The neurotoxin 1-methyl-4-(2'-aminophenyl)-1,2,3,6-tetrahydropyridine (2'-NH(2)-MPTP) damages forebrain serotonin (5-HT) and norepinephrine (NE) nerve terminals while sparing striatal dopaminergic innervation. Previous studies suggest that 2'-NH(2)-MPTP acts by a mechanism that involves uptake by the plasma membrane 5-HT and NE transporters. The present investigation further explores the molecular mechanism of 2'-NH(2)-MPTP with regard to cellular transport and effects on body temperature. Mice with genetically controlled decreases in serotonin transporter (SERT) expression were studied to corroborate pharmacologic evidence implicating SERT in 2'-NH(2)-MPTP-induced serotonin neurotoxicity. To investigate whether sequestration by the intracellular vesicular monoamine transporter type 2 (VMAT2) occurs, mice with reduced VMAT2 expression or mice receiving the VMAT2 inhibitor Ro 4-1284 (2-hydroxy-2-ethyl-3-isobutyl-9,10-dimethoxy-1,2,3,4,6,7-hexahydrobenzo[alpha]chinolizin hydrochloride) were treated with 2'-NH(2)-MPTP. Body temperature was measured as a function of reduced SERT or VMAT2 expression. 2'-NH(2)-MPTP caused a 2 degrees C drop in temperature that was attenuated by decreased SERT but not VMAT2. In addition, complete loss of SERT attenuated cortical and hippocampal depletions in 5-HT but not NE. In contrast, mice with a 50% reduction in VMAT2 exhibited similar 5-HT and NE toxicity when compared with wild-type mice at higher doses of 2'-NH(2)-MPTP, whereas a slight potentiation of toxicity was observed at very low doses of 2'-NH(2)-MPTP. Pharmacologic inhibition of VMAT2 caused minimal potentiation of neurotransmitter depletions in response to moderate doses of 2'-NH(2)-MPTP. Thus, 2'-NH(2)-MPTP seems to be similar to MPTP in its requirement for selective plasma membrane transport and the expression of acute hypothermia; however, unlike MPTP, VMAT2 does not appear to play a major role in the toxic mechanism of 2'-NH(2)-MPTP.