Purpose: Brain imaging: To investigate the effects of moderate and heavy ecstasy use, as well as prolonged abstinence from ecstasy use, on the density of Beta-CIT binding to serotonin transporter, including an examination of possible gender differences in ecstasy-induced changes in Beta-CIT binding.

Design: Non-experimental (retrospective) 2 x 4 between-groups design, with gender (men or women) as one between-group factor and extent of ecstasy use (current moderate ecstasy use, current heavy ecstasy use, former heavy ecstasy use, or non-user controls) serving as the other between-group factor. Beta-CIT binding ratio served as a dependent measure. All participants received SPECT scans.

Subjects: 15 moderate ecstasy users, 23 heavy ecstasy users, 16 former ecstasy users and 15 non-user controls residing in the Amsterdam (Netherlands) area, and recruited via flyers distributed at locations associated with the dance event (rave) scene. Matching - All 4 groups were matched by gender, age, and approximately matched on estimated verbal intelligence (via DART) and occurrence of depression. Men and women were matched on estimated verbal IQ and approximately matched on occurrence of depression.

Criteria for Inclusion, Moderate Ecstasy Users - Having taken 1 to 50 ecstasy tablets in a lifetime. Heavy Ecstasy Users - Having taken at least 50 tablets in a lifetime. Former ("Ex") Ecstasy Users - Having taken at least 50 tablets in a lifetime, but having abstained from further ecstasy use for at least 1 year prior to study date. Non-users - Never having used ecstasy, but use of other drugs permitted. All Groups - Healthy, being between the ages of 18 and 45, as apparently indicated through medical examination and no past or current major psychiatric illness, no use of medication with affinity for serotonin transporter, not pregnant, and abstinent from all psychoactive drugs for at least 3 wks before study day, with abstinence verified through urinary drug screen.

Drug Use Parameters - Moderate Ecstasy Users - Moderate ecstasy users reported an average lifetime use of 28.6 +/- 17.8 tablets, with an average dose per use of 1.4 +/- 0.5 tablets. Average duration of ecstasy use, in months, was 49.2 +/- 31.2 months, with no information provided concerning frequency of use. Time since last use for moderate users, in days, was reported at 108 +/- 177 days prior to study day, with abstinence verified through self-report and urinary screen on study day. Heavy Ecstasy users reported average lifetime use of 530 +/- 621.1 tablets, at an average dose of 2.2 +/- 0.7 tablets per use. Average duration of use, in months, was 55.2 +/- 25.2 months, with no information presented about frequency of use per month. Time since last use, in days, was an average of 69 +/- 72 days, with abstinence verified via self-report (and via urinary screening on study day). Former ecstasy users - reported average lifetime use of 268 +/- 614.3 tablets, at an average dose of 2.1 +/- 1 tablets per use. Average duration of use, in months, was 55.2 +/- 31.2 months, with no information presented about frequency of use per month. Time since last use, in days, was an average of 870 +/- 612 days (approx. 2.4 years), with abstinence verified via self-report and via urinary screening on study day. Other Drugs - Average alcohol consumption per week was 13.04 +/- 10.16 units for moderate ecstasy users, 9.56 +/- 5.95 units for heavy ecstasy users, 6.2 +/- 4.65 units for former ecstasy users and 10.36 +/- 9.92 units for non-user controls. Average tobacco use per day was 10.36 +/- 7.58 cigarettes for moderate ecstasy users, 9.34 +/- 10.18 cigarettes for heavy ecstasy users, 12.55 +/- 8.55 cigarettes for former ecstasy users and 9.93 +/- 4.79 cigarettes for non-users. Average cannabis use (joints per last 3 mo) was 53.58 +/- 24.54 joints for moderate users, 81.64 +/- 128.56 joints for heavy ecstasy users, 134.7 +/- 239.85 joints for former ecstasy users and 3.47 +/- 2.95 joints for non-users. Average amphetamine consumption (times in last 3 mo) was 0.24 +/- 0.48 in moderate ecstasy users (men only, actual figures 0.4 +/- 0.8), 3.7 +/- 6.49 times for heavy ecstasy users, with no amphetamine use reported for former ecstasy users or non-users. Average amphetamine dose per use in last 3 mo (g) was 0.14 +/- 0.10 g for moderate ecstasy users, 0.35 +/- 0.3 g for heavy ecstasy users, 0.85 +/- 0.6 g for former users, and non-users did not use amphetamine. Average cocaine use, in times per last 3 mo, was 0.76 +/- 0.43 times in moderate ecstasy users (only men reported use, actual figures 1.2 +/- 1.1 uses), and 4.3 +/- 3.09 times for heavy ecstasy users. Former ecstasy users and non-users did not report any cocaine use during this interval. In a previous paper with similar samples, heavy ecstasy users and former ecstasy users both reported some use of LSD and psilocybin-containing mushrooms.

Group Demographics and Matched Variables - Men and women in each group were matched for estimated verbal IQ. Drug use groups were matched for gender and age, and approximately matched for estimated verbal IQ and incidence of depression. Gender, as M/F ratio - Moderate users, 9/6, heavy users, 12/11, former users, 8/8 and non-users, 7/8. Age - Average age for moderate users was 24.44 +/- 5.62 years, 26.10 +/- 5.09 years for heavy users, 25.25 +/- 5.45 years for former users and 26.1 +/- 3.91 years in non-users. Men were older than women across all groups. Estimated Verbal IQ - Average estimated verbal IQ, as assessed via DART IQ, was 111.6 +/- 10.14 for moderate users, 105.28 +/- 8.71 for heavy users, 103.95 +/- 9.75 for former users and 105.87 +/- 6.85 for non-users; moderate users had higher estimated verbal IQ than all other groups. Depressive Symptoms - 3 moderate users (1 M, 2 F), 4 heavy users (2 M, 2 F), 4 former users (1 M, 3 F) and 2 non-users (0 M, 2 F) had current signs of depression. Education - No information is provided concerning years of education attained within drug-use group or across gender.

Measures: Estimated Verbal IQ - Assessed via DART (Dutch version of NART), used for matching only.

Imaging - SPECT scans were performed 4 h after injection of 3.8 mCi (140 MBq) [123I]-Beta-CIT. An investigator blind to condition constructed a standard template with regions of interest. Selected ROIs were in frontal, temporal, parieto-occipital, and occipital areas, as well as in thalamus and midbrain. Another template with cerebellum selected as comparison area due to presumed low serotonin transporter binding. [123I]-Beta-CIT binding ratios were calculated by division of ROI binding with binding in the cerebellum.

Depressive symptoms - This was assessed by an unspecified dichotomous measure of self-reported depression, though individuals with clinical depression were apparently excluded from study.

Analyses: A mixed linear model regression was used to compare 123I]-Beta-CIT binding ratios at six specific brain sites across drug use parameters (non-users, moderate users, heavy users and former heavy users) and gender (men or women). Within-individual binding ratios were compared using covariance within model. The model also contained interaction terms for group and gender, and included occurrence of depression (dichotomous) and age as additional (covariant) factors.

Results - Significant Differences: Overall [123I]-Beta-CIT binding ratios were significantly lower in heavy female users, compared with female non-user controls and with ex-MDMA users (non-users = moderate users = ex-users > heavy users). [123I]-Beta-CIT binding within brain regions tended to be highly correlated with each other. Binding ratios were significantly lower in frontal, temporal, parieto-occipital, midbrain and thalamus in female heavy ecstasy users, when compared with female non-user controls and with female former ecstasy users. There was a trend for female former ecstasy users to have lower [123I]-Beta-CIT binding ratios in parieto-occipital areas when compared with female non-users. (There was a trend for female former uses to have lower binding in occipital areas, but it did not reach significance). There was a significant correlation between log transformed lifetime use of ecstasy and [123I]-Beta-CIT binding ratio in women only.

Results - No Effects Found: [123I]-Beta-CIT ratio binding did not differ in male and female control subjects. Male and female moderate ecstasy users did not differ from controls in overall [123I]-Beta-CIT binding ratios. There were no significant differences in [123I]-Beta-CIT binding ratios due to drug use parameters for any of the male participants (non-users = moderate users = heavy users = ex-users). When compared with non-users, there were no differences in [123I]-Beta-CIT binding ratios in men in all groups and women moderate and former users in the following areas: midbrain, thalamus, frontal, temporal and occipital areas. Participant's age or presence (versus absence) of self-reported current depression did not affect levels of [123I]-Beta-CIT binding ratios. There was no significant correlation between l[123I]-Beta-CIT binding ratios in any brain area and log-transformed lifetime use of ecstasy for men.

Overall Effects: An imaging study comparing brain serotonin transporter site density by performing SPECT scans with [123I]-Beta-CIT found gender differences in binding in ecstasy users. Specifically, women reporting lifetime use of at least 50 ecstasy tablets had lower Beta-CIT binding ratios compared with women reporting no use of ecstasy, whereas ecstasy use did not change levels of [123I]-Beta-CIT binding in men. After abstinence from ecstasy for a year or more, women with a lifetime use of at least 50 tablets no longer had lower [123I]-Beta-CIT binding ratios than female non-user controls. Men and women reporting no use of ecstasy did not have different [123I]-Beta-CIT binding ratios, and neither did men and women reporting lifetime use of under 50 ecstasy tablets. Participants' age and presence of current depression did not affect differences in [123I]-Beta-CIT binding across drug use history, and participant's age also had no impact on [123I]-Beta-CIT binding ratios in either gender. Reported lifetime use of ecstasy was related to binding ratios for women, but not for men, with higher lifetime use associated with lower [123I]-Beta-CIT binding. When compared across specific areas, women, but not men, reporting use of 50 or more ecstasy tablets had lower ligand binding in all brain areas examined, both when compared with female non-user controls and when compared with women former ecstasy users. However, there was a trend for female (but not male) former ecstasy users to have lower [123I]-Beta-CIT binding sites in parieto-occipital areas and perhaps occipital areas. Overall, the report suggests that women may be more vulnerable to the effects of ecstasy on brain serotonin transporter sites, but that these long-term effects may be reversible and dose-dependent in terms of lifetime exposures.

Comments: This report is the second of two studies suggesting an increased sensitivity to ecstasy-related harm or changes to the serotonergic system in women. McCann and colleagues also found lower 5-HIAA in female, but not male, ecstasy users (McCann et al, 1994). Reneman is also the second researcher to use [123I]-Beta-CIT binding in human studies, with Semple previously publishing results from [123I]-Beta-CIT SPECT scans, albeit with a different procedure (Semple et al, 1999). In contrast with the findings reported here, Semple and colleagues found that lower [123I]-Beta-CIT binding in men who had used ecstasy compared with male non-users. This paper is now one of at least two reports suggesting that repeated ecstasy use produces effects only after a "threshold" number of exposures(see Vollenweider, 2001). Since Beta-CIT binds both to serotonin and dopamine transporters (see discussion in Semple et al, 1999), it is possible that all studies using this measure may be affected by differences in dopamine transporter sites as well as serotonin transporter sites. Like many other retrospective studies, the findings in this report could still be attributable to increased use of other drugs (specifically cannabis and amphetamines) in heavy ecstasy users, and with the self-selected nature of the samples. It is also possible that a more sensitive (continuous) measure of depression might have detected differences in Beta-CIT binding due to depression. In contrast with the findings reported here, there are no reports of gender differences in the effects of ecstasy on mood or cognitive function, save perhaps findings reporting less vulnerability to reduced verbal memory after ecstasy use in women compared with men reported in Bolla (1998).