Drug use data used for correlations with MRI data and PET data were acquired in MRI and PET scan days, respectively. In and , one subject was excluded due to lack of precise information about the number of days of abstinence prior to the MRI investigation day, but the person was eligible for other analysis due to a negative urine sample. Our a priori region of interest was the amygdala. At first, we therefore restricted the correction for multiple comparisons to the amygdala ROI, as defined by the SPM anatomy toolbox ; p-values are provided as p . We set the significance level for activated voxels at p<0.05 corrected for multiple comparisons using the family-wise error correction . The entry threshold was set to p<0.001 uncorrected with an extent threshold of five contiguous voxels. Second, a whole-brain analysis was performed. The significance level for activated voxels was set at p<0.05 corrected for multiple comparisons . The threshold was the same as in the ROI analysis.To test whether associations between ecstasy usage and BOLD response were mediated by SERT, a path analysis was used to decompose the total effect of MDMA usage into direct and indirect effects. The direct effect of ecstasy exposure on the mean BOLD response across the amygdala is the conditional effect adjusting for SERT binding. The indirect effect of MDMA on the mean BOLD response is the difference in the ecstasy effect between a model, where SERT BP is controlled for compared to when it is not. This difference in effect is equivalent to the product between the effect that ecstasy has on SERT and the effect that SERT has on BOLD response. Linearity assumptions were assessed graphically. Standard errors of the indirect effect were calculated by the delta method and were validated by comparison with 95% quantiles from a parametric bootstrap.

To our knowledge,how to dry cannabis this is the first study to examine the effects of long-term ecstasy use on the neural responses to emotional face expressions. Relative to neutral face stimuli, main effects of emotional processing were found bilaterally in the amygdala, showing increased neural activity, especially in response to fearful and angry faces. This concurs well with a number of studies showing that viewing emotional faces, fearful faces in particular, activates the amygdala . While there was no ecstasy effect on task performance, ecstasy users did, as hypothesized, show higher amygdala activity with increased lifetime ecstasy use during angry face processing; that is, the more ecstasy tablets the ecstasy users had taken during their lifetime, the more activation they displayed in amygdala when watching angry faces. In the ecstasy user group, SERT binding correlated negatively with amygdala activity in response to angry faces. Non-significant statistical trends for activity during processing of angry and sad face processing suggested that amygdala activity waned with increasing time since the last intake of ecstasy. Neither the analyses of emotional expressions other than anger nor the whole-brain analysis revealed any significant results. Thus, our results support the hypothesis that long-term ecstasy use alters the neural basis of emotional face processing. This effect is dose-dependently related to lifetime consumption of ecstasy and appears to be reduced with increased time since last use. Interestingly, the linear relationship was consistently expressed for angry faces but not for other aversive facial expressions. This observation is in line with the results of Bedi et al. who found that acute MDMA intake alters the amygdala response to angry, but not fearful, facial expressions. The limited sample size of this study does, however, not allow us to conclude that there is not an effect of lifetime ecstasy intake on processing of other aversive facial expressions. While acute MDMA intake has been shown to diminish amygdala activation , we found the opposite effect in long-term ecstasy users. This supports our hypothesis that long-term ecstasy users are in a chronic, albeit potentially reversible, serotonin-depleted state and therefore in accordance with studies showing that serotonin depletion, as induced by acute tryptophan depletion, leads to elevated amygdala activity when processing negative facial expressions .

When including the lifetime amphetamine use in the model, the effect of the lifetime intake of ecstasy tablets on amygdala activity was no longer significant. This may be due to high correlation between ecstasy and amphetamine use . Since the lifetime amphetamine use in itself did not have a significant effect on amygdala activity during angry face processing, our interpretation of the results is that the effect of ecstasy use on emotional processing would be present also in the absence of amphetamine use. The present study was carried out on a sub-sample of our previous study sample of chronic ecstasy users , and we confirmed a negative correlation between SERT binding and accumulated ecstasy use. Hence, it could be speculated that our present fMRI results, showing a positive correlation between lifetime use of ecstasy tablets and left amygdala activity, was mediated by SERT density; that is, a larger lifetime intake of ecstasy tablets was associated with lower SERT binding levels , possibly leading to a higher degree of amygdala activation during angry face processing. In the ecstasy user group, SERT binding was indeed negatively correlated with amygdala reactivity to angry faces, which is in line with Rhodes et al. , showing a negative correlation in the left amygdala between SERT density and activity during emotional face processing. Post hoc mediation analysis did, however, not support the mediation hypothesis, although these results need to be interpreted with caution given the small sample size and hence the low statistical power. In short, our study suggests that there are functional consequences of a chronically depleted serotonin system as indexed by lowered SERT. Of note, an augmented amygdala response to angry faces has also been observed in mood disorders and could within a population with reduced serotonergic tone represent a sub-clinical vulnerability marker for such conditions. In line with several other studies , we have recently reported that recovery of sub-cortical—but not cortical—SERT availability takes place after termination of ecstasy use. Importantly, here, we found trends showing that days of abstinence from ecstasy correlated negatively with left amygdala activity during angry face processing and with right amygdala activity during sad face processing.

Since lifetime use of ecstasy tablets correlated positively with amygdala activity during angry face processing, the trend toward a negative correlation between days of abstinence from ecstasy and amygdala activity during angry face processing might be a potential sign of functional reversibility.There are limitations to the current study. Because of the cross-sectional nature of our study, it cannot be ruled out that the exaggerated amygdala response to angry faces and/or the low cerebral SERT among heavy ecstasy users represents preexisting traits associated with an increased preference for the use of ecstasy. We consider this less likely because, as discussed already, interventional animal studies have shown that administration of MDMA lowers cerebral SERT levels, and data from our group and others support the presence of an ecstasy dose–response relationship and recovery of SERT binding with abstinence from ecstasy . As for all investigations of the long-term consequences of illicit drug use,best way to dry cannabis especially the use of ecstasy, there will be uncertainties about the precision of the users’ reporting of drug use and actual content of substance taken. As explained in more details elsewhere , hair analysis for MDMA, use of systematic semi-structured questionnaires, and access to systematically acquired data on the content of Danish ecstasy pills in the period of data collection was employed to minimize these factors. MDMA has several effects on the serotonergic system, such as inhibiting tryptophan hydroxylase, the rate-limiting enzyme for serotonin synthesis, and serotonin degradation by monoamine oxidase B . It is possible that it is not the specific effect of MDMA on SERT, but other effects of prolonged MDMA use on the serotonergic neurotransmitter system that mediate the effect of MDMA use on brain responses to emotional faces. MDMA also has noradrenergic and dopaminergic effects that could affect amygdala activation. An additional limitation of our study is that we did not record hormonal contraception or menstrual-cycle phase for the two females in each group. These factors have been shown to affect face processing . However, we do not have any reason to suspect differences in contraceptive use or cycle phase between groups, why the lack of this information is considered as added noise, potentially reducing the power of the study. In conclusion, these results emphasize the important role of serotonergic neurotransmission in the amygdala for processing angry face expressions. We show that long-term ecstasy use has a dose-dependent effect on the amygdala response to angry faces. Importantly, on the basis of earlier work on amygdala responses to emotional face stimuli after manipulation of serotonin levels, this finding is in support of the hypothesis that recreational use of ecstasy can cause serotonin depletion. The decreased SERT binding among ecstasy users in the current as well as in several previous samples further supports this notion.

The fact that changes observed in the current study showed signs, although at a trend level, of reversibility with sustained abstinence is also in agreement with previous PET/SPECT imaging studies. With the recent focus on MDMA as a potential therapeutic tool in psychiatry , it is important to emphasize that heavy use of the same substance in a recreational setting is associated with functional and molecular—possibly reversible—changes related to serotonergic neurotransmission.Use of non-cigarette tobacco is increasing among youth. Past 30-day use of electronic cigarettes among US high school students recently rose substantially, more than doubling in two years, from 11.7% in 2017 to 27.5% in 20191,2. Similarly, the use of conventional smokeless tobacco in 2018 nearly equaled the prevalence of cigarette smoking among male US high school students 1 . Use of e-cigarettes and smokeless tobacco exemplify a larger trend, in which a broadening range of non-cigarette and non-combustible tobacco products threatens to erode public health gains in reducing youth tobacco use1 . Tobacco product characteristics, such as flavors, nicotine strength, e-cigarette device type , or smokeless tobacco cut , can signal properties of tobacco products to potential consumers, including youth. Perceived properties might relate to the taste, potency, or relative safety of the product. To the extent that specific tobacco product characteristics lead to youth viewing certain tobacco products as more appealing or associated with fewer risks, those characteristics represent plausible targets of regulation or other restrictions intended to reduce youth use. A combination of branding, product design, and real or perceived properties likely operate individually and collectively to shape youth tobacco related attitudes and decision-making. Research that identifies and quantifies the contributions of specific tobacco product characteristics is potentially appealing to regulators seeking to reduce youth use without outright bans on entire classes of products. Discrete choice methods stem from economic theory that consumer preferences are based on the multiple intrinsic characteristics of goods or products, and have recently been applied to tobacco control and tobacco regulatory science. Discrete choice experiments are designed to identify the independent contributions of component parts of a good or service to potential consumers’ overall preferences and/or beliefs. In surveys, participants are often asked to choose between two different products or scenarios, each representing a composite set of relevant attributes at varying levels , allowing quantification of how these characteristics independently contribute to respondents’ choices. Recent work has examined adults’ preferences related to water pipe tobacco and e-cigarettes, as well as youth e-cigarette preferences. In the latter study, youth were more likely to prefer e-cigarettes with non-tobacco flavors and less likely to choose products with Food and Drug Administration warning labels or ‘cigalike’ devices. The present study expands on previous discrete choice studies by including a community-based sample of youth, assessing both e-cigarettes and smokeless tobacco, and considering multiple specific perceived properties, such as danger and ease of use. The study objective is to evaluate the extent to which specific characteristics of e-cigarette and moist snuff smokeless tobacco products convey product qualities to youth, especially those perceived qualities that may lead to greater youth appeal and product use. Such product characteristics are plausible targets of potential FDA regulation or local policy designed to reduce youth tobacco use.