This article details the process of obtaining a whole-cell configuration for electrophysiological recordings from brain slices. It outlines the steps necessary to ensure accurate positioning and sealing of the recording pipette with the target neuron.
Take a recording pipette with a wire electrode in an electrolyte solution.
Position the pipette above an immobilized brain slice.
Adjust the pipette current to reduce background signals.
Use a microscope to locate the target neuron and move the pipette toward the neuron.
Apply a slight positive pressure and lock it.
The positive pressure disturbs the tissue surface, indicating proximity to the cell.
Bring the pipette closer until a dimple forms on the membrane, signifying correct positioning.
Release the positive pressure.
Observe an increase in resistance as the cell forms a seal with the pipette.
Hold the membrane potential at a constant negative voltage for stability.
Pull the micropipette away to remove excess pressure.
Adjust the pipette's fast and slow electrical responses for accuracy.
Apply brief suction to break through the cell, establishing a direct electrical connection.
The whole-cell voltage clamp configuration is now ready for electrophysiology recording.
To obtain whole-cell configuration, place the recording pipette just above the slice, and offset pipette current in the Voltage Clamp mode. Apply slight positive pressure to the pipette and lock the stopcock.
Next, select a healthy cell with an intact membrane and approach the tissue with the pipette. The positive pressure should cause a slight disturbance on the tissue. Slowly bring the pipette closer to the cell in a diagonal motion until a small dimple is formed on the cell surface. Then, release the positive pressure lock. The cell will begin to form a seal and the resistance will increase above one gigaohm. In voltage clamp, hold the cell at -68 millivolts.
Subsequently, slightly pull the pipette away from the cell diagonally to remove excess pressure. Compensate for the fast and slow pipette capacitance. Apply a brief suction through the tube connected to the pipette holder to break through the cell and obtain a whole-cell configuration.
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