Purpose: Brain imaging: to investigate the effects of ecstasy use alone and in combination with amphetamine use on dopamine function by comparing Beta-CIT binding in SPECT scans of ecstasy users, ecstasy and amphetamine users and non-ecstasy using controls.

Design: Retrospective (non-experimental) 3-group between-subjects design, with drug use (ecstasy, ecstasy + amphetamine or no drugs) as the between-subjects factor, with all participants receiving SPECT scans with the radioligand [123I]-Beta-CIT.

Subjects: 29 ecstasy users, 9 ecstasy and amphetamine users and 15 non-drug user controls residing in the Amsterdam (Netherlands) area, and recruited via flyers distributed at locations associated with the dance event (rave) scene. Matching - Matched on gender distribution. (Some participants in this study also appeared in two prior studies by the authors; Reneman et al. 2001b; Reneman et al. 2001c)

Criteria for Inclusion, Ecstasy Users - No information provided; in previous paper, criteria was self-reported lifetime use of at least 50 ecstasy tablets. Ecstasy + Amphetamine Users - Reported having used ecstasy (perhaps no less than 50 times) and intentional use of amphetamines at least once in 3 months prior to study. Non-user controls - No self-reported lifetime use of ecstasy or amphetamine, though use of other drugs was permitted. All Groups - Healthy, as apparently indicated through medical examination and no past or current psychiatric illness, between the ages of 18 and 45 years old, not pregnant, and abstinent from all psychoactive drugs for 3 wks before study day, with abstinence verified through urinary drug screen.

Drug Use Parameters - Ecstasy Use - Ecstasy users reported taking a total lifetime dose of 324 +/- 527 tablets, reporting an average dose per occasion of 1.7 +/- 0.7 tablets. Information on frequency of use is not provided. Duration of use, in months, was 61.2 +/- 34.8 months, and time since last use, in days, was 102 +/- 138 days. Ecstasy + amphetamine users reported a total lifetime dose of 358 +/- 610 tablets, with an average dose per occasion of 2.3 +/- 0.7 tablets. No information on frequency of use is provided. Duration of use, in months, was 52.8 +/- 22.8 months, and time since last use, in days, was 30 +/- 10 days. Amphetamine use - Ecstasy + amphetamine users reported using amphetamine 47.5 +/- 45 times a year, with average dose, in grams, was .41 +/- .31 grams. Other drug use - Alcohol use, reported in units per wk, was 10.3 +/- 8.6 units for ecstasy users, 12.8 +/- 13.9 units for ecstasy + amphetamine users and 10.6 +/- 9.8 units for non-user controls. Tobacco use, in cigarettes per day, was 8.5 +/- 8.0 cigarettes for ecstasy users, 12.7 +/- 13.2 cigarettes for ecstasy + amphetamine users and 10 +/- 5 for non-user controls. Cannabis, in joints per 3 months prior to study, was 54.1 +/- 107 joints for ecstasy users, 87 +/- 102.5 joints for ecstasy + amphetamine users, and 1.7 +/- 3.1 for non-users; ecstasy + amphetamine users smoked significantly more joints than non-user controls. Both ecstasy users and ecstasy + amphetamine users reported using LSD and "magic mushrooms" at least once during a lifetime.

Group Demographics and Matched Variables - Members of all groups matched on gender. Gender, as M/F ratio - Ecstasy users, 15/14, ecstasy + amphetamine users, 6/3, non-ecstasy users, 7/8. Age - Average age for ecstasy users was 26.1 +/- 5.6 years, for ecstasy + amphetamine users was 22.1 +/- 2.8 years and 26.1 +/- 5.5 years for non-users. Education - No information provided concerning average years of education in each group, though previous studies by Reneman et al report education of approximately 12 years for ecstasy users and approximately 14.5 years for non-user controls.

Measures: SPECT scans were performed 20 h after injection of 3.8 mCi (140 MBq) [123I]-Beta-CIT. Participants were supine during SPECT scan, with heads parallel to orbitometeal line. A standard template with L and R striatum as ROI used to examine SPECT slices, with template derived from MRI, and the cerebellum used as a control ROI (assumed to have low DA binding). ROI analysis conducted by an investigator blind to participant's drug use history. [123I]-Beta-CIT binding ratios were calculated by division of ROI bindings by binding in the cerebellum.

Analyses: Differences in demographics compared with one-way analysis of variance (ANOVA) with drug use (ecstasy, ecstasy + amphetamine, no ecstasy) serving as a between-group factor. A standard linear regression model was used to analyze DA transporter binding, with [123I]-BetaCIT binding (estimated DA transporter density) serving as the dependent measure and group (3 levels; ecstasy, ecstasy + amphetamine, no ecstasy), gender (two levels; male or female), age and past experience with cannabis as predictors.

Drug Use Parameters and Binding Ratios - Pearson's correlation performed, with Beta-CIT binding ratios in striatum (estimated DA transporter density), past ecstasy use and past amphetamine use entered as factors

. Results - Significant Differences Found: Participant's age predicted (was significantly related to) Beta-CIT binding ratio (relationship unspecified); all analyses of ligand binding controlled for age. Ecstasy users had significantly higher Beta-CIT binding ratios than non-user controls. Beta-CIT binding ratios were significantly lower in ecstasy + amphetamine users than in ecstasy users.

Results - No Significant Differences: Participant's gender and past use of cannabis did not predict (were unrelated to) Beta-CIT binding ratio. Beta-CIT ratios in ecstasy + amphetamine users were not significantly different from binding ratios in non-user controls.

Drug Use Parameters and Binding Ratios - Beta-CIT ratios were not significantly correlated with past ecstasy use. Beta-CIT binding ratios were not significantly correlated with extent of past amphetamine use, though there was a slight trend, with binding ratios decreasing with increased amphetamine use.

Overall Effects: After controlling for the effects of participant's age, sole ecstasy users were found to have higher Beta-CIT striatum/cerebellum binding ratios than either individuals who had never used ecstasy or ecstasy users who reported intentionally using amphetamine as well as ecstasy. People who reported using ecstasy and amphetamines had lower Beta-CIT ratios than ecstasy users, but not non-user controls. Past ecstasy use did not correlate with Beta-CIT binding ratios, and while there was a trend for past amphetamine use to be associated with lower binding ratios, this was not statistically significant. Past cannabis use was not associated with Beta-CIT binding ratios. While gender was not associated with Beta-CIT ratios, age was associated with Beta-CIT binding ratios. Since binding ratios are intended to assess dopamine transporter binding site density, the findings of these studies fail to support a simple, linear relationship between past ecstasy use and number of DA transporter binding sites. On the other hand, it is possible that the combined use of ecstasy and amphetamine, or even the use of amphetamine alone, may reduce number of DA transporter binding sites.

Comments: This paper is one among several attempting to assess the contributions made by substances other than ecstasy to differences found between ecstasy users and non-users on measures of psychiatric and cognitive function, and (in this case) measures of transporter. The findings reported here do not support any links between the appearance of parkinsonism and ecstasy use. This possible relationship had been hypothesized in a case report (Mintzer et al. 1998), although this interpretation was disputed by others (Baggott et al. 1999; Sewell et al. 1999), including the patient (Borg et al. 1999). Parkinsonism has been caused by an illicitly-produced synthetic opiate (MPTP) and news reports about this and other "designer drugs" may have led to confusion between MPTP and MDMA in the 1980s. It is worth noting that there are apparently no published reports confirming (or denying) any association between Parkinsonism and either amphetamine or methamphetamine use, both of which produce dopaminergic axonal damage in animals. Like many other comparisons across substance using populations, this study is limited by non-random assignment and retrospective design. In addition, the number of ecstasy + amphetamine users is smaller than that of the ecstasy users or non-ecstasy using controls. Others (see Semple et al, 1999) have described difficulties with using Beta-CIT as a measure of transporter binding, as it has affinity for both serotonin and dopamine transporter sites. Currently, there is no ready explanation for increased DA binding sites in sole ecstasy users, but it is possible that some ecstasy-mediated change in serotonergic function has increased the number of DA transporter sites.