill tumor cells, but the mechanism is unclear. This study was to probe the effects of different dosages of energy controllable steep pulses (ECSP) on intracellular concentration of dissociative calcium ion ([Ca2+]i) and cell membrane potential.

Methods: The breast carcinoma MDA-MB-231 cells were divided into control group and five ECSP (different dosages) groups. Ca2+ was labeled by Fluo-3/AM and cell membrane potential was labeled by DiBAC4(3). The mean fluorescence intensity in MDA-MB-231 cells was observed by laser confocal microscopy after ECSP treatment. The changes of calcium concentration and cell membrane potential after ECSP treatment were analyzed. The changes of intracellular [Ca2+]i after ECSP treatment were also observed either with or without Ca2+ outside of the cells.

Results: Ca2+ outflow was observed when the cells were treated with lower dosage of pulse in quiet state; the outflow was enhanced with the dosage increase. In real-time kinetic detection, intracellular Ca2+ concentration was increased with the increase of pulse electric field intensity when cells were treated with lower dosages of ECSP. When the voltage was 285 V, frequency was 100 Hz, [Ca2+]i decreased obviously. The intracellular Ca2+ concentration was obviously lower in the cells without outside Ca2+ than in cells with outside Ca2+, but it still increased gradually. Low dosage of ECSP induced the increase of cell membrane potential, indicating the depolarization of cell membrane. With increase of the dosage, cell membrane potential was attenuated, indicating the superpolarization of cell membrane.

Conclusion: Lower dosage of ECSP can induce the depolarization of cell membrane and the inflow of outside Ca2+; higher dosage of ECSP can directly destroy the cell membrane and induce the superpolarization of cell membrane, then induce the outflow of intracellular Ca2+ which causes the necrosis of tumor cells.