Variations of Intracellular Ca²⁺ Mobilization Initiated by Nanosecond and Microsecond Electrical Pulses in HeLa Cells.

Abstract

Herein, the variations in transient Ca²⁺ mobilizations in HeLa cells exposed to a single, non-thermal pulsed electric field (PEF) are described. Three PEF waveforms categorized by pulse duration and intensity were used to deduce the kinetics involved in Ca²⁺ mobilization. A fast microscopic fluorescent imaging system and a fluorescent molecular probe were used to observe transient intracellular Ca²⁺ mobilization after pulse exposure. The sources and pathways in the transient Ca²⁺ mobilizations were investigated using an inhibitor of inositol-1,4,5-trisphosphate receptor (IP₃R) on the endoplasmic reticulum (ER) along with a Ca²⁺-free buffer. When exposed to the 10-μs-long PEF, the Ca²⁺ concentration increased mainly at the cathodic region near the membrane. However, Ca²⁺ concentration increased at both anodic and cathodic regions when Na⁺ concentration in the buffer was reduced. Ca²⁺ concentration increased only in the presence of extracellular Ca²⁺. These results suggest that the 10-μs PEF takes a large amount of extracellular Na⁺ into the cell through the electropermeabilized plasma membrane, especially at the anodic side, resulting in the suppression of the Ca²⁺ influx. On the contrary, the 20-ns-long PEF increased Ca²⁺ concentration in the surrounding region of the nucleus only in the presence of extracellular Ca²⁺. The PEF exposure with inhibition of the IP₃R indicates that increased Ca²⁺ ions are released from the ER via the activated IP₃R. These mechanisms could induce specific cell responses, such as Ca²⁺ oscillations, Ca²⁺ waves, and Ca²⁺ puffs.