The interaction of amphetamine (AMPH) with the dopamine (DA) transporter (DAT) is thought to be critically important for the DA-elevating actions of this drug. It is commonly believed that DA elevations are involved in the rewarding͞reinforcing properties of AMPH and other drugs of abuse. Here, we found that DAT deletion did not eliminate the rewarding effects of AMPH as measured by conditioned place preference (CPP). In fact, mice in which the DAT gene has been deleted (DAT-KO mice) exhibited AMPH-induced CPP for many weeks after the time when extinction occurred in WT mice. Moreover, systemic AMPH still increased extracellular DA in the nucleus accumbens (NAc) of mice lacking the DAT, although local infusion of AMPH into the NAc did not have this effect. By using voltammetry in NAc slices, we found that AMPH did not decrease the rate of DA clearance. The rate of ventral tegmental area DA neuron firing was dramatically inhibited by AMPH in brain slices from WT mice, but there was no inhibition of firing in DAT-KO mice. AMPH-induced CPP was abolished by pretreatment with WAY-100635, a serotonin 5-HT 1A receptor antagonist, in DAT-KO mice, but the drug did not change AMPH place preference in WT mice. Therefore, despite the absence of the DAT, AMPH displays rewarding effects and causes an increase in extracellular DA in the NAc of DAT-KO mice, acting indirectly in this case. The 5-HT system may be involved in the rewarding effects of AMPH in these mice.voltammetry ͉ electrophysiology ͉ microdialysis ͉ release ͉ uptake T he stimulating and reinforcing effects of psychostimulants are thought to result from enhanced extracellular dopamine (DA) concentrations in specific brain areas, including the caudate putamen (CPu) and nucleus accumbens (NAc). In fact, the magnitudes of stimulant-induced psychomotor activation and DA response in these regions are highly correlated (1-3). Amphetamine (AMPH) increases extracellular DA by several different mechanisms. Like other stimulants, AMPH decreases uptake of released DA by means of inhibition of the plasma membrane DA transporter (DAT) (4). AMPH decreases DA uptake not only by direct competition for the DAT, but also by reducing the available cell-surface DAT (5). In addition, AMPH displaces DA from synaptic vesicles to the cytoplasm (6) and promotes nonvesicular DA release by reverse transport through the DAT (7,8). AMPH also inhibits monoamine oxidase (9), thereby increasing cytosolic DA levels and leading to greater release of DA through reversal of the DAT. Finally, AMPH exhibits DAT-dependent influences on midbrain glutamatergic neurotransmission and ionic currents that may enhance terminal release of DA (10-12). Consequently, convergent evidence suggests that the striatal DAT is critically important for the extracellular DA-elevating action of AMPH as well as its stimulating and rewarding effects. This is consistent with earlier hypotheses that neural substrates for processing the reinforcing and stimulant properties of psychostimulants may be similar, if not identical (...