The current prospective study investigated the effect of MDMA on cognitive performances in incipient MDMA users over the course of a 2-year period. The results of the first follow-up assessment showed significant deficits in MDMA users in visual paired association learning after a period of 1 year (Wagner et al., 2013). Based on those results it was hypothesized that the sustained use of MDMA over a period of 2 years would lead to further decline in cognitive performance, especially in heavy MDMA users. The same cognitive test battery which was used during the first assessment, including tests of the subject's attention and information processing speed, memory, working memory and executive functioning, were also used during the second assessment. Out of the initial 149 subjects who participated in the first assessment, 96 subjects participated in the second assessment after 2 years. The analyzed data showed no differences concerning possible confounding variables, including the subjects' cannabis use, alcohol use, and their current medical-treatment, participation in sports, diet, satisfaction with sleep, and feelings of subjective well-being. However, the groups did differ with regard to the subjects' mean age, their use of cocaine, solvents, inhalants, and amphetamines, as well as differences with regard to the subjects' general intelligence. Therefore, these variables were considered as possible confounding variables.

The repeated measures ANOVA addressing episodic memory indicated a specific effect regarding paired association learning and MDMA use. A marginally significant interaction effect of group and time for both immediate and delayed recall memory subtests from the Lern- und Gedächtnistest (LGT; Bäumler, 1974) was suggested. The analyzed data showed differences in performance among the groups between the baseline and the first assessment for the immediate recall memory test. Based on these results, it seems that the non-user and moderate-user groups increased in performance between the baseline and the first assessment. Furthermore, it appears that the performance of the heavy-user group decreased in the first follow-up period. These results are in line with the already published findings from the study of Wagner et al. (2013) that demonstrated a dose-related neurotoxic effect of MDMA on visual paired associates learning. However, it seems that with respect to the second follow-up period, this effect no longer occurred. Based on the results of the second follow-up assessment, it appears that the performance of all groups improves with regard to the immediate recall test after a period of 2 years. The findings of the current study for the first follow-up period are congruent with previous results from other investigations that found deficits in associative learning with poorer immediate and delayed recall abilities in MDMA users compared to non-users (Daumann et al., 2005; Montgomery et al., 2005; Quednow et al., 2006; Parrott, 2013a). However, as already mentioned, the analyzed data showed a similar improvement in performance for the immediate recall test for all groups between the first assessment and the second assessment. These results are consistent with the findings of de Sola Llopis et al. (2008). In their study, they also found an improvement in the performance of MDMA users after a follow-up period of 2 years. However, the overall performance of the MDMA users was still lower when compared to the non-users. They postulated that the subjects' improvement in performance could be due to practice effects through their repeated exposure to the test material. Therefore, it may also be possible that the participants in the current study performed better because they were better acquainted with the test material. The use of parallel versions from neurocognitive tests could be a solution to prevent such practice effects in future studies. Perhaps another explanation for the improvement in performance in the heavy-user group could be a decrease in their MDMA use between the first and second assessments (see Figure 1). Similar results were found in other longitudinal studies: Heavy-users improved their performance on an immediate recall test in which the average use of MDMA had decreased between the first and second assessments (Thomasius et al., 2003; Gouzoulis-Mayfrank et al., 2005). A last explanation for the marginal significant effect of MDMA-use between the groups could be that the non-user group also could have been impaired in cognitive performance before participating in this study due to their little experience with MDMA. Because even a small cumulative dose of MDMA, is associated with a decline in cognitive performance (Schilt et al., 2007). For the future, it would be interesting to add a group without any drug experience to rule out preexisting cognitive impairments due to MDMA-use in the control-group.

With respect to performance on the delayed recall test, it seems that the groups changed in performance between the baseline assessment and the second assessment. It appears that an improvement in performance occurred among the non-users and moderate-users, but not in the heavy-user group. More precisely, a performance decline in heavy-users took place between baseline and the first assessment, with no further deterioration in performance between the first and the second assessments. This could indicate a dose-related neurotoxic effect of MDMA within the first follow-up period—as was already found in the study of Wagner et al. (2013)—but not for the second-follow-up period. Therefore, it seems that continuing MDMA, use did not lead to further decrease in performance on the delayed recall test after 2 years, a result which corresponds with the previous findings of Gouzoulis-Mayfrank et al. (2005). Furthermore, no significant changes in performance within the 2-year follow-up period were found, results which are similar to the findings of Brown et al. (2010). Brown and colleagues also established significant effects on association learning tasks with regard to MDMA users and non-users, but no differences concerning other memory tasks. However, it has not been established whether these other measures of cognitive performance may also decrease over time with prolonged use of MDMA over the subject's lifetime. Therefore, longitudinal studies including follow-up assessments over a period of several years are needed.

As already established in our analysis of the data from the first follow-up period (Wagner et al., 2013), a significant intercorrelation was found between the subjects' use of MDMA and amphetamine in the second follow-up period. It still remains unclear whether or not the impairments in visual pair associates learning can be ascribed to the subjects' use of MDMA alone or whether it is also due to their concomitant use of amphetamine. However, in their study, Gouzoulis-Mayfrank et al. (2003) attributed the subjects' performance on the same immediate and delayed recall test to their use of MDMA rather than to their polydrug-use of MDMA and amphetamine. More research concerning memory deficits in humans with respect to the neurotoxic effects of amphetamine are needed. Furthermore, there was also a significant intercorrelation between the subjects' use of MDMA and cocaine. However, the confounding effect of cocaine did not fully account for the differences between the groups on the visual paired association task. Even after excluding cocaine as a covariate, the results on the visual paired association task remained marginal significant. Even though dose-related neurotoxic effects of MDMA have been established, among incipient MDMA users in their first year of regular use, it seems that this effect no longer exists or at least diminishes after a second year of MDMA use. In this respect, the findings of the current study could be interpreted as contradicting the MDMA-related memory decline in continuing users. Otherwise, a further decrease in performance within the second year, would be expected. Another explanation could be that the neurotoxic potential of MDMA displays its effect on memory at the beginning of regular MDMA use in novice users without an exponential deterioration through continued use. In a recent study, Taurah et al. (2013), argued that prolonged use of MDMA is a weak predictor for memory deficits. Instead, they argue that it is a combination of MDMA use with other drugs that seems crucial for memory deficits in recent and past MDMA users (Taurah et al., 2013). However, most studies reveal that prolonged lifetime use of MDMA leads to memory deficits in MDMA users (Parrott et al., 2002; Parrott, 2013a). Moreover, the performance of MDMA users gets worse when they are confronted with supraspan tasks, where their memory systems become overloaded beyond their normal capacities (Parrott, 2013a). In this respect, the LGT 3 is such a supraspan task (Bäumler, 1974). This test has been shown to be sensitive in detecting performance deficits in MDMA users (Gouzoulis-Mayfrank et al., 2003).

The recent literature shows that impairment in MDMA users can be found in almost all cognitive domains (Parrott, 2013b). In their study, Quednow et al. (2006) found deficits in verbal declarative memory in MDMA users compared to non-users. The subjects' verbal declarative memory was assessed with the Rey Auditory Verbal Learning Test (RAVLT; Heubrock, 1992), which was likewise used in our current study as an assessment tool. However, as compared to our study, Quednow et al. (2006) looked at a group of former MDMA users with a longer lifetime and higher cumulative use of MDMA. Regarding to the longer lifetime use and higher cumulative use of MDMA, it might be possible that the performance of MDMA users on the declarative memory task in the current study may continue to decline if they continue to use MDMA in the following years.

The findings on the tests of executive functioning are consistent with the previous study of Halpern et al. (2011). They also found no differences on tests of executive functioning (Trail-Making Test A and B), except with the digit span test backwards from the WAIS 3. Although MDMA users showed impairments on the digit span test backwards, significant differences were only established between the non-user and moderate-user groups, and not between the heavy-user and non-user groups. This is inconsistent with the dose-related neurotoxic effect hypothesis of MDMA. Moreover, no differences between the groups were found in performance on the Stroop Tasks, a result which corresponds with our findings. In summary, the results of the current study indicate a MDMA-related performance decrease in heavy MDMA users on a visual paired association task. A reason for the performance decrease in heavy MDMA users on visual paired association tasks could be hippocampal dysfunction, because the hippocampus plays a crucial role in learning processes and memory, and it is particularly vulnerable to the neurotoxic effects of MDMA (Hatzidimitriou et al., 1999; Daumann et al., 2005; Brown et al., 2010; Pergola and Suchan, 2013). Interestingly, in a recent prospective fMRI study by Becker et al. (2013), continuing MDMA users showed, in contrast to abstinent MDMA users, a decrease in encoding-related activity in the left parahippocampus during an association memory task. However, no differences in performance on this task were found between the groups. This suggests that MDMA users are still able to perform in a normal range despite an MDMA-specific dysfunction of the parahippocampus. One possible explanation could be that MDMA users do not want to believe that their results on memory performance tests could be negatively affected by their MDMA use. Consequently, they may try harder to reach normal results and thereby compensate for the dysfunction of their parahippocampus. In a recent functional near-infrared spectroscopy (fNIRS) study by Roberts, Wetherell, Fisk and Montgomery (Roberts et al., 2015) they found also no behavioral performance differences on a high mental workload task between MDMA-users and non-users. However, in comparison to non-users, MDMA-users showed an aberrant neuronal functioning in the dorsolateral prefrontal cortex (DLPFC), suggesting that MDMA-users have neurological impairments in these networks. Also in Electroencephalography (EEG) studies, neuronal changes in MDMA-users are found in the absence of a decrease in behavioral performances (Burgess et al., 2011; Roberts et al., 2013). Therefore, it seems that performance on cognitive tasks between MDMA-users and non-users alone are not sensitive enough to measure cognitive impairments induced by MDMA-use.

Despite the empirical evidence found in animal studies for the neurotoxic effect of MDMA in the hippocampus and other brain regions, the underlying molecular mechanisms have not been fully clarified. There are different factors that seem to contribute to the neurotoxic effect of MDMA. For example, heightened ambient temperature influences the metabolites of MDMA which are associated with the induction of 5-HT toxicity (Moon et al., 2008; Capela et al., 2009) and decreases the activity of tryptophan hydroxylase (TPH), an enzyme that is required for the synthesis of 5-HT (Moon et al., 2008). Oxidative stress induced through MDMA administration leads to an increase in reactive oxygen species (ROS) and decreases the antioxidant defense of cells, which leads to cell damage and mitochondrial dysfunction (Moon et al., 2008; Capela et al., 2009). As a consequence of an increased monoamine oxidase metabolism of monoamine neurotransmitters (dopamine, noradrenaline, adrenaline, and serotonin), more toxic metabolites can be found in cells. This subsequently leads to cell damage.

However, the underlying mechanism of MDMA, which can cause neurodegeneration and changes in the serotonin system, is still difficult to identify in humans (Green et al., 2012). Most evidence regarding the underlying molecular mechanisms came from studies using laboratory species. This creates difficulties in translating such information to human MDMA users (Green et al., 2012). Furthermore, there are findings which are inconsistent with the neurodegeneration hypothesis. For example, serotonergic fibers have been found to remain in rats even after MDMA administration and indeed to recover from the neurotoxic effects of MDMA (Hatzidimitriou et al., 1999; Kish et al., 2010; Biezonski and Meyer, 2011). With respect to the empirical evidence of the neurodegenerating effects of MDMA on human users, it is therefore important to consider that serotonergic deficits are not necessarily the result of axonal damage. Therefore, Biezonski and Meyer (2011) noted that “MDMA can certainly be considered as neurotoxic in terms of causing serotonergic dysfunction.”

It should be noted that the current study is not free of certain methodological limitations, as we have already discussed in our previous paper. Like other open-trial studies, this study was not experimental, and, therefore, it provides no causal interpretation of the effect of MDMA on brain regions which are associated with specific cognitive performance. This study began with a large sample of 149 subjects. However, only 96 subjects remained for the second follow-up assessment, and most of these were non-users or moderate-users. Therefore, the subjects were unevenly distributed over the groups, with a small sample size of 10 subjects in the heavy-user group. In addition, the remaining 10 participants in the heavy-users group might be actually highly motivated and that might constitute a bias in terms of performance and might partially explain the results found in the present study. Furthermore, in the recent study MDMA-users consumed cannabis almost twice as much as non-users. Even though the concomitant use of cannabis did not differ significantly between the groups, a possible confound of cannabis cannot be completely ruled out. Some investigators have suggested a possible neuroprotective effect of Tetrahydrocannabinol (THC) against MDMA neurotoxicity in laboratory animals (Tourino et al., 2010; Lopez-Rodriguez et al., 2014). In this regard, cannabis might be preventing a performance decline in moderate-users but not in heavy-users due to their excessive use of MDMA. In a fMRI-study of Daumann et al. (2003) pure MDMA-users presented lower activations of different brain regions during a working-memory task in comparison to polyvalent- and non-users, suggesting a neuroprotective effect of cannabis against MDMA neurotoxicity.

Furthermore, in the recent study a significant difference between the groups in their use of cocaine was found. Although, no interaction effect of MDMA and cocaine was found on cognitive functioning, a confounding effect of cocaine cannot completely ruled out, too. Investigators found cognitive impairments on different cognitive domains in cocaine-users (Potvin et al., 2014). Thus, in the recent study, the impairments in visual pair associates learning cannot only be attributed to MDMA alone but also to the concomitant use of cocaine and cannabis, which is a common limitation to prospective MDMA-studies. Further limitations, which are common to all longitudinal, MDMA research in humans have already been intensively discussed. For these, we refer the reader to our already published study (Wagner et al., 2013).

In conclusion, the already published findings of the first follow-up period seem to be consistent with the findings of the current study. In this study, we found an MDMA-related decline in memory performance on a visual paired association test in the first follow-up period in heavy MDMA users, suggesting an MDMA-related dysfunction of the hippocampal regions. However, no further deterioration was observed in the second follow-up period, that could be due to methodological limitations of the recent study, which are inherent to open-trial studies and mentioned below.