Using a computerized infrared activity analysis system, the dose-response relationship, timing, and duration for stimulation of motor activity after a single dose of methylphenidate was studied in Sprague-Dawley rats. After 5 days of acclimation and 2 days of monitored baseline activity, rats received a single subcutaneous injection of vehicle or of 0.6, 2.5, 10 or 40 mg/kg methylphenidate 1 h into the dark cycle. Recording was then resumed for an additional 36 h. Five locomotor indices were analyzed. Each locomotor parameter monitored different aspects of motor activity. The doses of 2.5, 10 and 40 mg/kg significantly increased (P < 0.01) locomotor activity. The time to maximal effect (20, 50, and 90 min) and duration of effect (70, 210, and 280 min) increased with dose respectively. Ten mg/kg had the maximum effect on locomotor activity, while the largest dose, 40 mg/kg, elicited a more focused stereotyped activity that limited the amount of forward ambulation. Single injections of methylphenidate did not alter motor activity the next day. Pharmacological parameters and specific locomotor parameters describing the effects of methylphenidate at the beginning of the dark cycle can later be used in chronopharmacologic studies. They will also provide the basis for investigation of adaptive mechanisms during repeated or chronic administration of methylphenidate.