Intrathecal injection is a technique used to deliver drugs, viral vectors, or tracers directly into the cerebrospinal fluid (CSF) via the spinal canal. The protocol involves anesthetizing the subject (e.g., mouse or rat), positioning it properly to access the lumbar region, inserting a fine-gauge needle into the intrathecal space, slowly injecting the substance, and monitoring for post-procedure recovery. This method is widely used in pain research, gene therapy, and neurological studies.