Hyperthermia exacerbates and hypothermia attenuates methamphetamine (METH)-induced dopamine (DA) neurotoxicity. The mechanisms underlying these temperature effects are unknown. Given the essential role of the DA transporter (DAT) in the expression of METH-induced DA neurotoxicity, we hypothesized that the effect of temperature on METH-induced DA neurotoxicity is mediated, at least in part, at the level of the DAT. To test this hypothesis, the effects of small, physiologically relevant temperature changes on DAT function were evaluated in two types of cultured neuronal cells: (1) a neuroblastoma cell line stably transfected with human DAT cDNA and (2) rat embryonic mesencephalic primary cells that naturally express the DAT. Temperatures for studies of DAT function were selected based on core temperature measurements in animals exposed to METH under usual ambient (22°C) and hypothermic (6°C) temperature conditions, where METH neurotoxicity was fully expressed and blocked, respectively. DAT function, determined by measuring accumulation of radiolabeled DA and 1-methyl-4-phenylpyridinium (MPP ϩ ), was found to directly correlate with temperature, with higher levels of substrate uptake at 40°C, intermediate levels at 37°C, and lower levels at 34°C. DAT-mediated accumulation of METH also directly correlated with temperature, with greater accumulation at higher temperatures. These findings indicate that relatively small, physiologically relevant changes in temperature significantly alter DAT function and intracellular METH accumulation, and suggest that the effect of temperature on METHinduced DA neurotoxicity is mediated, at least in part, at the level of the DAT.Key words: dopamine; dopamine transporter; temperature; methamphetamine; MPP ϩ ; neurotoxicityThe psychostimulant methamphetamine (METH) is a potent dopamine (DA) neurotoxin in rodents, nonhuman primates (Gibb et al., 1994;Lew et al., 1998) and, possibly, humans (McCann et al., 1998Volkow et al., 1999). METH administration leads to longlasting reductions in a number of DA axonal markers including DA, dihydroxyphenylacetic acid (DOPAC), homovanillic acid, tyrosine hydroxylase activity, and the DA transporter (DAT). Recent studies indicate that the striatal vesicular monoamine transporter is also reduced on a long-term basis after METH (Frey et al., 1997;Villemagne et al., 1998). Morphological studies indicate that loss of these presynaptic DA axonal markers is related to destruction of DA axons and axon terminals (Ellison et al., 1978;Lorez, 1981;Ricaurte et al., 1982Ricaurte et al., , 1984Bowyer et al., 1994;Broening et al., 1997;Fukumura et al., 1998), generally with sparing of DA nerve cell bodies (Ricaurte et al., 1982;Woolverton et al., 1989; but see Sonsalla et al., 1996;Hirata and Cadet, 1997). The mechanism by which METH induces dopaminergic neurotoxicity is not known. However, there is compelling evidence that the DAT plays an essential role (Marek et al., 1990;Pu et al., 1994;Fumagalli et al., 1998). The DAT is an integral membrane protein of DA neu...