Single nanosecond pulsed power induced structural modifications of medaka fish embryo

Abstract

The effects of nanosecond pulsed electric field on the medaka fish egg development in low and high conductivity mediums are investigated. A pulsed power modulator using a magnetic compression circuit (MPC) was employed to generate 0.5 to 20 kV pulses with 50 nanosecond pulse duration. Input voltage and current were measured by using a high voltage probe, a current monitor, and an oscilloscope. Fertilized eggs of strain d-rR medaka were used. The ages of the experimental eggs were 1, 6, and 48 hours post fertilization (hrPF). Fluorescein isothiocyanate (FITC)-Dextran conjugates with different molecular weights and a florescence microscope were used to study the effects of the pulsed electric field on the blastomeres and the extend of eggs structure damage. The FITCs were injected to experimental eggs by a micro-injection system, which can deliver nano-liters of FITC without producing any damage to egg structure. In group A, 10 ng of FITC was injected to blastomere of eggs at 1 hrPF. In group B, 30 ng of FITC was injected to yolk sphere at 1 hrPF. Group C eggs were kept as control. After injection, each egg was set at the middle of a 2 mm or 4 mm cuvette and a single electric pulse was applied. The control eggs were sham treated. After pulse application the eggs were observed under the fluorescent microscope until they hatched or died. By applying low electric field pulses (up to 10 kV/cm) in 17 mS/cm medium (PBS), the eggs showed small size temporary pore formation. These eggs could recover and grew as fish. By increasing the electric field (10 to 20 kV/cm) more FITC flow from blastomere were observed, embryos showed abnormal growth with deformed body structure or missing eyes. For higher electric fields (over 30 kV/cm) immediate extended damage were observed.