Injection of horseradish peroxidase solution in calibrated volumes (nanolitre) into the spinal cord

A method for the injection of horseradish peroxidase (HRP) into the spinal cord (SC) is described, based on the principle of fluid expansion upon application of heat. HRP solution in the tip of a glass micropipette (tip diameter 10-15 microm) was extruded when the sealed stem of the pipette, containing distilled water, was warmed. The pipette was calibrated (temperature/volume relation) before it was inserted into the SC and the injection of HRP was made according to the calibration curve. Using the pipette as an electrode it was possible to locate its tip with reference to recorded field potentials. After the injection, the volume of HRP solution actually injected was determined.