Single Cell Electroporation in Organotypic Brain Slice Cultures: An In Vitro Technique to Deliver Plasmids Inside Targeted Neurons in Brain Hippocampal Slices
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Begin electroporation by taking a glass pipette containing the desired plasmid solution. Attach it to a micropipette holder fitted with a silver wire electrode, ensuring that the electrode is inserted into the micropipette. Take an electroporation chamber containing an ultrathin hippocampal section submerged in artificial cerebrospinal fluid.
Dip the ground electrode into the fluid present in the chamber for subsequent electroporation. Lower the pipette tip into the electroporation chamber and record the baseline resistance. Use a microscope to position the pipette tip near the target cell. Once the pipette is in contact with the neuronal membrane, its resistance increases sharply.
Apply positive air pressure to create a small invagination in the neuronal membrane. Rapidly apply mild suction to push the membrane up, creating a loose seal between the pipette tip and the membrane. Repeat this process a few more times to condition the membrane and avoid cell lysis.
After conditioning, apply strong suction to form a tight seal with a membrane called gigaohm seal. Use electric pulses to form transient pores in the sealed area of the membrane. These openings facilitate the entry of plasmids inside the targeted neuron. Post electroporation, the pores reseal, entrapping plasmids inside the neuron.
