Protocol for making a set of three serotonin drug plates and a set of three naloxone drug plates. Serotonin (20 mM) is added to molten agar, whereas naloxone (10 mM) is made into a 10X stock solution in water first before being spread over agar plates to achieve the final concentration. Each 35 mm drug plate contains 3 mL low peptone NGM agar and the drug, and is always made up fresh the day before imaging experiments. For naloxone methods, refer to https://www.ncbi.nlm.nih.gov/pubmed/25898004.