Accumulating evidence suggests that genetic factors contribute to the vulnerability to methamphetamine (MAP) abuse and associated psychiatric symptoms. Chronic MAP abuse leads to psychosis, which may be of a transient or a prolonged type. Serotonergic dysfunction has been proposed as one of the contributory factors in the development of MAP psychosis. Our PET studies revealed that the serotonin transporter (5-HTT) density in global brain regions is significantly lower in MAP abusers. In this study, we examined the role of a functional polymorphism in the 5′ flanking region of the 5-HTT gene (5-HTTLPR) in the development of MAP psychosis in a Japanese population. We analyzed DNA samples from 166 MAP patients (95 with transient and 71 with prolonged psychosis) and 197 age-, sex-, and geographic-origin-matched healthy controls. Patients were also subdivided according to the presence (n = 119) or absence (n = 148) of spontaneous relapse. We observed significant genotypic association of the 5-HTTLPR polymorphism with MAP psychosis (P = 0.022), particularly in patients who show prolonged psychosis. The frequency of the S allele in patients with prolonged psychosis was significantly higher than that of the controls (P = 0.045); it was further higher in patients with prolonged psychosis with spontaneous relapse (P = 0.004). 5-HTTLPR has been suggested to regulate the transcriptional activity of 5-HTT, with S alleles showing lesser transcriptional efficiency and also lower 5-HT1A receptor-binding potential. Prolonged MAP use, combined with the high frequency of 5-HTTLPR S-alleles, may lead to reduced 5-HTT levels and 5-HT1A receptor-binding potential in the brain, resulting in the dysfunction of the serotonergic system. Thus, we suggest a possible role for the 5-HTTLPR polymorphism in MAP psychosis.