The dopamine transporter is the primary means of inactivating synaptic dopamine as well as a major site of action for psychostimulants (such as cocaine and amphetamine) and for neurotoxins that induce parkinsonism. In the present study, a human dopamine transporter partial cDNA clone obtained by polymerase chain reaction exhibited 87% and 89% identity at the nucleic acid and amino acid levels, respectively, with transmembrane domains 3-5 of the rat homolog. This clone was used to quantitate human dopamine transporter mRNA by nuclease protection assay. The postmortem content of dopamine transporter mRNA in the substantia nigrae of 18-to 57-yr-old subjects was relatively constant, while in subjects >57 yr old, a precipitous (>95%) decline in substantia nigra dopamine transporter mRNA was evident. In contrast, tyrosine hydroxylase mRNA in the same samples declined in a linear manner with increasing age. In situ hybridization experiments confirmed the profound loss of dopamine transporter gene expression in melanin-positive (presumptive dopamine) nigral neurons. These data may begin to shed light on compensatory changes occurring in human dopamine neurons during normal aging.The actions of many neurotransmitters are terminated by rapid reuptake into presynaptic nerve endings via neurotransmitter-specific, high-affinity, Na+-dependent membrane transporter proteins. The transporter for the neurotransmitter dopamine (DA) is apparently the receptor through which psychostimulants such as cocaine and amphetamine exert their potent reinforcing properties, leading to psychostimulant abuse and dependence (1). In addition, the DA transporter mediates the active accumulation of neurotoxins such as 1-methyl-4-phenylpyridine into DA neurons (2, 3), resulting in DA cell death and a parkinsonian syndrome. Thus, understanding the regulation of human DA transporter gene expression is an important step toward elucidation of the molecular bases of psychostimulant drug abuse and of neurotoxin-induced (and quite possibly idiopathic) parkinsonism.Ligand binding to the human DA transporter decreases with age (4, 5), in keeping with other evidence for an age-related loss of DA neurons (6). Nevertheless, surprisingly little is known about the nature of normal age-related changes in human DA neurons, a process apparently distinct from the neuronal changes seen in Parkinson disease (6, 7). In the present study, a human DA transporter cDNA clone was obtained by polymerase chain reaction, exploiting predicted sequence homology with the recently cloned, functionally related human norepinephrine transporter (8). With the use of this clone, a precipitous, age-related loss of DA transporter mRNA was observed in human substantia nigra, while tyrosine hydroxylase mRNA, another phenotypic marker of DA neurons, decreased more linearly with age. These data may shed light on some compensatory changes occurring in human DA neurons during the normal aging process.
MATERIALS AND METHODSHuman substantia nigrae used for biochemical analyses (n = 36) w...