Rat: 20 mg/kg per day, PO, 35 days.

HEK-hDAT and HEK-hSERT cells were incubated in modified Eagle’s medium supplemented with 5% fetal bovine serum, 5% calf bovine serum, 0.05 U penicillin/streptomycin and puromycin (2 μg/mL). HEK-hNET cells were incubated in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum, 0.05 U penicillin/streptomycin and geneticin (300 μg/mL). Cells were grown until confluent on 150 mm diameter tissue culture dishes in a humidified 10% CO2 environment at 37◦C. [125I]RTI-55 (Perkin Elmer Life Sciences, Boston, MA) was used in competition binding assays characterizing DOV 102,677 affinity for the three monoamine neurotransmitter transporters. Aliquots (50 μL) of the suspended cells were added to assay tubes containing drugs and Krebs-HEPES assay buffer in a final assay volume of 0.5 mL. Uptake inhibition studies were conducted using triplicate determinations for each test substance. [3H]DA, [3H]5-HT, or [3H]NE (Perkin Elmer Life Sciences, Boston, MA, 56, 26.9, 60 Ci/mmol, respectively, 20 nM final concentration) was added after a 10 min pre-incubation of the isolated cells in a 25◦C water bath, and the assay incubated an additional 10 min. The reaction was terminated by vacuum filtration over Whatman GF/C filters using a 96-well Tomtec cell harvester (Hamden, CT). Specific uptake was defined as the difference in uptake observed in the absence and presence of 5 μM mazindol (hDAT and hNET) or 5 μM imipramine (hSERT).