External Excitation of One-Dimensional Patterned Neuronal Cultures

Prior to an electric stimulation, place the seated coverslip into an imaging chamber. To stimulate the cells with an electric field of a constant direction, use two parallel electrodes separated by 11 millimeters and located at opposite sides of the culture plated on the 13-millimeter coverslips. Then, assemble the culture imaging chamber and matching electrode holder such that the wire electrodes are immersed in the extracellular media and positioned one millimeter above the culture.

Then, plug the single electrode pair to the operational amplifier that amplifies the signal from the generator. Apply a single pulse of bipolar square wave with a 50% duty cycle and amplitude of 22 volts. Use a bipolar wave with no DC component to avoid electrolysis that may occur at the electrodes. Apply the pulse lasting from 10 microseconds to four milliseconds to ensure effective stimulation, but prevent cell injury.