Rats raised in an isolated condition (IC) are impulsive and hyperactive compared to rats raised in an enriched condition (EC), suggesting that isolation rearing may be a preclinical model of attention-deficit/hyperactivity disorder (ADHD). The current study determined if administration of methylphenidate (MPH), a dopamine transporter (DAT) blocker used in the treatment of ADHD, reduces the hyperactivity observed in IC rats toward levels observed in EC rats. Another goal was to determine if chronic MPH treatment differentially alters DAT function in EC and IC rats in medial prefrontal cortex (mPFC) or orbitofrontal cortex (OFC). IC and EC rats were treated with either MPH (1.5 mg/kg, p.o.) or vehicle from postnatal days (PND) 28-51. On PND 28 and 51, rats were evaluated for MPH-induced locomotor activity. On PND 55-63, in vitro [(3)H]DA uptake assays were performed in mPFC and OFC. At both PND 28 and 51, IC rats were hyperactive compared to EC rats. At PND 28, MPH increased activity in EC rats only. At PND 51, MPH did not alter locomotor activity in IC or EC rats. Beginning at PND 55, basal uptake of [(3)H]dopamine in IC rats was higher in mPFC and lower in OFC compared to EC rats. The basal differences in DAT function were normalized by MPH treatment in mPFC, but not in OFC. These findings suggest that isolation rearing may not represent a valid predictive model for screening effective medications in the treatment of hyperactivity associated with ADHD.