To confirm previous results obtained in the overlapping study sample , we used linear regression analysis to test whether the log2 to the accumulated lifetime intake of ecstasy tablets correlated with SERT binding from the amygdala. Since effects of lifetime ecstasy intake on SERT become smaller with increasing abstinence from ecstasy , we controlled for this by also including the log2 to the number of days since the last use of ecstasy in the analysis. Also using a linear regression analysis, we tested whether the log2 to the number of days since the last use of ecstasy correlated with SERT binding from the amygdala . The possible difference between ecstasy users and controls in amygdala SERT binding was tested with an independent samples t-test.Functional data were preprocessed and analyzed using the statistical parametric mapping software package . Preprocessing included spatial realignment, co-registration to the anatomical image, segmentation, and normalization to the standard Montreal Neurological Institute template, and smoothing using a symmetric 6-mm Gaussian kernel. Subject-level models were constructed using five emotional face regressors , together with regressors modulating the events by their RT values. In addition, the subject-level models included a regressor for incorrect sex discrimination answers, and 24 nuisance regressors to correct for movement artifacts,sub irrigation cannabis including first- and secondorder movement parameters and spin history effects . T-contrasts comparing BOLD responses to emotional and neutral images were created for each participant.
These maps were used in group-level models assessing: the main effects of emotional face processing across all participants, differences in BOLD responses between ecstasy users and control subjects, correlations between BOLD responses and the log2 to the accumulated lifetime ecstasy intake in ecstasy users, correlations between brain activity and regional SERT binding in the amygdala in ecstasy users and control subjects, and correlations between BOLD responses and the log2 to the number of days since the last use of ecstasy in ecstasy users. 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. All activations are reported at peak level and are in standard MNI stereotactic space.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, 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,vertical grow 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, 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.Oleoylethanolamide is a fatty acid ethanolamide and a natural analog of the endogenous cannabinoid anandamide. There is no detailed research on the role of endocannabinoids in sleep in humans. Anandamide is known to engross slow-wave sleep by increasing adenosin levels in the forebrain of rodents . This is blocked by the cannabinoid CB1-receptor antagonist rimonabant.Murillo-Rodriguez et al. supposed oleoylethanolamide, which—unlike oleamide— activates the nuclear peroxisome proliferator-activated receptor-a to increase alertness and to participate in the regulation of waking. Up to now, elevated levels of oleoylethanolamide and anandamide were found in human microdialysates within the first day of ischemia as well as following neural injury or other stressors associated with necrosis. Furthermore, massive increases in FAEs and their precursor phospholipids have been found during the acute phase of stroke in the adult rat brain .