MDMA’s Neurotoxicity: What the Research Shows & How to Reduce Your Risk (Detailed)

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The neurotoxicity of 3,4-methylenedioxymethamphetamine (MDMA) has been a controversial topic for decades. Even though a lot of research has been published, we still don’t fully understand what the harm potential is. As it currently stands, there’s good evidence for serotonergic dysfunction (not necessarily neurotoxicity) with heavy use.

A lot of problems exist with the literature. Despite that, animal and human results suggest heavy use (total use and/or frequency) is likely connected to some form of serotonergic disruption and potentially toxicity.

There’s reason to believe certain behaviors and supplements may be able to reduce the risk.

Contents

Main Claims
Problems & Limitations
Animal Evidence
Human Evidence
Potential Routes of Dysfunction
Drugs That May Enhance The Risk
Drugs That May Decrease The Risk
Other Factors
References

Main Claims

MDMA appears capable of resulting in a long-term reduction of serotonin, reduction of 5-HIAA (a serotonin metabolite), reduced SERT availability, and cognitive impairment. It may also cause an extended decline in tryptophan hydroxylase.

Some studies suggest that even if serotonergic markers return to normal, cognition could be impaired for years.

Although it’s been claimed MDMA use could result in mood disorders, there’s not much evidence to support this, particularly in reasonable users.

In the acute stage, the drug may damage serotonin axons, with the terminals appearing swollen and fragmented. From there, one hypothesis (the neurodegeneration hypothesis) claims the terminals are destroyed. This process has been referred to as an “axotomy.”

Doubts have been raised regarding the true neurotoxic nature of MDMA. It’s possible that the serotonergic changes we’ve seen are actually due to lasting protein and/or gene effects, rather than actual degeneration of neurons. Certain studies have also shown a recovery of serotonergic markers, at least in some brain regions.

Problems & Limitations

The research has long been complicated and controversial. Often the animals are given far too much MDMA and humans are studied after using excessive amounts of the substance.

It’s also possible there’s a publication bias towards negative effects and if only minor/moderate negatives are found in a study, the authors may provide a much more dramatic interpretation of the results.

Non-representative samples

One of the greatest issues with the human research is that the majority of serotonergic/cognition studies involve people who self-report very heavy levels of use. Much of the time they’ve been taking the drug more than once per month and have a cumulative use of hundreds of tablets.

Dosing

The bulk of the evidence currently supports giving a larger mg/kg dose to animals. However, the doses used in some animal studies go well beyond imitating human exposure and their results are therefore of questionable value.

Papers

(Green, 2009) – Comparing rats and humans

Humans received MDMA orally
Rats received MDMA via oral, IP, and SC
- The data between routes was combined since a similar Cmax was reached despite the route.
Human metabolism
- 1 mg/kg
  - Cmax – 0.10 mg/L
- 2 mg/kg
  - Cmax – 0.45 mg/L
- Auto-inhibition of metabolism exists, meaning the pharmacokinetics of MDMA change nonlinearly.
  - This likely contributes to findings of over 1 mg/L in some hospital cases.
Rodent metabolism
- The 2 mg/kg Cmax in humans is only reached when rats are given around 7 mg/kg.
Comparing functional measures
- 0.6°C core temperature increase using data from (Farre, 2007) and (Colado, 1995)
  - 1.4 mg/kg oral in humans and 5 mg/kg IP in rats
Author interpretation
- “The results presented here make clear that the “direct extrapolation of dose” technique is naive and with no credibility.”

(Mueller, 2009) – Comparing squirrel monkeys and humans

Methodology
- 6 squirrel monkeys
  - Received different doses orally, around 6 weeks apart, which were calculated to be equivalent to 0.4, 0.8, 1.6, and 2.8 mg/kg in humans.
- 9 humans
  - Received either placebo, 1.0, or 1.6 mg/kg with each administration separated by at least a week.
Results
- For a pharmacokinetic comparison, 1.6 mg/kg in humans and 2.8 mg/kg in monkeys was picked due to comparable Cmax values.
  - The half-life was much shorter in monkeys. Monkeys also showed around half the AUC of humans.
  - The Tmax was also shorter in monkeys.
  - Humans (1.6 mg/kg)
    - Cmax: 254.7 ng/mL
    - AUC: 3070.6 ng x h/mL
    - Half-life: 8.4 hours
  - Monkeys (2.8 mg/kg)
    - Cmax: 312.7 ng/mL
    - AUC: 1314.2 ng x h/mL
    - Half-life: 2.1 hours
- Another comparison looked at concentrations of MDMA and its metabolites (HHMA, HMMA, and MDA).
  - This used interspecies dose-scaling, resulting in 1.6 mg/kg for humans and 5.7 mg/kg for monkeys.
    - This produces similar AUC values, but a higher Cmax in monkeys.
  - A similar proportion of MDMA to HHMA was seen between species, while monkeys generated more HMMA.
  - In both species, the fraction of MDMA converted to MDA was under 5%.
  - The proportion of MDMA to metabolites varied by dose.
    - At the lowest dose in monkeys, the Cmax and AUC of HMMA exceeded that of MDMA.
    - In humans, the Cmax of HMMA after 1.0 mg/kg was higher than MDMA, but not to a significant degree.
    - At higher doses, the amount of MDMA exceeded that of HMMA and HHMA in both species.
  - Results
    - Humans (1.6 mg/kg)
      - AUC: 3070 ng x h/mL
      - Cmax: 254.7 ng/mL
    - Monkeys (5.7 mg/kg)
      - AUC: 3866 ng x h/mL
      - Cmax: 723.6 ng/mL
Author interpretation
- “Because of the shorter half-life of MDMA in the squirrel monkey, it is impossible to achieve comparable Cmax and AUC values simultaneously in the 2 species.”
- It appears CYP activity for MDMA > HHMA is similar in the species, while COMT activity is higher in monkeys.
- Since the HHMA pharmacokinetics remained similar despite increasing the dose of MDMA, there is evidence of nonlinear pharmacokinetics.
  - As the dose of MDMA increases, MDMA levels increased and exceeded those of HHMA and HMMA, which were remaining constant.
  - Once plasma MDMA was at 125 – 150 ng/mL, the HHMA and HMMA levels remained constant despite boosting the MDMA dose.

Animal Evidence

(Scheffel, 1998) – MDMA caused lasting serotonergic changes in a baboon for up to 13 months

Methodology
- 1 baboon
- Control baseline readings taken over a one month period.
- Comparative readings at 13 days, 19, days, 40 days, 9 months, and 12 months after MDMA
- Studied using (+)McN5652 binding
Dose
- 5 mg/kg (SC) twice per day for 4 days (over 8 hours between doses)
Results
- 13 days
  - Significant reduction in radioactivity accumulation from McN5652
  - Highest tracer accumulation in the hypothalamus
  - Intermediate accumulation in the thalamus and striatum
  - Lowest accumulation in cortical regions and the cerebellum
- 9 months
  - Significant recovery of binding in some parts of the brain, but not others.
  - Tracer concentrations increased back to pretreatment levels in the midbrain and hypothalamus.
  - All cortical regions still had reductions in binding.
- 13 months
  - Binding remained down in the frontal cortex (-62%), parietal cortex (-78%), and occipital cortex (-73%).
  - Binding actually increased in the pons (85% over control), midbrain (101% over control), and hypothalamus (123% over control).
Interpretation from authors
- MDMA can cause lasting SERT declines in the baboon, but it also appears to cause a reorganization of ascending serotonin axon projections.
  - Distal targets like the dorsal neocortex remain denervated while proximal targets like the hypothalamus become hyperinnervated.

(Hatzidimitriou, 1999) – Altered serotonin innervation patterns in the forebrain of monkeys for up to 7 years

Methodology
- 10 squirrel monkeys
- Animals were killed at either 2 weeks or 6-7 years post-treatment
- Before the animals were killed, they received 10 mg/kg IP of trans-2-phenylcyclopropylamine, an MAOI.
Dose
- 5 mg/kg SC twice daily for 4 days
Results
- Control animals showed no significant effect of age on serotonin axon density.
- Neocortex
  - 2 weeks
    - 83-95% reduction in serotonin axon density in all examined areas of the cerebral cortex.
  - 7 years
    - Serotonin axon density was still decreased in all neocortical regions, though there was some recovery.
    - All neocortical cell layers appeared equally denervated overall.
    - In the primary visual cortex and occipital cortical regions, the largest reduction in fibers was in layer IVC.
      - Decline in density in the IVC made it a no longer discernible layer.
- Hippocampus
  - CA1 – CA3
    - 2 weeks
      - Reduction in axon density in all three fields.
    - 7 years
      - Significant declines still in CA1 and CA2, with partial recovery in CA3.
  - Dentate gyrus
    - Similar overall denervation between groups, though different pattern of serotonergic innervation.
    - 2 weeks
      - Dentate hilus had moderate density of spared axons compared to the severely denervated molecular layer.
    - 7 years
      - Hilus had fewer axonal fibers remaining.
      - While molecular layer had some recovery in density.
  - Subiculum
    - 2 weeks
      - 93% decline in axon density, with a small number of remaining axons in the molecular layer.
    - 7 years
      - Out of every brain region in the study, this area has the great (80%) remaining denervation.
- Striatum
  - Caudate
    - 2 weeks
      - All regions have substantial reduction in density.
    - 7 years
      - Partial recovery
  - Putamen
    - 2 weeks
      - Reduction in axon density.
    - 7 years
      - Partial recovery
  - Globus pallidus
    - 2 weeks
      - Significant reduction in axon density throughout the length of the globus pallidus, with a more profound reduction in external segment.
    - 7 years
      - Both segments show evidence of hyperinnervation, especially in external segment.
      - Most noticeable along the medial aspect of the globus pallidus adjacent to the internal capsule.
- Amygdaloid complex
  - 2 weeks
    - 62-95% reduction in serotonin immunoreactivity among the six nuclei examined.
    - Lateral nucleus has the greatest reduction and is one of the most serverely lesioned regions overall at 2 weeks.
  - 7 years
    - 16-57% reduction in immunoreactivity in the amygdala, with the exception of the central nucleus, which has a full recovery.
- Hypothalamus
  - 2 weeks
    - Slight to moderate reduction in axons, depending on the location of the particular hypothalamic nucleus relative to the medial forebrain bundle.
    - Lateral nucleus was largely unaffected (-6%).
    - Ventromedial nucleus had 31% reduction, dorsomedial nucleus had 39% reduction, and dorsal nucleus had 55% reduction
  - 7 years
    - Complete recovery in all nuclei.
- Thalamus
  - 2 weeks
    - 81-94% reduction in axon density in the seven nuclei examined.
  - 7 years
    - Most nuclei had complete recovery.
    - Dorsoanterior nucleus and the lateral-dorsal nucleus are the main exceptions, with both having 44% reduction in axon terminals.
- Raphe nuclei
  - No apparent loss of cell bodies or change in serotonin or tryptophan hydroxylase immunoreactivity in the dorsal, median, or B9 cell groups at 2 weeks or 7 years.
- Axon morphology
  - At 7 years, there was a shift to fine axon type in the hyperinnervation regions, namely the globus pallidus.
- Catecholaminergic fibers
  - No difference from control in density of axons.
Author interpretation
- MDMA leads to lasting changes in axon density for serotonin system.
- One factor for differences between areas, in general, appears to be distance of the axonal terminal field from its nerve cell body of origin.
  - With distal fields showing persistant deficits on average relative to more proximal fields that can have full recovery or hyperinnervation.
  - Though this doesn’t apply in all cases.

(O’Hearn, 1988) – MDMA leads to selective loss of serotonin axon terminals in rat forebrain

Rats
Dose
- 8 injections of MDA or MDMA
  - SC injections every 12 hours for 4 days, with 20 mg/kg per injection.
Rats killed at 2 weeks or 1-3 days.
Results
- There is extensive loss of serotonergic axons in the cerebral cortex, striatum, and thalamus.
- Axon terminals appear selectively vulnerable, with no damage in preterminal axons, fibers of passage, and raphe cell bodies.
- MDA per dose led to greater denervation.

(Schmidt, 1986) – Loss of serotonin/5-HIAA at 6 days

Rats
Given a single 10 mg/kg injection
Results
- Acute serotonin and 5-HIAA decline that remained present 6 days later, with a 65% of control value for serotonin in the neostriatal region.

(Baumann, 2006) – Different doses lead to variable serotonergic and temperature changes

Rats
Three IP injections of either 1.5 or 7.5 mg/kg, with one dose every 2 hours.
Rats were killed 2 weeks later.
Results
- Temperature
  - Repeated 7.5 mg/kg doses led to persistent hyperthermia acutely.
  - 1.5 mg/kg didn’t result in hyperthermia.
- Serotonin depletion
  - 7.5 mg/kg regimen led to decrease in serotonin levels in frontal cortex, striatum, and olfactory tubercles.
    - Reductions were over 50%.
  - 1.5 mg/kg failed to alter serotonin concentrations.
Interpretation
- Consistent with (O’Shea, 1998) which found high doses of 10-15 mg/kg in rats led to long-term serotonin depletion, but not 4 mg/kg IP.

(O’Shea) – Effects of different doses and frequencies

Rats
Given MDMA IP
Results
- Temperature
  - 4, 10, and 15 mg/kg IP all led to a rise in temperature.
- Effect on serotonin markers 7 days later
  - Single 15 mg/kg dose led to a greater than 50% loss of serotonin and 5-HIAA in the cortex, hippocampus, and striatum.
    - This was accompanied by a loss in SERT binding.
  - There were also neurotoxic changes at 10 mg/kg.
  - But no significant effect at 4 mg/kg.
- Effect of 4 mg/kg given once or twice per day for 4 days on temperature
  - The first dose of 4 mg/kg led to a temperature increase
  - But all subsequent doses (whether twice per day or just once) failed to elicit hyperthermia.
- Effect of 4 mg/kg given once or twice per day on serotonin markers.
  - A 4 mg/kg dose for 4 days led to a small and generally non-significant effect on serotonin, 5-HIAA, and SERT binding 7 days after the last dose.
  - When the frequency switched to twice per day for 4 days, substantial negative effects were seen.
- Effect of 4 mg/kg twice weekly for 8 weeks on body weight
  - Animals given MDMA 4 mg/kg twice per week for 8 weeks had a slowing of weight gain.
- Effect of 4 mg/kg twice weekly for 8 weeks on serotonin markers
  - 4 mg/kg twice weekly led to no loss of serotonin, 5-HIAA, or SERT binding 7 days after the last dose.

Human Evidence

(Win)

71 participants
- 33 heavy ecstasy users
  - Mean age: 23
  - Mean lifetime: 322 tablets
  - Mean time since last use: 8.2 weeks
  - Mean initiation age: 17.7 years
  - Mean dose: 2.7 tablets
- 38 non-ecstasy users
- Both groups had a lot of variation in terms of the type/amount of other drugs.
Results
- A significant of ecstasy, but not other drugs, located for:
  - Reduced CIT binding in the thalamus
  - Reduced fractional anistropy in the thalamus
  - Increased rrCBV in the thalamus
Author interpretation
- Lower CIT binding is likely to mean lower serotonin transporter densities.
- Lower Fractional anistropy could be a sign of axonal damage, though potentially a sign of increased brain perfusion in the thalamus instead.
- Increased rrCBV could be sue to serotonin depletion resulting in vasodilation/decreased vasoconstriction.

(Daumann, 2005)

12 participants with MDMA use, matched to 12 controls
- The two groups had similar cannabis usage patterns, although the average daily dose was higher in MDMA users
- Ecstasy users
  - Mean age: 26.27
  - 6/12 MDMA users reported concomitant use of amphetamines
  - Mean lifetime use: 202 tablets
  - Mean duration of regular use: 32 months
  - Average frequency of use: 2.15 times per month
  - Average dose: 2.69 tablets
Results
- MRI
  - Significantly lower activity level in the left hippocampus in the users during retrieval (cognitive test).
  - However, there was no significant correlation between the different BOLD response and task performance (retrieval accuracy) or other drug use patterns in ecstasy users.
  - And there were no other significant fMRI differences.
Author interpretation
- Found it surprising that the users had similar retrieval accuracy.
  - Although it was considered possible that BOLD response differences could precede functional differences.
- It’s not the case that a smaller BOLD signal is clearly negative as there have been studies showing the opposite in other settings.
  - So the finding could be treated as higher neural efficiency in MDMA users.
  - But it could also be a sign of damage.
- Problem with polydrug use
  - No significant difference found in results between amphetamine and non-amphetamine users, but that could be from small sample size.

(den Hollander, 2012)

10 ecstasy users vs. 7 non-users
- Ecstasy users
  - Mean age: 25.4 years old
    - Note: The mean age was 4.1 years higher in ecstasy using group,
  - Average lifetime use: 281 tablets
  - They did use more amphetamine and cocaine, but there were no significant differences.
Results
- MRI
  - Hippocampal volume in the ecstasy group was on average 10.5% smaller, with no significant differences between left and right hippcoampal volume.
  - No different in overall white matter volume, but a 4.6% lower grey matter volume for the total brain.
Author interpretation
- Chronic use may lead to hippocampal damage
- And the grey matter effect may be suggestive of damage beyond the hippocampus.
- Not interpreted to be from 5-HT axon terminal damage directly given they take up well under 1% of volume
  - But, it could be from changed neuronal and glial structures from lack of growth factors mediated by serotonin.
  - Other factors could be vasospasic ischemia and preexisting differences in hippocampal volume.

(Daumann, 2003)

22 ecstasy users split into heavy and moderate groups, 11 in each
Compared to 11 non-users
Excluded for regular use of opiates, benzodiazepines, or heavy use of alcohol.
- Though amphetamine and cannabis use were present in many users.
Ecstasy users
- Heavy
  - Mean age: 27
  - Lifetime dose: 258 tablets
  - Duration of regular use: 53 months
  - Average frequency: 2.78 times per month
  - Average dose: 1.91 tablets
  - Average onset: 21 years old
- Moderate
  - Mean age: 23
  - Lifetime dose: 27 tablets
  - Duration of regular use: 16 months
  - Average frequency: 1.8 times per month
  - Average dose: 1.57 tablets
  - Average onset: 20 years old
Results
- Performance
  - Heavy users tended to perform slower than 2-back condition, but there was no significant interaction between memory load and group.
- MRI
  - 1-back test
    - For a p<0.01, both user groups had significantly larger activation of right parietal cortex.
      - No significant differences between user groups
    - Activation differences were lateralized to the right and localized in the supramarginal gyrus.
    - For a p<0.001, only the difference between moderate users and controls was significant.
  - 2-back test
    - For a p<0.01, both user groups had increased activation of right parietal cortex.
      - Activation was more superior in moderate users and more inferior in heavy users.
    - Heavy MDMA users relative to controls had higher activation in right BA40.
    - Heavy users had less activation in the left superior temporal lobe
      - Also slightly lower in left superior frontal gyrus and anterior cingulate
    - Compared to moderate users, heavy users had lower signal in several left-lateralized frontal areas, most significant in the superior BA6 and BA9, and inferior frontal gyrus.
    - For a p<0.001, only the lower activation in the left superior temporal lobe between heavy users and controls could be verified.
Author interpretation
- Found it interesting that the groups didn’t have significant n-back performance differences, although the heavy users tended to perform more slowly on the 2-back.

(Roiser)

Four groups
- MDMA users (30)
  - Age: 22.4
  - Lifetime use: 274.6 tablets
  - Time since last use: 75 days
  - Mean highest regular frequency: 4.8 times per month
- ex-MDMA users (20)
  - Age: 27.5
  - Lifetime use: 792.6 tablets
  - Time since last use: 1,021 days
  - Mean highest regular frequency: 9.9 times per month
- Polydrug controls (30)
- Drug-free controls (30)
Ex-MDMA users had more lifetime ecstasy use, more frequent use, and longer abstinence.
Ex-MDMA users also had greater lifetime use of amphetamine than current users.
Results
- IVE questionnaire
  - ex-MDMA users found to be significantly more impulsive than drug-free controls
    - But there was no differences between ex-MDMA users, current MDMA users, and polydrug controls.
  - Impulsiveness was correlated with monthly tobacco use, lifetime cannabis use, lifetime ecstasy use, frequency of ecstasy use, and lifetime amphetamine use.
- Pattern recognition memory (PRM)
  - Delayed PRM latency correlated with total ecstasy use, lifetime amphetamine use, and lifetime cocaine use.
  - Current MDMA users didn’t differ from either control group
- Executive function
  - Current MDMA users didn’t differ from either control on the Mental Rotation test, Tile Manipulation test, or DMTS.
  - ex-MDMA users made more moves per problem than drug-free controls at the “Copy” stage of Tile Manipulation test, but not the “Mirror” or “Mental Rotation” stage.
Author interpretation
- Significant differences between ex-MDMA users and drug-free controls on IVE impulsiveness scale and the Copy stage of the Tile Manipulation test.
- The lack of differences between polydrug controls and current MDMA users could have been from low statistical power.

(Hoshi)

105 participants
- 26 ex-ecstasy users
- 25 current ecstasy users
- 29 polydrug controls
- 25 drug-free controls
Current ecstasy users
- Mean lifetime use: 288 tablets
- Mean dose: 3.86 tablets
- Mean frequency: 2.92 days
- Mean time since last use: 15 days
Ex-ecstasy users
- Mean lifetime use: 253 tablets
- Mean dose: 2.52 tablets
- Mean frequency: 5.35 days
- Mean time since last use: 987 days
Results
- None of the groups differed with aggressive interpretation bias.
- Suggesting that any ecstasy changes on that measure are just transient.

(Selveraj)

12 former MDMA users, 9 polydrug controls, and 19 drug-free controls
Examining SERT
MDMA users
- Mean lifetime use: 243 tablets
- Mean dose: 2.75 tablets
- Mean frequency: 4.75 times per month
- Mean time since last use: 2.74 years
Results
- SERT binding
  - No significant correlations between MDMA use and DASB binding.
Author interpretation
- MDMA users who are abstinent for at least a year don’t show altered SERT availability relative to polydrug controls or drug-free controls.
- This could mean MDMA lacks an effect entirely or it could mean recovery is possible.

(Quednow)

19 chronic MDMA users, 19 drug-free chronic cannabis users, and 19 drug-free controls
All participants were drug-free for at least 3 days before testing.
MDMA users had much more amphetamine use during their history (208 times during life and 0.82 times per week), without about half as much cannabis use.
MDMA users
- Age: 24.21
- Mean Lifetime use: 458 tablets
- Mean Last consumption time: 17 days
- Mean years of use: 3.66 years
- Mean tablets per week: 1.97 tablets
Results
- Memory
  - MDMA users had significantly worse learning performance on RAVLT (rey auditory verbal learning test) than cannabis users or drug-free controls
    - Only weak trend for worse performance on the first trial, but significantly worse on trials 6 and 7
  - Both greater and longer MDMA use were more significantly correlated with worse memory performance.
  - MDMA users showed deficits in learning, consolidation, recall, and recognition during a verbal memory test.
    - Also a worse organization of memory information as seen by high inconsistency of recall and a high lose after interference.
Author interpretation
- MDMA users appear to have a fronto-temporal dysfunction.

(Morgan, 1999)

25 ecstasy users vs. 22 polydrug controls vs. 19 drug-free controls
Looking at memory effects
On testing day, participants had to go through the Rivermead Behavioral Memory Test (RBMT), which involves repeating word-for-word a news story.
Ecstasy group
- Average lifetime use: 50 tablets
- Average time since last use: 65 days (ranging from under a week for 5 participants to over six months in 3 participants)
Results
- Marked impairment of MDMA users vs polydrug group for recall performance.
  - For both delayed and immediate recall.
- Some trend towards a negative association between immediate recall performance and the lifetime use and duration of use of ecstasy.
- Recall performance had a significant association with time since last use
  - Those who hadn’t used for over 6 months had significantly better performance than those who used in the last month or those last using between 1 and 6 months
- 21/25 in the MDMA group self-reported their use being connected to long-term negative effects
  - Mainly anxiety, depression, mood swings, sleep problems, reduced ability to concentrate, and impaired memory

(Yip)

100 ex-ecstasy users vs. 100 non-users
None had regular smoking or alcohol use, a history if illicit drug use, or regular use of other pharmacological drugs.
Ecstasy users
- Mean total use: 36 tablets
- Mean dose per week: 3.70 tablets
- Mean duration of use: 2.38 months
- Mean time of abstinence: 2.23 months
Results
- Former ecstasy users were significantly impaired on measures of verbal and non-verbal memory, switching attention, verbal fluency, and figural fluency.
- Ecstasy users had similar performance to non-users for working memory and attention control.
- Interestingly, ecstasy users performed higher for RFFT-unique.
- No difference found between groups for auditory attention.
- Correlation found between total use and performance rather than with period of abstinence, suggesting consumption is a greater factor when it comes to impairment.

(Gerra, 2000)

15 MDMA users vs. 15 controls
- Tested 3 weeks after last use and after 12 months of abstinence
MDMA users
- Total exposure count: 69 times
- Duration of use: 15 months
- Frequency of use: 4.7 times per month
- Usual dose: 106 mg
Results
- Behavioral
  - MDMA users had significantly higher scores on the BDHI direct aggressiveness scale, the BDHI guilt scale, the HDRS, and on the novelty seeking TPQ scale after 3 weeks.
    - After 12 months, users maintained higher scores on the HDRS and novelty seeking TPQ, but not on BDHI direct aggressiveness or guilt scales.
- Markers of serotonin function
  - At 3 weeks, there were no significant basal level differences between users and control for prolactin.
    - Cortisol levels also didn’t vary at basal.
  - At 12 weeks, there were still no significant basal differences.
  - D-Fenfluramine test
    - Prolactin
      - Prolactin rise was significantly impaired in MDMA users at 3 weeks and after 12 months.
        Prolactin AUC was significantly lower in users than in controls at 3 weeks.
      - At 12 months, prolactin AUCs were significantly lower in users than in controls.
      - There were no significant prolactin response differences between 3 weeks and 12 months in MDMA users.
    - Cortisol
      - At 3 weeks, cortisol response was much lower in MDMA group, with a significant lower AUCs.
      - At 12 months, cortisol jumped back up and there was no significant difference between MDMA users and control group.

(Erritzoe, 2011)

24 MDMA and/or hallucinogen users
21 controls
Measuring SERT and 5-HT2A binding in MDMA and hallucinogen users
MDMA users
- Mean lifetime sessions: 236
- Mean lifetime use: 1296 tablets
- Mean tablets per session: 4.3
- Mean frequency of use: 3.3 days per month
- Mean length of regular use: 72 months
- Mean time since last use: 57 days
Hallucinogen users
- Mean lifetime sessions: 18
- Mean lifetime use: 60 tablets
- Mean tablets per session: 1.8
- Mean frequency of use: 0.3 days per month
- Mean length of regular use: 57 months
- Mean time since last use: 122 days
Results
- SERT
  - Significantly lower SERT binding in MDMA users than hallucinogen users for: pallidostriatum, neocortex, and amygdala.
  - There was a significant negative correlation between lifetime use of MDMA and SERT binding.
  - There was also a positive correlation between abstinence time and SERT binding in some areas but not others
- 5-HT2A
  - For MDMA+hallucinogen users vs controls, there was slightly lower neocortical 5-HT2A binding.
    - However, when removing 2 control subjects with much higher binding values than other control participants, the significance of the difference went away.
  - When comparing MDMA to hallucinogen users, there was no significant group difference between MDMA users, hallucinogen users, and controls.

(Obrocki, 1999)

7 MDMA users vs. 7 controls
- Took the drug for 1-39 months and had cumulative uses of 12-840 tablets
- PET scan conducted 2-16 months after last use.
Results
- Lower glucose metabolic activity within ecstasy group
  - Hippocampus, amygdala, and cingulate
- Elevated glucose metabolic activity within ecstasy group
  - Brodmann’s areas 10 and 11, the putamen, and caudate nucleus bilaterally
- Greatest statistically significant results were in the hippocampus and Brodmann’s area 11
  - Most prominent was in the left hippocampus
  - These differences were in both hemispheres
- There was no observed correlation between PET results and psychopathology, possibly due to small sample size.

(Kish, 2010)

49 ecstasy users vs. 50 controls
Ecstasy users
- Years of use: 4.1
- Lifetime consumption: 206 pills
- Dose per use: 2.2 pills
- Times per month: 2.2 times
- Days abstinent: 45.2
- Also a high rate of other drugs use, including alcohol, tobacco, cannabis, ketamine, cocaine, and methamphetamine.
All ecstasy users had MDMA in their hair
- 47% also had cocaine and methamphetamine was present in 65%, indicating there was underreporting when just asking about methamphetamine use.
Results
- Decreased DASB binding was restricted to the entire cerebral cortices and hippocampus, with the worst in the occipital cortex.
  - No effect on striatum.
  - Analysis indicated ecstasy use was connected with decreasing binding independent of methamphetamine use as detected by hair or cocaine, cannabis, ketamine, psilocybin, or tobacco use.
- There were no significant differences in whole brain volume, size of ventricles, white matter volume, or that in any of the cortical or subcortical regions.
- No difference between males and females.

(Buchert, 2003)

Four groups
- 30 current ecstasy users
- 29 former ecstasy users
- 29 polydrug controls
- 29 drug-free controls
Ecstasy users
- Mean Cumulative use: 827 tablets
- Mean Duration of use: 54 months
- Mean Abstinence time: 24 days
Former ecstasy users
- Mean cumulative dose: 793 tablets
- Mean duration of use: 55 months
- Mean abstinence time: 514 days
Results
- Distribution volume ratios (DVRs)
  - DVR in current ecstasy users was reduced (per b-CIT binding) in all SERT-rich regions, particularly mesencephalon and thalamus.
  - No significant differences in white matter
  - Abstinence had an effect
    - DVR in former ecstasy users was very close to that of drug-naive controls.
  - DVR wasn’t lower (was actually non-significantly higher) in polydrug controls.

(Schilt, 2007)

Prospective study
Initially recruited 188 healthy non-users
- 58 were then examined after using ecstasy
- 60 were examined after remaining abstinent
Ecstasy users
- Mean cumulative use of 3.2 tablets
- Range of 0.5 to 30 tablets during the study period
Results
- It was found that the Ecstasy group had performance within the normal range, but the change scores on immediate and delayed verbal recall and verbal recognition were significantly lower.
  - This means the scores didn’t decline, but they didn’t increase.
- This kind of change was potentially seen with as little as 1 use

(Reneman, 2001)

Investigating the impact of dose, sex, and abstinence.
Groups
- 15 moderate MDMA users
- 23 heavy MDMA users
- 16 ex-MDMA users
- 15 controls
MDMA use
- Moderate
  - Mean lifetime dose: 28.6 tablets
  - Abstinence time: 3.6 months
- Heavy
  - Mean lifetime dose: 530 tablets
  - Abstinence time: 2.3 months
- Ex
  - Mean lifetime dose: 268 tablets
  - Abstinence time: 29 months
MDMA users also reported more amphetamine and cocaine use than controls.
Men were older by 3.1 years and on average consumed more alcohol each week.
Men in the heavy MDMA group had used significantly more tablets and the usual dose per kg was higher than females.
Results
- Binding
  - CIT binding was significantly lower in female, but not male, heavy MDMA users than in controls
    - Though there were non-significantly lower binding in males
  - CIT binding didn’t differ between controls and moderate male or female MDMA users.
  - CIT binding was significantly higher in female ex-MDMA users than female heavy MDMA users, though not higher than controls.
    - Though there was still a trend in female ex-MDMA users for a reduction in binding in parieto-occipital cortex and the occipital cortex.
Author interpretation
- Females may be more susceptible.
- MDMA use at moderate level might not result in lower SERT availability.
- There may be a recovery effect with abstinence, as seen by ex-MDMA females.

(Wagner, 2013)

Prospective study
109 participants
- Inclusion criteria of high probability of future ecstasy use and only limited preexisting use of under 5 pills.
Participants
- 43 (non-users) didn’t use any other illicit substance other than cannabis during the year.
- 23 (MDMA users) had used more than 10 pills during the past year
  - Mean use: 33.6 pills
  - Range use: 10 – 62 pills
  - Mean occasions: 13.5 times
  - Range occasions: 4 – 36 times
- Remaining subjects left out due to fewer than 10 pills being used.
No considerable difference in cocaine, hallucinogen, sedative, solvent/inhalant, or opioid use.
MDMA users had more use of amphetamines
Hair samples taken randomly confirmed self-reported substance use in every subject but 1, who was then also excluded from analysis.
Results
- No significant impact on attention or information processing speed.
- Significant negative effect on immediate and delayed recall in episodic memory.
- No significant effect on any AVLT variable.
- No significant effect on frontal/executive functioning.
- When entering amphetamine use as a covariate into the multivariate analysis, the effects did not remain significant.
  - However, a multivariate analysis for amphetamine use as a grouping variable (0 vs. 5 grams) didn’t yield any significant effect.
Author interpretation
- MDMA seems to result in specific impairments on episodic memory.

(Taurah, 2014)

Measuring depression, impulsiveness, sleep, and memory in past and present users
Six groups
- 182 in ND – People with no recreational drug use history.
- 172 in AN – People with alcohol and/or nicotine use.
- 163 in CAN – People with cannabis use with/without alcohol and/or nicotine
- 169 in PD – People who’ve never used MDMA, but have used amphetamine, cocaine, heroin, or ketamine.
- 154 in MDMA – Polydrug users who’ve taken MDMA in the past 6 months, but not for at least 3 weeks (mean abstinence of 4 weeks)
- 157 in Ex-MDMA – Polydrug users with ecstasy use history, but abstinent for at least 4 years (mean abstinence of 4.98 years)
MDMA use
- MDMA
  - Mean age of initiation: 18.25 years
  - Mean usual dose: 170 mg
  - Mean lifetime use: 69 grams
- Ex-MDMA
  - Mean age of initiation: 23.47 years
  - Mean usual dose: 90 mg
  - Mean lifetime use: 36 grams
Results
- Depression, impulsiveness, and sleep
  - Past and present ecstasy users had higher mean BDI, BIS-11, and PSQI scores than the control groupos.
  - No difference between the two ecstasy groups for the BDI or PSQI
    - Compared with present users, ex-users had lower BIS-11 scores.
  - For control groups
    - BDI, BIS-11, and PSQI increased progressively between ND, AN, CAN, and then PD group.
    - All drug groups showed significant impairment vs. ND, with exception of AN for the BDI.
- Memory
  - General, verbal, visual, and delayed memory
    - Both ecstasy groups had greater impairment than poldydrug users and other controls
    - No significant difference between ecstasy groups
  - Attention memory
    - Past ecstasy users were more impaired than current ecstasy users
    - Though neither ecstasy using group differed significantly from the polydrug users
- Correlations and regression analyses
  - Correlations on test scores were strong for lifetime ecstasy use and age
  - Regression
    - Ecstasy as a dominant predictor for BDI, BIS, PSQI Global, BDI Somatic, BIS Motor, nonplanning, and attention
      - But not for the WMS-R
    - Ecstasy was a weak predictor for memory tests
Author interpretation
- Ecstasy users appeared to have increased psychological disturbances compared to polydrug users.
- There was limited evidence for a reduction in impairment with prolonged abstinence.
- On ten of the psychometric measures, past and present ecstasy users had greater impairment vs. polydrug users and other controls.
- Lifetime ecstasy use was significantly correlated with performance on all tests.
- Regression analysis only revealed ecstasy use was strong predictor for depression, impulsiveness, and sleep
  - With a weak predictor for memory, with some control drugs being moderate to strong predictors.
- It appears ecstasy on its own is negative for depression, impulsiveness, and sleep, while polydrug use is important for memory deficits.

(Bhattachary, 2001)

Looking for evidence of cognitive impairment.
Groups
- 20 non-users
- 18 novice users: 1-5 uses and never more than once per month, with at least one use in past 21 days.
  - Days since last use: Mean of 8.56 days
- 26 regular users: At least 5 uses and at least twice within past 21 days
  - Days since last use: Mean of 7.42 days
- 16 abstinent users: Had been regular users but didn’t take it in past 30 days, though none were abstinent longer than 120 days.
  - Days since last use: Mean of 46.25 days
Results
- Verbal memory
  - For immediate recall they were impaired vs non-users, with no difference between regular and abstaining users.
  - For delayed recall, non-users did better than regular and abstaining users, but not differing from novice users.
    - Abstaining users had better recall than regular users.
  - Lifetime consumption had a correlation with memory scores, with heavier use correlated with poorer delayed and immediate recall scores.

(MacInnes, 2001)

Demonstrating lasting depressive symptoms, albeit mild.
24 former chronic ecstasy users vs. 29 controls.
Ecstasy users
- Weeks since last use: 26
- Total number of tablets: 527
- Maximum number in one 12 hour period: 5.4
Results
- Depressive symptoms
  - Former users had significantly higher BDI than controls.
    - Total population score of 9.2 in users vs. 5.2 in non-users.
- Indices of life-stress
  - Female users had significantly elevated life stress units and locus of control scores compared to males.
  - No different on the Daily Hassles scale.
- Depression was correlated with peak usage in 12 hours as well as life stress.

(McCann, 1999)

22 ecstasy users vs. 23 controls
Ecstasy group
- Number of uses: 215 (30-725 range)
- Duration of use: 4.52 years
- Frequency of use: 5.72 times per month (0.8-15 times)
- Usual dose: 272 mg (100 – 100 mg)
- Abstinence time: 13.91 weeks (3-157 weeks)
All participants abstained for drugs for at least 3 weeks before testing, with drug-free status confirmed by urine and blood drug screens.
Tests:
- CSF measurement of monoamines, sleep studies, pain testing, personality assessment, and pharmacological studies with the mixed serotonin agonist m-CPP.
Results
- Cognitive
  - There was a group effect for the Logical Reasoning and Code Substitution,
  - and a group-x-time effect on Serial Add/Substract tests
  - No other differences on the other WRAIR PAB tasks.
  - Group differences for Logical Reasoning and Code Substitution had lower overall scores in MDMA group, with no significant differences in throughput scores between groups at any particular time.
  - There were similar Serial Add/Subtract scores at baseline for the groups, but significantly lower Day 2 an 3 scores in MDMA users, indicating impaired learning and lower peak performance.
  - For the Delayed Recall test, a post comparison showed significantly lower baseline scores in MDMA users.
- Accuracy
  - Post hoc analysis showed the MDMA group was significantly less accurate at baseline for the Code Substitution and Delayed Recall tests.
- Speed
  - Post hoc analysis showed the MDMA group was slower than controls on Day 2 and Day 3.
- Association between MDMA use and cognitive performance
  - Association found between total MDMA exposure and peak performance on Code Substitution
- CSF monoamines
  - MDMA users had significantly lower CSF 5-HIAA concentrations with controls, with no CSF HVA or MHPG differences.
  - There was no significant correlation between 5-HIAA levels and cognitive performance.
Author interpretation
- MDMA users abstaining for at least 3 weeks had impairments on 4/7 Walter Reed Army Institute of Research Performance Assessment Battery
  - Deficits in sustained attention, short-term memory, and working memory.
  - Working memory deficits were directly associated with extent of previous MDMA use.

(McCann, 1994)

30 MDMA users vs. 28 controls
MDMA users
- Number of uses: 94.4 (25-300)
- Duration of use: 4.98 years
- Frequency of use: 4.16 times per month (0.15-20)
- Usual dose: 170 mg
- Abstinence time: 17.9 weeks (2-104)
MDMA users had higher amphetamine, cocaine, benzodiazepine, and hallucinogen use.
Results
- CSF monoamines
  - 5-HIAA
    - MDMA users had significantly lower 5-HIAA levels
    - Within the MDMA group, the reduction was much lower in females (46% vs 20%)
    - 5-HIAA levels were negatively correlated with the number of MDMA uses, but not to significance.
    - 5-HIAA levels not correlated with duration, frequency of use, or abstinence time.
  - HVA
    - MDMA females had lower HVA levels than control females, but the levels between males in either group weren’t difference.
  - MHPG
    - No effect
- Basal prolactin and response to tryptophan
  - Basal
    - No variation in basal levels.
  - Tryptophan
    - No significant drug between for prolactin response.
- Pain sensitivity
  - No effect of pain sensitivity or pain endurance.
- Personality
  - MPQ (multidimensional personality questionnaire)
    - MDMA users had less impulsivity (higher control scores)
    - MDMA females had higher controls scores than all other groups
    - A near significant effect on harm avoidance, with MDMA users having higher harm avoidance.
    - MDMA females had higher harm avoidance than all others.
  - BDHI (buss durkee hostility inventory)
    - Both MDMA groups had significantly less indirect hostility than control females, but neither MDMA group was significantly different from male controls.
  - EPQ (eysenck personality questionnaire)
    - No impact on sociability or impulsivity.
- Correlations between biological and personality measures
  - MPQ
    - For females, 5-HIAA was positively correlated with alienation, with a near significant negative correlation on control.
    - For males, positive correlation between 5-HIAA and harm avoidance, with a near significant positive correlation with constraint.

(Daumann, 2004)

Investigating neural mechanisms of working memory in continuing vs. abstaining ecstasy users
17 people at baseline and then 18 months later
- Abstinent from ecstasy and amphetamine: 8 people
- Still using ecstasy and amphetamine: 9 people
Use at baseline
- Abstaining
  - Lifetime use: 243 tablets
  - Duration of use: 31.50 months
  - Average frequency of use: 3.43 times per month
  - Average single night use: 1.66 tablets
  - Abstinence time: 487.50 days
- Continuing
  - Lifetime use: 149.44 tablets
  - Duration of use: 42 months
  - Average frequency of use: 1.72 times per month
  - Average single night use: 1.89 tablets
  - Abstinence time: 15.67 days
Use at 18 months during interim
- Abstaining
  - N/A
- Continuing
  - Interim use: 35.56 tablets
  - Duration of regular use: 14.86 months
  - Average frequency of use: .89 times per month
  - Average single night use 1.97 tablets
  - Abstinence time: 69.38 days
Results
- No significant change in abstinence users between timepoints in cortical activation for n-back conditions.
- Continuing users showed an activation increase in two clusters in the parietal cortex compared to baseline for most difficult 2-back test
- For continued users, a stronger enhancement of the hemodynamic response compared to baseline was associated with a higher average single night dose.
- No correlations found between fMRI changes and other overall or interim drug use for other substances.
Author interpretation
- Interim abstinent users don’t show changes after 18 months of not using the drug, but that’s of unclear importance.
- Continued use may lead to greater parietal activation during working memory tests.
- The lack of impairment in ecstasy users suggests that altered BOLD response could come before clear impairment.

(Gouzoulis-Mayfrank, 2005)

Measuring memory performance between abstinent and continuing users
38 people compared between baseline and 18 months later
- 17 had no or very minimal interim use (0-5 pills)
- 21 had continued use of 20-400 pills
Use stats
- Abstinent
  - Total before
    - Total use: 146.88 pills
    - Duration of use: 24 months
    - Frequency of use: 2.53 times per month
    - Average dose: 1.60 pills
    - Abstinence time: 471.25 days at baseline
  - Interim
    - Total use: 1.31 pills
    - Duration of use: 0.8 months
    - Frequency of use: 0.27 times per month
    - Average dose: 0.34 pills
    - Abstinence time: 798 days
- Continued
  - Total before
    - Total use: 357 pills
    - Duration of use: 36.25 months
    - Frequency of use: 2.95 times per month
    - Dose: 2.21 pills
    - Abstinence time: 40.45 days at baseline
  - Interim
    - Use: 53.85 pills
    - Duration of use: 12.90 months
    - Frequency of use: 2.16 times per month
    - Dose: 1.79 pills
    - Abstinence time: 58.76 days
- Continuing users had higher amphetamine use during the interim.
Results
- Performance remained stable between timepoints in abstinent group.
  - No tendency to improve or deteriorate during the time
- Continued users also didn’t show deterioration, and they actually higher performance on one memory test (city map, immediate recall) at 18 months
- No association between test performance and other drug use.
Author interpretation
- Considered surprising to see no change positive/negative in either group.
- Data doesn’t support or rule out memory decline from MDMA use.

(Parrott, 2000)

Showing possible psychological problems in heavy ecstasy polydrug users
Groups
- Non users (22)
- Light users (16)
  - Mean number of uses of 6.8 (1 – 20)
- Heavy users (12)
  - Mean number of uses of 371 (30 – 1000)
Ecstasy users had significantly higher cocaine, amphetamines, and hallucinogens use.
Results
- Heavy users
  - Significantly higher scores on the SCL-90 for somatisation, obsessionality, anxiety, hostility phobic-anxiety, paranoid ideation, psychoticism, poor appetite, and restless/disturbed sleep.
- Light users
  - Only significantly different on SCL-90 for paranoid ideation and psychoticism.

(Reneman, 2001)

Examining SERT and verbal memory scores
22 recent users vs. 16 ex-users vs. 13 controls
- Recent users abstinent for 2.4 months
- Ex-users abstinent for 29 months
Significantly lower level of education in ecstasy groups, but not DART-IQ test.
MDMA groups
- Users
  - Total use: 485 tablets
  - Usual dose: 2.2 tablets
  - Duration of use: 5.5 years
- Ex-users
  - Total use: 268 tablets
  - Usual dose: 2.1 tablets
  - Duration of use: 4.6 years
Results
- Cortical SERT binding significantly lower in recent MDMA users, but not in ex-users.
- Recent and ex-users recalled significantly fewer words on immediate recall test
  - 47 for recent users, 48 for ex-MDMA users, and 60 for controls
- Recent and ex-users recalled significantly fewer words on delayed recall test
  - 9.8 for recent users, 10.1 for ex-users, and 13.1 for controls
- Higher lifetime use of MDMA correlated with greater decrements in immediate verbal memory.
Author interpretation
- People abstinent for over a year don’t show different SERT densities, but they still show memory declines.

(Di Iorio)

15 female MDMA users vs. 10 female non-users
Studied using labelled setoperone to determine 5-HT2A status in cerebral cortex
Data from 1 female user was exclueded due to presence of cocaine in hair sample.
MDMA users
- Lifetime use: 13.50
- Lifetime use: 1400 mg
- Abstinence time: 689 days
All participants had to have no used amphetamines, cocaine, or LSD for at least 90 days.
Results
- MDMA users had increased 5-HT2A binding potential in 5 cortical clusters: occipitoparietal, temporal, occipitotemporal-parietal, frontal, and frontoparietal regions, but also limbic areas.
  - Greatest between-group difference was in the temporal cluters, where 5-HT2A availability was 20.5% higher in MDMA users.
  - Other regions:
    - 19.7% higher in occipitoparietal
    - 18.3% higher in occipitotemporal-parietal
    - 16.6% higher in frontal
    - 18.5% higher in frontoparietal
  - Regions with increased binding potential involved more brain right hemisphere regions than left, especially for frontal regions.
  - No regions had lower binding potential for MDMA users.
- Lifetime MDMA use (in mg) was positively associated with receptor binding in 4 cortical clusters: frontoparietal, occipitotemporal, frontolimbic, and frontal.
- When including duration of abstinence in the regression model, the results were still significant, indicating lack of effect on abstinence on the impact that MDMA use overall has on 2A binding potential.
- No other drugs had significant associations with binding potential
Author interpretation
- Female MDMA users have chronic changes that may be reflective of serotonergic transmission reduction, which could lead to 2A upregulation.
  - More research needs to validate the validity of higher 2A binding potential as a marker of serotonin denervation.
- Other studies have found that while 2A levels in MDMA users are lower in the weeks after use, that changes in months after use.

Potential Routes of Dysfunction

Metabolism

(Esteban, 2001) – MDMA can induce monoamine release, but not toxicity, with central injection

Rats given MDMA via central injection
- The dose was intended to replicate what would be locally seen with IP administration.
Results
- MDMA (100, 200, and 400 uM) perfusion led to dose-dependent serotonin increase in the hippocampus, with no change in temperature during perfusion.
- Acute
  - MDMA at 400 uM led to rapid and substantial DA rise in striatum
- Seven days later
  - MDMA at 400 uM in hippocampus led to no change in serotonin concentration of rats kept in normothermic or hyperthermic conditions.
  - MDMA at 400 uM with perfusion led to no difference in serotonin or DA concentrations in the striatum compared to controls.
Author interpretation
- Injecting a dose of MDMA that leads to concentrations similar to IP administration known to be neurotoxic doesn’t lead to neurotoxicity
- It appears metabolism is required for toxicity

(Jones, 2004) – Showing presence of neurotoxic thioether metabolites in rat brain following SC MDMA

Rats
Given MDMA SC and in some tests acivicin, an inhibitor of y-GT.
Results
- Quantification of N-methyl-a-methyldopamine thioether conjugates in the striatum
  - Maximum 5-glutathion-s-yl-n-methyl-a-methyldopamine and 2,5-bis-glutathion-s-yl-n-methyl-a-methyldopamine concentrations were reached at 20-40 min following SC injection
    - The peak level was around 78 to 102 nmol for 5-glutathion-s-yl-n-methyl-a-methyldopamine after just one injection
  - Maximum levels of 5-n-acetylcystein-s-yl-n-methyl-a-methyldopamine and 2,5-bis-n-acetylcystein-s-yl-n-methyl-a-methyldopamine levels reached in 80 – 100 min
- Acivicin pretreatment raises levels of conjugates in the brain and also reduces hyperthermia
  - Pretreatment with a y-GT inhibitor, acivicin, led to higher levels of the N-methyl-a-methyldopamine thioether metabolites in the striatum
  - Acivicin also managed to eliminate the hyperthermia response to MDMA.
    - Earlier research showed the y-GT inhibition could boost neurotoxicity
- Correlation between metabolites and neurotoxicity
  - Strong positive correlation between striatal N-acetylcysteine metabolites and the degree of neurotoxicity
  - While strongest for the striatum, there was also a correlation between metabolites and declines in 5-HT/5-HIAA in the cortex, hippocampus, and hypothalamus.
  - Animals with greater striatal 5-n-acetylcystein-s-yl-n-methyl-a-methyldopamine and 2,5-bis-n-acetylcystein-s-yl-n-methyl-a-methyldopamine levels had greater serotonin and 5-HIAA declines.
- 5-n-acetylcystein-s-yl-n-methyl-a-methyldopamine is a serotonergic neurotoxin
  - Intrastriatal administration of 5-n-acetylcystein-s-yl-n-methyl-a-methyldopamine leads to significant declines in striatal and cortical serotonin and 5-HIAA.
  - Despite injection into striatum, there were declines in the hippocampus and hypothalamus, indicating difusion.
  - 5-n-acetylcystein-s-yl-n-methyl-a-methyldopamine also had a milder effect on dopamine, with the highest (21 nmol) dose resulting in lower DA levels in the striatum and cortex by 16 and 18%, respectively.
Author interpretation
- Systemic administration of MDMA can lead to thioether metabolites of N-methyl-a-methyldopamine in the striatum of rats.
- At least one of the metabolites, 5-n-acetylcystein-s-yl-n-methyl-a-methlydopamine, is directly toxic when injected into the striatum.
- Neurotoxicity of MDMA appears to be coming, at least partly, from N-methyl-a-methyldopamine’s oxidation to the quinone and conjugation with glutathione.
- Acivicin can potentiate the neurotoxicity while also eliminating hyperthermia, indicating hyperthermia isn’t required for toxicity, but it can be a potentiator.
- The persistance of the NAC conjugates in the brain suggests there could be accumulation following multiple drug administration.
- Interestingly, behavioral responses are seen with the glutathione and NAC conjugates, but not with a-methyldopamine itself.
- Since MDA is a minor metabolite of MDMA, N-methyl-a-methyldopamine could be more problematic.

(Perfetti, 2009) – Found thioether conjugates in human urine after MDMA

15 human participants
Dose
- 1.5 mg/kg (75 to 100 mg) in a controlled setting
Results
- Catechol-thioether metabolites of MDMA were identified and quantified
  - 5-(N-acetylcystein-S-yl)-n-methyl-a-methyldopamine and 5-(N-acetylcystein-S-yl)-a-methyldopamine
- Fraction of MDMA dose recovered in urine at 4 hours as thioether adducts is 0.002%
  - But this could be an underestimate of exposure.
Interpretation
- These results show known neurotoxins are found in human urine after MDMA is given.
- CYP2D6 and COMT polymorphisms could impact this.
  - COMT is more likely to be important.

(Capela, 2006) – In vitro examination of neurotoxicity of metabolites and the role of hyperthermia

Studying n-methyl-a-methyldopamine, a-methyldopamine, and 5-glutathion-s-yl-a-methyldopamine
In vitro test using rat cortical neurons
Results
- Impact of MDMA and metabolites on neurotoxicity and the role of concentration, time, and temperature
  - Cell viability assessed by the MTT test
  - After 24 hours of incubation, MDMA, n-methyl-a-methyldopamine and a-methyldopamine had toxicity at normothermic and hyperthermic conditions.
  - Toxicity was greater with all drugs at 40°C compared to 36.5°C
  - Normothermic
    - Toxicity from metabolites was greater than that of MDMA
    - 5-glutathion-s-yl-a-methyldopamine is the most toxic
  - Increasing the incubation to 48 hours boosts the level of neurotoxicity, still with metabolites worse than MDMA itself.
- Protective effect of NAC (1 mM) and PBN (100 uM)
  - They were given 1 hour prior to 5-glutathion-s-yl-a-methyldopamine.
    - They protected against neurotoxicity at 24 hour measure.
  - Similar protection against N-methyl-a-methyldopamine and a-methyldopamine after 38 hours of incubation.
- MDMA metabolites seem to lead to apoptotic cell death
  - Investigating the mechanism of cell death found it was typical of apoptotic cell death.
  - Neurons exposed to increasing metabolite concentrations during a 24 hour period showed: neurite disintegration, chromatin condensation, membrane blebbing, cytoplasmic shrinkage, nuclear fragmentation, membrane integrity loss, and neuritic processes.

(Johnson, 1992) – Metabolites of MDMA lead to extended tryptophan hydroxylase decline

Rats
Exposed to 3,4-dihydroxymethamphetamine (135 ug) or 2,4,5-trihydroxymethamphetamine (50-200 ug) through intracerebroventricular (ICV) injection
Rats were killed 5 days later
Results
- 2,4,5-trihydroxymethamphetamine
  - Dopamine
    - Reduced striatal tyrosine hydroxylase activity in a dose-dependent manner, without significantly affecting substantia nigra activity
      - 50, 100, or 200 ug led to reduction of tyrosine hydroxylase: 21%, 56%, and 76%
    - Dopamine and DOPAC concentrations were also reduced in the striatum
      - 50 ug – DOPAC fell to 77% of control and HVA fell to 74% of control
      - 200 ug – DA fell to 36% of control, DOPAC fell to 44% of control, and HVA fell to 37% of control
  - Serotonin
    - Impact on tryptophan hydroxylase activity
      - 50 ug: 58% of control in the hippocampus and 132% of control in median raphe
        Normal activity in the frontal cortex and striatum
        Trend towards an increase in the dorsal raphe with 123% of control
      - 200 ug
        53% of control in frontal cortex, 70% of control in striatum, and 10% of control in hippocampus
      - 100 and 200 ug doses led to increase in tryptophan hydroxylase activity in the dorsal raphe to 154% and 144% of control, respectively.
        Failed to alter activity in median raphe
      - 5-HT and 5-HIAA
        50 ug
        Cortical serotonin fell to 89% of control
        Hippocampal serotonin fell to 79% of control
        100 ug
        Hippocampal serotonin fell to 60% of control
        200 ug
        Cortical serotonin fell to 70% of control
        Striatum serotonin fell to 83% of control
        Hippocampal serotonin fell to 30% of control
Interpretation
- Study shows 2,4,5-trihydroxymethamphetamine leads to an extended decline in tyrosine hydroxylase and tryptophan hydroxylase, along with reductions of DA and 5-HT, going against what centrally administered MDMA appears to result in.

(Elayan, 1993) – MDMA metabolites reduce tryptophan hydroxylase activity

Rats
Studying the impact of 2,4,5-trihydroxyamphetamine (THA) and 2,4,5-trihydroxymethamphetamine (THM)
1 uMol was given for each chemical, injected into the right lateral ventricle
Animals sacrificed 3 hours after injection
Results
- Effect on hippocampal and striatal TPH
  - THM
    - Failed to reduce frontal cortex TPH activity
    - Decreased hippocampal TPH activity to 54% of control
    - Decreased striatal TPH activity to 87% of control
  - THA
    - Decreased hippocampal TPH activity to 8% of control
    - Decreased striatal TPH activity to 79% of control
- Effect on striatal tyrosine hyxroxylase activity
  - THM
    - Led to no significant change.
  - THA
    - Led to decline in TH activity to 75% of control.
- Effect on striatal DA, serotonin, and their metabolites
  - Dopamine
    - THM
      - Did not alter DA levels, but it brought DOPAC to 515% of control and HVA to 366% of control.
    - THA
      - Despite decreasing TH activity, it actually increased DA levels to 135% of control in the contralateral striatum, which was accompanied by 287% of control for DOPAC and 237% of HVA for control.
  - Serotonin
    - THM
      - Decreased 5-HT level to 74% of control
      - 5-HIAA increased to 149% of control
    - THA
      - Brought 5-HT level to 115% of control
      - No impact on 5-HIAA
- In vitro studies using different THA concentrations on incubated striatal and hippocampal tissues
  - Striatal TPH
    - 0.1, 0.5, and 5 mM
      - Led to reduction of TPH activity to 70, 54, and 17% of control
  - Hippocampus TPH
    - 0.01, 0.1, 0.5, and 5 mM
      - Led to reduction of TPH activity of 71, 68, 47, and 3% of control
Author interpretation
- Both THA and THM can decrease tryptophan hydroxylase activity after ICV administration.
- Increase of DA and its metabolites from THA might be from release of dopamine.
  - It’s worth nothing this was on the contralateral side.
    - It’s possible the increase came from compensation for a reduction in enzyme activity on the injection side.

(Miller, 1997) – Locating a potentially neurotoxic MDA metabolite

Rats
Examining the effects of systemic MDA and central ICV administration of metabolites
Results
- Behavior in rats
  - ICV administration of 5-glutathion-s-yl-a-methyldopamine and 5-n-acetylcystein-s-yl-a-methlydopamine
  - Led to hyperactivity, aggressiveness, displayed forepaw treading, Straub tails, and most displayed splayed hind limbs
- Effect on serotonin and DA concentrations
  - MDA (93 umol/kg SC) led to significant hippocampal, striatal, and cortical serotonin declines 7 days later
  - No declines see with 5-(glutathion-s-yl)-a-methyldopamine (4 x 720 nmol ICV) or 5-(n-acetylcystein-s-yl)-a-methyldopamine (4 x 100 nmol ICV)
  - Yet, 2,5-bis-(glutathion-s-yl)-a-methyldopamine did lead to neurotoxic signs at 4 x 475 nmol ICV
    - Produced serotonergic neurotoxicity 7 days after last dose
      - Ipsilateral cortex – 29% decrease
      - Contralateral cortex – 40% decrease
      - Ipsilateral hippocampus – 65% decrease
      - Contralateral hippocampus – No change
      - Striatum was less affected
        Down 24% for ipsilateral and 16% for contralateral
  - No adverse effect from 2,5-bis-(glutathion-s-yl)-a-methyldopamine on dopamine system.
Author interpretation
- Very similar behavior response from MDA subcutaneous or 5-(glutathion-s-yl)-a-methyldopamine ICV
  - This appears to be a serotonin syndrome response.
- 5-(n-acetylcystein-s-yl)-a-methyldopamine required only 7 nmol to produce overt MDA-like behaviors, though no lasting neurotoxicity was seen.
- 5-(glutathion-s-yl)-a-methyldopamine can form 2,5-bis-(glutathion-s-yl)-a-methyldopamine
  - Dropped serotonin levels in striatum, hippocampus, and cortex to 24%, 65%, and 30%
- Believed that 2,5-bis-(glutathion-s-yl)-a-methyldopamine neurotoxicity may come from further downstream metabolites, 2,5-bis-(cystein-s-yl)-a-methyldopamine and 2,5-bis-(N-acetylcystein-s-yl)-a-methyldopamine.

(McCann, 1991) – Major MDA metabolites aren’t responsible for toxicity

Rats
Dosing
- Experiment 1
  - Carbidopa (25 mg/kg IP)
  - Followed by either a-methyldopamine or 3-O-methyl-a-methyldopamine (either at 200 mg/kg SC)
    - These were given twice daily for 4 days.
  - One group was given both drugs.
- Experiment 2
  - a-methyldopamine and 3-O-methyl-a-methyldopamine were given ICV
    - Doses ranging from 100 to 400 ug/10 uL
- Experiment 3
  - a-methyldopamine was given intrastriatally at 400 ug/uL
Rats examined 10-14 days later.
Results
- Neither a-methyldopamine nor 3-o-methyl-a-methyldopamine led to a lasting serotonin depletion after SC administration.
- Meanwhile MDA SC led to a profound depletion of serotonin.
- Rats given intraventricular a-methyldopamine or 3-o-methyl-a-methyldopamine didn’t show region brain serotonin depletion at 14 days.
Author interpretation
- Neither a-methyldopamine nor 3-o-methyl-a-methyldopamine seemed to cause neurotoxicity.
  - Those are the main MDA metabolites.
- MDA itself does lead to neurotoxicity.

(Bai, 1999) – Multiple MDA conjugate metabolites can cause toxicity centrally

Rats
Dosing (all given four times at 12 h intervals)
- 5-glutathion-s-yl-a-methyldopamine (200 nmol and 400 nmol)
- 2,5-bis-glutathion-s-yl-a-methyldopamine (150 and 300 nmol)
- 5-n-acetylcystein-s-yl-a-methyldopamine (7 and 20 nmol)
Given into the left striatum, cortex, or hippocampus
Rats killed 7 days later.
Results
- MDA (93 umol/kg SC)
  - Reduction in serotonin levels in striatum, cortex, and hippocampus of 43, 38, and 52%
  - DA and NE not affected
- 5-glutathion-s-yl-a-methyldopamine
  - Intrastriatal
    - Decreased in serotonin to 74% of control with 200 nmol and to 68% of control with 400 nmol
    - Concentrations in the striatum contralateral to the injection were unaltered
    - Concentrations in the ipsilateral cortex were also significantly decreased, with no hippocampal impact.
  - Intracortical
    - Decrease in serotonin to 62% of control with 200 nmol and to 54% of control with 400 nmol
    - Concentrations in the cortex contralateral to the injection were unaltered.
    - Concentrations in the striatum ipsilateral to the injection were also significantly reduced, with no impact on hippocampus.
- 2,5-bis-glutathion-s-yl-a-methyldopamine
  - Intrastriatal
    - Decrease in serotonin of 47% with 300 nmol
    - Concentrations in the striatum contralateral to injection unaffected.
  - Intracortical
    - Significantly decreased serotonin concentrations in the cortex and striatum ipsilateral to injection, without impact on hippocampus.
- 5-N-acetylcystein-s-yl-a-methyldopamine
  - Intrastriatal
    - At both 7 and 20 nmol, it caused a decline in striatal serotonin only at the site of injection.
  - Intracortical
    - At 7 and 20 nmol, it caused a decline in cortical serotonin only at the site of injection.
  - Intrahippocampal
    - 20 nmol led to a decline in hippocampal concentrations of serotonin.
Author interpretation
- Direct injections into striatum, cortex, or hippocampus of 5-glutathion-s-yl-a-methyldopamine, 5-N-acetylcystein-s-yl-a-methyldopamine, or 2,5-bis-glutathion-s-yl-a-methyldopamine lead to significant declines in serotonin 7 days later.
- This goes against other results using ICV administration of 5-glutathion-s-yl-a-methyldopamine and 5-N-acetylcystein-s-yl-a-methyldopamine.
- Order of potency
  - 5-N-acetylcystein-s-yl-a-methyldopamine > 2,5-bis-glutathion-s-yl-a-methyldopamine > 5-glutathion-s-yl-a-methyldopamine

Cellular energy disruption

MDMA could lead to serotonergic changes by raising neuronal energy utilization or through disruption of energy metabolism, which may raise intracellular calcium in the mitochondria. It could also deplete brain glycogen and it’s been found to activate glycogen phosphorylase, an enzyme responsible for glycogen breakdown, in astroglial rich primary cultures.

(Darvesh, 2005) – Addressing cellular energy status can block MDMA-induced changes

Rats
Drugs administered intrastriatally
- MDMA (100 uM) and malonate (100 mM) were perfused into striatum for 8 hours.
- Nicotinamide (1 mM) was given 2 hours before MDMA until 6 hours after MDMA/malone perfusion ended.
In other tests
- Rats received MDMA (10 mg/kg IP) at 2 hour intervals for 4 injections.
  - They were also given nicotinamide (1 mM) or ubiquinone (100 uM) directly into striatum beginning 2 hours before MDMA until 6 hours after last injection.
Rats were killed 5 days later.
Results
- Intrastriatal
  - Local perfusion of MDMA and malonate led to 54% reduced serotonin concentrations and 69% reduced dopamine concentrations.
    - Perfusion of MDMA itself didn’t significantly alter serotonin or dopamine.
    - Malonate itself didn’t significantly affect serotonin but it did reduce dopamine by 24%.
  - Rats given nicotinamide before MDMA/malonate didn’t have significantly altered serotonin concentrations.
    - While dopamine was still reduced, it was significantly less.
- Systemic
  - Striatal serotonin was reduced by 48%.
  - Nicotinamide resulted in serotonin concentrations significantly higher in the side adjacent to the probe vs. on the contralateral side only exposed to MDMA.
  - Ubiquinone also blocked the reduction of serotonin in the striatum
    - Comparison found the serotonin concentration on the side exposed to both was much higher than the contralateral side only exposed to MDMA.
  - Impact on hippocampus
    - Nicotinamide significantly attenuated the lasting depletion of serotonin from MDMA.
      - Concentration of serotonin was much higher in tissue exposed to both than to contralateral side only given MDMA.
  - ATP effects
    - ATP concentration significantly reduced after last MDMA injection
      - 30% at 12 hours, and 26% at 24 hours
      - No significant depletion seen at just 1 hour after last injection
  - ATP in the hippocampus was also significantly reduced after MDMA , with a drop of 33% at 12 hours.
Interpretation
- Central MDMA alone doesn’t lead to neurotoxic-like changes, but combining it with striatal malonate, a mitochondrial inhibitor, can result in depleted DA and serotonin concentrations.

Dopamine-related

(Sprague, 1995) – MAO-B inhibition protects against serotonergic changes

Rats
Doses
- MDMA: 40 mg/kg SC
- L-deprenyl: 2 mg/kg IP
- MDL-72974: 1.25 mg/kg IP
L-deprenyl or MDL-72974 was given 30 min before MDMA.
Results
- L-Deprenyl and MDL-72974 both protected against the loss of SERT and loss of serotonin/5-HIAA seven days after MDMA.
Author interpretation
- Attenuation of neurotoxicity by these MAO-B inhibitors suggests deamination of dopamine is a contributing factor.
  - Possible deamination within the serotonergic terminal could lead to higher intracellular levels of hydrogen peroxide.

(Yuan, 2002) – Significant dopamine depletion failed to protect against changes

Rats
Doses
- MDMA: 20 mg/kg IP every 2 hours for 4 injections
- Reserpine: 5 mg/kg IP given 18 hours before MDMA
- AMPT: 150 mg/kg IP every 4 hours for 2 injections, beginning 18 hours after reserpine and 3 hours before MDMA
  - This was given in a separate test to see how inhibiting tyrosine hydroxylase, which would ensure cytoplasmic and vesicular DA depletion, would impact the results.
Other animals were just given reserpine to see its effect on dopamine.
Animals killed 4 weeks or 6 weeks after treatment.
- Left hemisphere cortex, hippocampus, and striatum evaluated for DA, DOPAC, 5-HT, and 5-HIAA.
- Right hemisphere hippocampal and cortical sections used for binding tests
Results
- 22°C
  - Reserpine only
    - Marked dopamine and serotonin reduction at 18-36 hours after injection, meaning it’s effective during the time period of MDMA administration.
  - MDMA only
    - Significant and persisting reductions in cortical serotonin axonal markers.
  - Reserpine pretreatment
    - Significantly reduced the negative effects of MDMA.
      - However, hypothermia elicited by reserpine was seen at 22°C, meaning the effect could be from DA depletion or temperature reduction.
- 27°C
  - MDMA alone
    - Even greater reductions in brain serotonin axonal markers.
    - At this temperature, MDMA also led to long-term depletion of striatal dopamine axonal markers.
  - Reserpine pretreatment
    - Without any evidence of hypothermia this time, reserpine failed to attenuate the long-term negatives of MDMA.
    - Reserpine also didn’t protect against DA-related impact.
- Resperine + AMPT effects
  - Reserpine and AMPT only
    - Found to cause near total depletion of brain dopamine and serotonin.
  - Reserpine and AMPT before MDMA (at 33°C)
    - Despite near complete depletion of DA and serotonin, this were still significant serotonin axonal marker deficits at 6 weeks.
      - Magnitude of impact didn’t differ between MDMA alone vs. those with pretreatment.
Author interpretation
- Endogenous DA isn’t required for serotonin neurotoxicity once you control for the hypothermic response of reserpine.

(Breier, 2006) – Tyrosine may have a role

Rats were given a variety of drugs for different experiments.
In vitro tests also conducted to look at tyrosine’s generation of DOPA in a pro-oxidant environment.
Doses
- MDMA: 10 mg/kg IP every 2 hours for 4 hours
- MDMA: 100 uM for intrastriatal perfusion
- L-Tyrosine: 500 uM for intrastriatal
- L-Valine: 500 uM for intrastriatal
- NSD 1015: 100 uM for AADC inhibition
In vitro
- L-Tyrosine: 25, 62.5, 125, and 250 uM
Results
- Tyrosine effects in striatum
  - Systemic MDMA acutely increases extracellular tyrosine in the striatum, with a fivefold rise over baseline.
  - MDMA also leads to a 40% depletion of serotonin in the striatum 1 week later.
- Local perfusion of MDMA in striatum
  - Local administration of MDMA didn’t affect tyrosine concentrations nor did it deplete serotonin.
- Local perfusion of tyrosine in striatum during systemic MDMA
  - Tyrosine, but not valine, significantly enhanced the depletion of serotonin from MDMA.
- Tyrosine’s in vitro conversion to DOPA
  - Tyrosine in a hydroxyl radical-generating system led to DOPA.
  - DOPA was generated in a concentration-dependent manner.
- Inhibition of AADC in striatum
  - MDMA was given systermically, while NSD 1015 and tyrosine were given centrally.
  - Tyrosine poteniated MDMA’s increase in extracellular dopamine.
  - NSD 1015 fully blocked the MDMA-induced rise in DA.
  - NSD 1015 prevented tyrosine’s enhancement of the MDMA-induced DA rise.
- Systemic MDMA and striatal serotonin content
  - Testing central NSD 1015, tyrosine, or a combo with systemic MDMA.
  - NSD 1015 blocked, while tyrosine increased, MDMA’s depletion of serotonin.
  - Coinfusion of NSD 1015 and tyrosine blocked the exacerbation of MDMA’s serotonin depletion from tyrosine alone.
- Local perfusion of MDMA and serotonin content
  - MDMA didn’t affect serotonin content 1 week later.
  - However, combining it with tyrosine did lead to significant depletion of serotonin in the striatum 1 week later.
  - NSD 1015 added to MDMA/tyrosine blocked the depletion of serotonin.
- Tyrosine effects in the hippocampus
  - Systemic MDMA increased tyrosine concentrations in the hippocampus by 2.5-fold.
  - MDMA led to depletion of serotonin in the hippocampus (-60%) 1 week later.
  - MDMA also boosted DA.
  - Tyrosine potentiated the MDMA-induced rise in DA.
  - NSD 1015 prevented the MDMA-induced rise in DA.
  - A tyrosine hydroxylase inhibitor, AMPT, failed to alter MDMA’s effect.
  - NSD 1015 blocked or attenuated the MDMA-induced depletion of serotonin in the MDMA only and MDMA+tyrosine groups.
Author interpretation
- MDMA given systemically can increase tyrosine levels in the brain.
- Tyrosine can become DOPA in a pro-oxidant in vitro environment.
- Blocking DOPA’s conversion to DA prevents MDMA’s toxicity.
- Tyrosine-derived DA synthesis doesn’t appear reliant of tyrosine hydroxylase.
- Tyrosine seemingly contributes to toxicity, possibly through DA metabolism.
  - Perhaps it’s taken up into serotonin terminals and converted to DOPA in the presence of hydroxyl radicals.
    - MDMA’s metabolism could be a source of hydroxyl radicals.
  - DOPA then becomes DA from AADC in serotonin terminals.
- Hypotheses for why systemic but not central MDMA raises tyrosine
  - Stimulation of peripheral b-adrenergic receptors and the subsequent rise in LNAA transport into the brain, which could raise brain tyrosine.
  - Stimulation of adrenergic receptors by norepinephrine may facilitate the breakdown of skeletal muscle and rhabdomyolysis, leading to higher circulating tyrosine and other amino acids.
  - MDMA may disrupt BBB, such that the normally regulated transport of tyrosine from the brain to the periphery is impaired.
- Hippocampus may be more vulnerable because of lack of DA terminals, meaning local rise in tyrosine preferentially boosts levels of DA in serotonin rather than dopamine terminals.

Drugs That May Enhance The Risk

(da Silva, 2014) – MDMA with other amphetamines

Found the cytotoxicity of amphetamines increases significantly with hyperthermia.
Doses of any given amphetamine (including MDMA) that would otherwise be non-toxic can be made toxic when administered together.

(Khairner, 2010) – MDMA with caffeine

Using mice
Animals were killed 48 hours after last MDMA administration.
Methodology
- First day
  - MDMA (4 x 20 mg/kg IP) at 2 hour intervals
    - Either alone or with caffeine (2 x 10 mg/kg IP) 30 minutes before the first and last MDMA dose.
- Second day
  - Two administrations of vehicle or caffeine (10 mg/kg) 12 hours apart.
- Third day
  - One administration of caffeine.
Results
- GFAP in the striatum and SNc
  - GFAP was significantly higher in the striatum from MDMA and MDMA+caffeine
  - Caffeine significantly boosted the level of GFAP immunoreactivity compared to MDMA alone.
  - No effect in the SNc
  - Caffeine itself didn’t change GFAP
- CD11b immunoreactivity in the striatum and SNc
  - Significantly higher in the striatum and SNc in MDMA and MDMA+caffeine conditions
  - Caffeine significantly boosted the level of immunoreactivity in the striatum, but not the SNc.
  - Caffeine itself didn’t modify CD11b levels
Author interpretation
- MDMA leads to a neuroinflammatory process in mice that’s characterized by microgliosis in the striatum/SNc and astrogliosis in the striatum.
- Caffeine seems to exacerbate the negative effects.
- Caffeine might affect the AUC of MDMA and potentiate hyperthermia.
- There could potentially be a boost to neuroinflammation from antagonizing A1 and A2a adenosine receptors with caffeine.

Drugs That May Decrease The Risk

Alpha-lipoic acid (ALA)

(Aguirre, 1999) – Protective effect

Rats
Drugs
- Control or a-lipoic acid (100 mg/kg IP twice daily) for 2 days
- Thirty minutes after the fourth a-lipoic acid dose or control, rats received either MDMA (20 mg/kg) or saline.
Rats were killed and examined 7 days after MDMA.
Results
- MDMA led to a significant decline in 5-HT content (-40-60%) in the frontal cortex, in the hippocampus, and in the striatum of rats at 7 days.
  - Repeated a-lipoic acid administration completely prevented the loss of 5-HT from MDMA in all brain regions examined.
- MDMA led to a significant decline in paroxetine SERT binding sites in the frontal cortex and striatum (-35%) and the hippocampus (-45%).
  - Repeated a-lipoic acid administration completely blocked the loss of SERT density in the examined brain regions.
- MDMA on its own led to hyperthermia and a-lipoic acid on its own led to hypothermia
  - a-lipoic acid did not prevent the hyperthermic action of MDMA.
- Glial response and GFAP immunoreactivity
  - MDMA led to a significant rise in GFAP immunoreactivity in the stratum lacunosum-moleculare (hippocampus) and the molecular layer of the dentate gyrus
    - The greatest change was in the pyramidal cell layer of the CA1 region.
  - a-lipoic acid completely prevented the glial response change and pattern of GFAP immunoreactivity from MDMA

Acetylcarnitine (ALCAR)

(Alves, 2009) – Protective effect without impacting hyperthermia

Rats
Drugs
- MDMA (10 mg/kg every 2 hours) for four injections
- ALCAR (100 mg/kg) once 30 minutes before injection
- Control animals received saline and ALCAR
Results
- Hyperthermia
  - ALCAR didn’t attenuate MDMA hyperthermia.
- Lipid peroxidation
  - ALCAR didn’t attenuate lipid peroxidation, which was assessed based on malondialdehyde equivalents formation in whole brain mitochondria 14 days after MDMA.
  - Both ALCAR and MDMA on their own also increased malondialdehyde equivalents.
- Protein carbonyls
  - MDMA produced a significant increase in protein carbonyls in whole brain mitochondria 14 days after.
  - ALCAR significantly reduced the increase from MDMA.
  - The levels in the combo group were similar to those in the ALCAR-only group, which were also higher than controls.
- Deletion of mtDNA
  - mtDNA from the prefrontal cortex, the striatum, the amygdala, the ventral mesencephalon, the hippocampus, and the raphe nuclei was analzyed.
  - Deletion of mtDNA was found to be significantly higher with MDMA.
  - Administration of ALCAR significantly attenuated the deletion.
- Expression of the mitochondrial subunit NDII and COXI
  - Expression of NDII was decreased in MDMA compared to control, ALCAR, and combo group.
  - Expression of COXI was also decreased.
  - Animals given ALCAR before MDMA had levels of expression not significantly different from controls.
- Serotonin content
  - MDMA led to reduced 5-HT levels in the prefrontal cortex, striatum, amygdala, VTA-SN, hippocampus, and raphe nuclei at 2 weeks.
    - This was accompanied by a decline in 5-HIAA levels, although it was only significant in the hippocampus and raphe nuclei.
  - ALCAR significantly reduced the decline in 5-HT levels
Author interpretation
- It appears MDMA neurotoxicity is linked to oxidative stress, particularly in the mitochondria.
- ALCAR likely increased lipid peroxidation
  - It’s possible this could be counteracted with a-lipoic acid while also boosting protection against MDMA

Vitamin E

(Johnson, 2002) – Investigating the impact of Vitamin E deficiency on neurotoxicity and hepatotoxicity

Mice
Given d-MDMA (5 or 10 mg/kg SC) every 2 hours for 4 injections
Measurements taken 72 hours after MDMA
Results
- Effect on brain DA and measures of neurotoxicity
  - d-MDMA dose-dependently reduced striatal DA in the vitamin E sufficient mice
    - A low-dose didn’t lead to significant reductions, but the higher dose led to a 80% reduction in DA.
    - A low-dose also didn’t raise striatal GFAP, but the higher dose boosted it by 300%.
  - In deficient animals
    - d-MDMA caused a larger magnitude DA and GFAP change
    - Low dose
      - a 47% reduction in striatal DA was seen
      - Elevated GFAP by 200%
    - High dose
      - Similar level of DA reduction as in sufficient mice (~80%)
      - And a similar level of GFAP rise
- Effect on brain and liver Vitamin E levels and other antioxidant measures
  - d-MDMA administration to sufficient animals
    - Dose-dependent reduction of three of brain antioxidant measures
      - 10 mg/kg
        56% reduced vitamin E
        33% reduced total antioxidant reserve
        24% reduced protein thiols
        Trend towards reduced glutathione concentrations
    - In liver, a similar dose-dependent change.
      - 10 mg/kg
        28% reduced total antioxidant reserve
        21% reduced glutathione
        No significant impact on protein thiols or Vitamin E
    - In deficient animals
      - Further reduction in antioxidant reserve in antioxidant reserve in liver and brain of about 30%
      - It did not further reduce vitamin E, glutathione, or protein thiol levels beyond the deficient diet itself.
      - There was no further reduction in brain Vitamin E levels
- Effect on liver cytoarchitecture
  - d-MDMA in sufficient animals
    - Dose-dependent increase in micro-vesicles replacing the cytoplasm of hepatocytes
      - 5 mg/kg only had a low incidence of microvesicles
      - 10 mg/kg led to numerous microvesicles replacing most of the cytoplasm in hepatic cells
  - In deficient animals
    - 5 mg/kg
      - Damage, but not as much as 10 mg/kg.
    - 10 mg/kg
      - Led to multiple foci of frank liver cell necrosis and areas with multiple microvesicles in the cytoplasm.

Vitamin C

(Shankaran, 2001) – Reduces hydroxyl radical formation and other negatives

Rats
Drugs
- Vitamin C (100 mg/kg IP) every 2 hours for five injections
- MDMA (10 mg/kg IP) every 2 hours for four injections
- Combination of the two, with Vitamin C given 1 hour before each MDMA injection and 1 hour after last MDMA injection.
Results
- MDMA’s effect on brain ascorbic acid and vitamin E concentrations
  - Striatum
    - Significant decline in vitamin E and vitamin C 6 hours post-MDMA
    - No decline in vitamin E or vitamin C at 2 and 12 hours post-MDMA
  - Hippocampus
    - Significant decline in vitamin E and ascorbic acid 6 hours post-MDMA
    - Significant decline in Vitamin C at 12 hours post-MDMA
- Vitamin C’s effect on MDMA’s hydroxyl radical formation, DA release, and hyperthermia
  - Hydroxyl radicals
    - MDMA led to a sustained and significant rise in extracellular 2,3-DHBA in the striatum, a marker of hydroxyl radical formation.
    - Vitamin C significantly attenuated the rise in 2,3-DHBA.
  - MDMA significant raised extracellular DA in the striatum, with no attenuation from Vitamin C.
  - MDMA significantly raised body temperature, with no attenuation from Vitamin C.
- Vitamin C’s effect on MDMA’s serotonin depletion (1 week after dosing)
  - MDMA led to a significant decline in serotonin in the striatum
  - Vitamin C prevented the serotonin depletion to the extent that there was no difference from controls.
- Vitamin C’s effect on functional consequences of serotonin depletion (1 week after dosing)
  - MDMA 7.5 mg/kg IP
    - Rats only given MDMA had a significantly lower rise in serotonin from a challenge dose of MDMA one week later.
    - The rise in serotonin was significantly greater if rats had been given both MDMA and Vitamin C.
      - There was no difference in the response from control rats that didn’t receive MDMA the first time.
  - MDMA 15 mg/kg IP
    - MDMA would normally lead to serotonin syndrome at this dose with headweaving, forepaw treading, and low body posture.
      - Those actions were significantly suppressed if they had received MDMA a week earlier.
    - The actions were much greater in those that received both MDMA and Vitamin C before.
    - Also, only having MDMA the first time attenuated the rise in temperature for the later challenge dose.
      - Vitamin C the first time pallowed hyperthermia to occur significantly more during the challenge.

N-acetylcysteine (NAC)

(Asl) – Protective effect of NAC on cognition and serotonergic changes

Rats
Groups
- Sham
- NAC (100 mg/kg) twice daily for 1 week
- MDMA at either 5, 10, or 20 mg/kg twice daily for 1 week
- NAC 1 hour before each MDMA dose twice daily for 1 week
Learning tests were complete on the same day as the last administration.
Results
- Temperature
  - NAC attenuated MDMA’s hyperthermia.
- Learning/memory
  - NAC reduced MDMA’s memory impairment, though with primarily nonsignificant effect.
- Histological analysis
  - MDMA significantly reduced neural density in the CA1 hippocampus.
  - NAC significantly decreased cell loss.
- Bax and Bcl-2
  - MDMA caused a dose-dependent reduction in Bcl-2, but only significant at 20 mg/kg.
  - NAC pretreatment increased Bcl-2 protein expression, though not significantly.
  - There was a significant rise in Bax expression from MDMA.
    - NAC pretreatment decreased Bax expression, but not significantly.
- Bcl-2/Bax ratio
  - MDMA led to a significant reduction in Bcl-2/Bax protein ratio.
  - NAC pretreatment caused a rise in the ratio, but it was only significant in the rats given 5 mg/kg MDMA.
- Caspase-3 expression
  - Different amounts of MDMA significantly upregulated caspase 3
  - NAC pretreatment significantly decreased caspase 3 protein expression compared to MDMA groups

(Asl) – NAC has protective effect on the cerebellum in rats

Rats
Groups
- Placebo: saline daily for a week
- MDMA: 10 mg/kg IP for a week
- Treatment: NAC 100 mg/kg 30 minutes before each MDMA dose.
Results
- MDMA led to increase in BAX protein expression, which was reduced by pretreatment with NAC, although the difference didn’t reach significance.
- MDMA led to decrease in Bcl-2 protein expression, which was significantly blocked by NAC.
Author interpretation
- MDMA can raise the levels of proapoptotic Bax protein and decrease anti-apoptotic Bcl-2 in rat cerebellum.
- NAC appears capable of protecting against these changes.

Tryptophan & 5-HTP (Warning: While this research provides useful information, these substances shouldn’t be used before MDMA due to acute safety concerns.)

(Sprague, 1994) – 5-HTP and tryptophan can block serotonergic changes

Rats
Groups
- Saline
- MDMA 20 mg/kg
- MDMA and tryptophan (400 mg/kg)
- MDMA and 5-HTP (50 mg/kg) and RO 4-4602 (50 mg/kg)
  - Latter substance to inhibit peripheral decarboxylase of 5-HTP.
MDMA was given SC, all others were given IP.
Pretreatments always given 30 min before MDMA.
Rats studies 1 week after administration.
Results
- MDMA led to a significant reduction of paroxetine-labeled SERT sites in hippocampus, striatum, and frontal cortex
  - Greatest in hippocampus
- Both tryptophan and 5-HTP protected the three brain regions
  - Greatest in the frontal cortex (-28%) and the least protection in the hippocampus (-32%)
- MDMA led to significant declines in 5-HIAA and serotonin levels, with protection from tryptophan and 5-HTP
  - MDMA
    - Serotonin: 43.3% of control
    - 5-HIAA: 69.3% of control
  - MDMA and tryptophan
    - Serotonin: 72.4% of control
    - 5-HIAA: 103.9% of control
  - MDMA and 5-HTP
    - Serotonin: 125.5% of control
    - 5-HIAA: 88.9% of control
Author interpretation
- MDMA’s serotonergic neurotoxicity requires neuron terminals to be depleted of serotonin.
  - It had also been shown that para-chloroamphetamine neurotoxicity could be blocked by tryptophan pretreatment.
- The depletion of serotonin in neuron terminals may then allow uptake of a toxicant like dopamine into the serotonin terminal.

Sildenafil

(Puerta, 2012) – Protective effect without affecting hyperthermia

Rats
Given 8 mg/kg sildenafil oral, 24 hours before MDMA, mimicking a 100 mg tablet in humans
MDMA was administered with 3 doses of 5 mg/kg IP, every 2 hours
Rats killed 7 days later
Results
- Impact on hyperthermia and serotonin deficits
  - Sildenafil partially prevented the loss of serotonin and 5-HIAA in the striatum, frontal cortex, and hippocampus.
  - Sildenafil fully prevented reduced SERT binding in all three brain regions.
  - These effects were independent of any effect on MDMA’s rise in body temperature.
- Impact on metabolism
  - No differences in MDMA, MDA, HMMA, or HMA levels between groups.
- Sildenafil induces the phosphorylation of ERK1/2
  - There was a significant rise in ERK1/2 phosphorylation with sildenafil, which was seen 1.5 hours after administration and persisted for at least 12 hours.
  - Fitting with post-transcriptional regulation, ERK1/2 protein levels weren’t affected.
- Impact on MnSOD expression, superoxide production, and nitrotyrosine formation
  - Sildenafil led to a rise in MnSOD expression and it was still higher 24 hours later.
    - This could protect against the superoxide radicals from MDMA that are normally dismutated by MnSOD.
  - MDMA led to a rise in superoxide and superoxide-derived oxidants in the hippocampus at 1 hour past last dose.
    - This was significantly prevented by sildenafil pretreatment.
  - MDMA also significantly increased concentration of nitrotyrosine in the hippocampus (nitrotyrosine is a marker of peroxynitrite formation).
    - Sildenafil led to nitrotyrosine levels similar to those in control rats.
  - Other antioxidant enzymes, thioredoxin and heme oxygenase-1, in the hippocampus, weren’t affected by sildenafil.
Author interpretation
- Sildenafil, a PDE5 inhibitor, leads to protection against MDMA toxicity without affecting hyperthermia.
- This appears related to increased expresion of MnSOD and an attenuation of superoxide and superoxide-derived oxidants, thereby reducing oxidative stress.
- Sildenafil also prevented the formation of nitrotyrosine.
- Though these effects didn’t fully prevent the loss of serotonin, suggesting other mechanism are playing a role.
- MDMA-induced serotonin depletion appears dependent on peroxynitrite (Darvesh, 2005), a highly reactive anion formed by reaction of nitric oxide with superoxide radicals.
  - This study showed sildenafil’s attenuation of superoxide production and nitrotyrosine concentrations, suggesting that’s the key mechanism.

(Puerta, 2009) – Investigating efficacy and potential pathways

Rats
Conditions
- The dose of sildenafil used in the study was intended to simulate someone taking 20-100 mg of sildenafil.
- Experiment 1
  - Rats given saline or sildenafil (1.5 mg/kg or 8 mg/kg) 30 minutes before ether saline or MDMA (3 doses of 5 mg/kg IP every 2 hours).
- Experiment 2
  - Rats given vardenafil (1.5 mg/kg) before either saline or MDMA.
- Other experiments
  - Rats given a dose of L-NIO (10 mg/kg IP) 30 minutes before sildenafil alone or in combination with MDMA
    - L-NIO is an endothelial nitric oxide synthase inhibitor
- To further investigate the mechanisms, some drugs were given intrastriatally.
Rats killed 7 days later
Results
- Impact of sildenafil on hyperthermia and 5-HT deficits
  - No impact on MDMA-induced hyperthermia with either dose of sildenafil.
  - Even just the low dose of sildenafil protected against serotonin depletion in the striatum.
  - There was a dose-dependent protection in the frontal cortex and hippocampus.
  - After seven days, MDMA alone led to a significant loss of paroxetine binding in all three brain regions.
    - This was prevented with 8 mg/kg of sildenafil, with the low dose not being tested.
- Sildenafil’s effect on cGMP
  - It produced an increase in hippocampal and striatal cGMP levels.
- Importance of the PKG pathway for protection
  - There is subsequent activation of PKG after the rise in cGMP levels from sildenafil.
  - Intrastriatal administration of KT5823 (2 nmol), a PKG inhibitor, was given 30 minutes before MDMA alone or in combo with sildenafil.
    - There was no impact of KT5823 on hyperthermia
    - Pretreatment with KT5823 didn’t alter MDMA’s striatal 5-HT or 5-HIAA depletions, but it did reverse the protective effects of sildenafil.
      - Analysis revealed significant differences for serotonin.
  - As a result, 8-Br-cGMP (a nitric oxide-independent PKG stimulator) was tested 30 minutes before MDMA
    - Found it could also protect against serotonin depletion, without any impact on hyperthermia.
- Importance of mitochondrial KATP channels for protection
  - Sildenafil is known to be cardioprotective via opening of mitochondrial KATP channels.
  - 5-HD, a KATP blocker, was used to see if the channels also played a role in this case.
    - 5-HD (40 nmol) was intrastriatally given 30 minutes before MDMA or MDMA/sildenafil
    - 5-HD had no effect on MDMA’s 5-HT depletion, but it fully reversed the protective effect of sildenafil on striatal serotonin and 5-HIAA concentrations.
- Sildenafil inducing the phosphorylation of Akt upstream from PKG and mitochondrial KATP channels
  - There was a significant rise in Akt phosphorylation after 3 and 6 hours, but not after 1 hour from sildenafil.
  - Neither KT5823 nor 5-HD blocked the Akt phosphorylation, indicating it occurs upstream from those effects.
- Importance of the Akt/eNOS/sGC pathway for protection
  - To look at relevance of the Akt phosphorylation, two things were tested 30 minutes before sildenafil:
    - Wortmannin (1 nmol intrastriatal): PI3K inhibitor
    - L-NIO (10 mg/kg IP): eNOS inhibitor
  - Wortmannin significantly reduced Akt phosphorylation from sildenafil, while L-NIO had no effect.
  - Neither wortmannin nor L-NIO blocked the protective nature of sildenafil.
  - Although, L-NIO was found to offer partial protection against MDMA-induced serotonin depletion.
- Testing vardenafil
  - Vardenafil (1.5 mg/kg PO) was given 30 minutes before MDMA
  - It also provided significant neuroprotection against 5-HT depletion without affecting hyperthermia
Author interpretation
- PDE5 inhibitors like sildenafil can prevent MDMA-induced serotonin depletion without affecting hyperthermia.
- It appears the relevant pathways are: PKG activation and mitochondrial KATP channel opening
- PDE5 is known to be expressed in the cerebral cortex, hippocampus, and the basal ganglia, all of which have dense 5-HT projections.
- PDE5 is connected to hydrolysis of cGMP and therefore regulates cGMP signalling.
  - Inhibition of PDE5 leads to downstream activation of PKG, one of the major intracellular receptors for cGMP.
    - Activation of PKG is known to induce antioxidative and antiapoptotic proteins
- The mechanism of relevance for KATP channel opening isn’t clear.

THC/cannabinoid system

(Tourino, 2010) – THC is protective (in mice)

Mice
- Wild-type (WT) or deficient in CB1 or CB2 receptors
Drugs
- MDMA (20 mg/kg IP) was given four times, every 2 hours
- THC (3 mg/kg IP) was administered one hour before each MDMA injection
- AM251, a CB1 antagonist, and AM630, a CB2 antagonist, were also administered at 1 mg/kg 15 minutes before each THC injection.
Animals were killed 48 hours after the last MDMA dose
Results
- THC reverses hyperthermia from MDMA at 21°C
  - MDMA led to a significant increase in body temperature
  - THC led to a significant reduction in body temperature
  - THC given before MDMA blocked the hyperthermia
- THC prevents microglia and astrocyte activation, seemingly via CB1.
  - MDMA led to marked glial activation
  - THC completely prevented microglia and astrocyte activation normally induced by MDMA
  - THC failed to perform in CB1 and CB1-CB2 mutant mice
  - THC partially suppressed microglial activation in CB1 knockout mice, with a similar impact on astrocyte activation.
- MDMA didn’t cause visiable damage to striatal DA terminals at 21°C
  - The structure of the striatum and tyrosine hydroxylase weren’t affected by MDMA at room temperature..
- THC reverses hyperthermia at 26°C
  - MDMA administration at 26°C led to significant hyperthermia
  - THC alone led to significant hypothermia
  - THC preadministration led to attenuation of MDMA-induced temperature increase, bringing the animals to a temperature similar to those given saline.
  - Analysis of the room and warm temperature conditions found warm temperatures potentiated the hyperthermic effect of MDMA but didn’t affect the hypothermic effect of THC.
  - AM251, a CB1 antagonist, blocked the effect of THC on body temperature and failed to block MDMA-induced hyperthermia
  - THC still induced hypothermia and prevented MDMA-induced hyperthermia when AM630, a CB2 antagonist, was given.
- THC prevents microglia and astrocyte activation at 26°C
  - THC suppressed MDMA-induced microglia and astrocyte activation in animals housed at warm temperature. .
- THC protects against DA terminal loss at 21°C, seemingly via CB1.
  - Unlike at room temperature, MDMA led to a notable alteration of the striatum and tyrosine hydroxylase levels were significantly reduced.
  - THC pretreatment led to normal levels of tyrosine hydroxylase
  - THC was only protective in wild-type and CB2 knockout mice, not in CB1 and CB1-CB2 deficient mice.
- MDMA didn’t result in tryptophan hydroxylase or SERT levels changes on its own in the striatum, prefrontal cortex, or hippocampus.
Interpretation
- THC at 3 mg/kg (which was intended to be a dose consumed by regular moderate cannabis users) could be useful in inhibiting neurotoxicity from MDMA
- It’d be particularly useful given cannabis is used by the majority of MDMA users
  - This is not the case for other hypothermic agents that also seem to inhibit neurotoxicity.

Memantine

(Chipana, 2008) – Protective against neurotoxicity

Using striatal synaptosomes from mice and in vivo tests in rats
Results
- Synaptosomes
  - MDMA (50 uM) led to ROS production that was entirely inhibited by Memantine (0.3 uM).
  - Memantine’s positive effect was entirely inhibited by PNU 282987 (0.5 uM), a specific agonist of alpha-7 nAChr.
- In vivo in rats
  - MDMA leads to persistent hyperthermia
    - Memantine pretreatment doesn’t prevent hyperthermia
  - MDMA also leads to reduced SERT binding in the hippocampus in animals killed at 24 hours and 7 days
    - Memantine fully prevents the reduced SERT binding at both time points

MK-801

(Miller, 1995) – Protective effect (in mice)

Mice
Drugs
- d-MDMA (20, 30, or 40 mg/kg SC) – Four injections, each 2 hours apart
- d-Fenfluramine (25 mg/kg SC) – Four injections, each 2 hours apart
- MK-801 (1.0 mg/kg SC) in some tests, given 30 min before first and third MDMA dose.
Results
- Impact of time
  - All MDMA doses, but not those of Fenfluramine, led to higher striatal GFAP
    - Over the next 21 days, these levels slowly declined toward baseline.
  - The rise in GFAP was accompanied by a decline in dopamine and tyrosine hydroxylase
    - 75% decline in both, with no recovery by 21 days.
  - Striatal serotonin was reduced by MDMA and fenfluramine, but returned to control levels by 21 days.
- Effect of lowering ambient temperature
  - 22°C
    - MDMA led to a significant rise in core temp.
    - Significant rise in striatal GFAP (over 300%) and lower DA (over 75% decline)
  - 15°C
    - Core temperature didn’t go above control mice at 22°C
    - No change in either striatal GFAP or DA
- Effect of MK-801
  - MK-801 pretreatment fully blocked MDMA neurotoxicity in both male and female mice.

Zingiber officinale (ginger)

(Mehdizadeh, 2012) – Ginger exerts protective effects

Rats
Groups
- Placebo
- MDMA (10 mg/kg IP) for one week
- Ginger (100 mg/kg IP) 4 hours before each MDMA dose
Ginger made with:
- 500 grams of dried rhizome powder extracted with 3 liters of 70% aqueous ethanol using the percolation method
- Filtered and then evaporated to dryness under reduced pressure
- Yielded a 33.28% dried extract
Learning test on the day after the last administration.
Results
- Effect on learning memory in Morris water maze
  - Ginger group had a significant decrease in the escape latency compared to the MDMA group.
  - Ginger group had a significant decrease in traveled distance to the escape platform compared to MDMA.
  - Ginger group spent significantly more time in the target quarter where the platform was located compared to MDMA group.
- Effect on neuronal density in CA1 hippocampus
  - MDMA significantly decreased neuronal density in the CA1 hippocampus.
  - Ginger led to a relative increase compared to MDMA in density, but it wasn’t significant.
- Effect on Bax and Bcl-2 protein expression
  - MDMA upregulated Bax and downregulated Bcl-2.
  - Ginger significantly decreased Bax expression compared to MDMA and it led to more Bcl-2 expression, though not significant.
- Effect on Bax and Bcl-2 gene expression
  - Ginger significantly reduced Bax gene expression increase compared to MDMA.
  - And the bcl-2 gene showed increased expression in ginger group compared to the MDMA group.
Author interpretation
- Appears ginger is protective against memory deficits and measures of brain damage in rats from MDMA.

Other Factors

Exercise

(Taghidazeh, 2016) – Exercise shows a protective effect against changes

Rats
Groups
- MDMA (5, 10, and 15 mg/kg IP) 30 minutes before each Morris water maze (MWM) trial each day
- Exercise for 30 min per day for 2 or 4 weeks at 5 days/week
- Exercise + MDMA, with exercise weeks completed prior to MDMA trials
- Control
Results
- Memory test
  - MDMA (5, 10, and 15 mg/kg) significantly increased latency and travel distance on the 4 MWM days, and decreased time spent in target quadrant in probe test of MWM, showing impairment in spatial memory.
  - Exercise at both durations prevented the increased latency and distance, as well as the decline in time spent in the target quadrant with 5 mg/kg of MDMA.
  - Only 4 weeks of exercise significantly attenuated the spatial memory impairment at 10 and 15 mg/kg of MDMA.
- Brain mitochondrial function
  - Mitochondrial ROS formation
    - MDMA at all doses significantly increased ROS formation in brain mitochondria.
    - 4 weeks of exercise significantly inhibited the rise in ROS formation at 5 mg/kg of MDMA, and significantly decreased the ROS formation at 10 and 15 mg/kg.
  - Mitochondrial membrane potential
    - Dose-related reduction in MMP with MDMA.
    - 4 weeks of exercise significantly inhibited the MMP collapse from 5 mg/kg MDMA, while there a decrease in the collapse at 10 and 15 mg/kg.
  - Swelling
    - All doses led to dose-dependent rise in mitochondrial swelling.
    - 4 weeks of exercise significantly suppressed mitochondrial swelling at 5 mg/kg, with a strong decline in swelling at 10 and 15 mg/kg.
  - Outermembrane damage
    - Dose-dependent damage in outer membrane
    - 4 weeks of exercise strongly prevented damage from 5 mg/kg, and significantly decreased the damage at 10 and 15 mg/kg.
  - Cytochrome c release
    - Dose-dependent significant rise in cytochrome c release in MDMA group
    - 4 weeks of exercise significantly inhibited the cytochrome c release from 5 mg/kg, and it strongly decreased the release at 10 and 15 mg/kg.
  - ADP/ATP ratio
    - Brain ADP/ATP ratio was increased in dose-dependent manner with MDMA
    - 4 weeks of exercise significantly attenuated the change in ratio
Author interpretation
- Physical exercise can significantly reduced MDMA’s negative effect on memory and oxidative measures
- MDMA would normally lead to impairment on spatial learning and memory
- Duration of exercise plays a role
- Other research in vitro has found NAC can significantly attenuate a drop in neuronal ATP from 5-glutathion-s-yl-n-methyl-a-methyldopamine (Capela, 2006)

Stress

(Johnson, 2009) – Raising basal glucocorticoid levels can enhance MDMA’s negative effects

Rats
Background info
- Prior exposure to chronic unpredictable stress (CUS) has been found to enhance methamphetamine hyperthermia.
- MDMA is known to raise cortisol/corticosterone (CORT) levels.
- Stress-induced CORT increases can cause neuronal atrophy in the hippocampus and cortex.
Regimen
- Rats stressed for 10 days, with variable stressors twice per day, providing CUS exposure.
Dosing
- Metyrapone (inhibiting CORT synthesis) was given at 50 mg/kg IP at 15 min before each stressor during CUS.
- Day after last stressor, 5 mg/kg IP was given of MDMA.
  - Either once or every 2 hours for 4 total injections.
For cortisol measurement, rats were killed 1 hour post-MDMA (about 18 hours post-CUS)
For serotonin/DA measurements, rats were killed 5 days after receiving 5 mg/kg four times.
Results
- Effect of CUS and metyrapone on basal CORT
  - Stress led to a significant rise in basal CORT
  - Metyrapone pretreatment significantly blocked the rise in basal CORT
- Effect of CUS and metyrapone on plasma CORT after MDMA
  - MDMA causes a significant increase in plasma CORT
  - Regardless of pretreatment, every group given MDMA had a significant CORT rise.
  - No difference between stressed and non-stressed rats in the increase from MDMA.
- Effect of CUS on MDMA’s impact on temperature
  - MDMA significantly increased temperature
  - Prior exposure to CUS led to a greater hyperthermic response
- Effect of CUS on MDMA’s reduction of serotonin and dopamine in the striatum, hippocampus, and frontal cortex
  - Striatum
    - Serotonin depletion was enhanced by CUS, with it being 55.5% greater compared to non-stressed rats given MDMA.
    - There was no depletion of dopamine in non-stressed rats, but there was in stressed rats.
  - Hippocampus
    - Serotonin depletion was enhanced by CUS, with it being 56% greater compared to non-stressed rats.
  - Frontal cortex
    - Serotonin depletion was enhanced by CUS, with it being 44% greater than in non-stressed rats.
    - There was no significant impact on dopamine concentrations.
- Effect of CUS and metyrapone on MDMA-induced hyperthermia
  - There was no difference in baseline temperature between the pretreated groups receiving either vehicle or metyrapone before stressors.
  - MDMA led to an increase in temperature in both stressed and non-stressed rats
  - CUS increased the impact of MDMA
  - Metyrapone significantly attenuated the stress-induced enhancement of MDMA’s hyperthermia.
- Effect of CUS, metyrapone, and MDMA on serotonin and dopamine concentrations
  - After 5 mg/kg four times, 5 days later
    - Striatum
      - Serotonin
        MDMA significantly depleted serotonin
        CUS increased the depletion from MDMA
        Metyrapone pretreatment blocked the enhancement of MDMA’s depletion from CUS.
        Stress itself didn’t significantly affect serotonin concentrations.
      - DA
        MDMA caused a significant depletion of dopamine only in stressed rats
        That depletion was blocked by metyrapone
    - Hippocampus
      - MDMA significantly depleted serotonin
      - CUS increased the depletion
      - Metyrapone blocked the enhancement of depletion from CUS
    - Frontal cortex
      - Serotonin
        MDMA significantly depleted serotonin
        CUS increased the depletion
        Metyrapone blocked the enhancement of depletion
      - DA
        No impact of metyrapone, stress, or MDMA on frontal cortex dopamine concentrations.
Author interpretation
- CUS for 10 days can enhance some of the negative aspects of MDMA.
- It appears higher basal CORT levels are responsible.

Temperature

It’s wise to avoid hot environments while on MDMA. If you’re in a hot environment, you should at least take regular breaks during which you’re away from crowds and no longer dancing.

(Malberg, 1998) – Different temperatures led to different core temperatures and serotonergic responses.

Methodology
- Exposed to MDMA at 20, 22, 24, 26, 28, and 30°C.
- Rats killed 2 weeks after administration.
Dose
- 20 or 40 mg/kg (SC)
Results
- Effects on core temperature
  - Ambient temperature itself didn’t impact core temperature.
  - 20 and 22°C
    - Hypothermia in MDMA animals vs controls
  - 28 and 30°C
    - Hyperthermia in MDMA animals vs controls
- Impact on neurotoxicity
  - 20, 22, and 24°C
    - No significant depletion of serotonin.
  - 26°C
    - Significant depletion of serotonin
      - 20 mg/kg
        90% of control in somatosensory cortex
      - 40 mg/kg
        87% of control in frontal cortex, 80% of control in hippocampus, and 72% of control in striatum.
        82% of control in somatosensory cortex
  - 28°C
    - Significant depletion of serotonin
      - 20 mg/kg
        80% of control in somatosensory cortex, 75% of control in hippocampus, and 71% of control in striatum.
      - 40 mg/kg
        75% of control in frontal cortex, 66% of control in somatosensory cortex, 65% of control in hippocampus, and 61% of control in striatum.
  - 30°C
    - Significant depletion of serotonin
      - 20 mg/kg
        74% of control in striatum, 70% of control in somatosensory cortex, 63% of control in frontal cortex, and 58% of control in hippocampus.
      - 40 mg/kg
        70% of control in frontal cortex, 66% of control in striatum, 65% of control in somatosensory cortex, and 42% of control in hippocampus.

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Category Topics

Adam
Here is a good page on MDMA neurotoxicity, two opposing points of view… http://ecstasy.org/info/interviews.html
- Seth Fitzgerald
  This is not really an opposing point of view, at least not an informed one. The TDC content on this topic covers Ricaurte’s work and does not dispute the findings in his primary studies, yet the interview is from 1995, meaning the vast majority of the research on MDMA’s toxicity was conducted after the interview was published.
  - Adam
    Yes its quite old bit I’m not sure what you mean by ‘this is not an opposing point of view’ given the title of the article and the content? Others have questioned MDMA neurotoxicity, see section 6 of this article from 2011…
    https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4058674/
    Most studies are with poly-drug users so how can we be sure that MDMA alone and not some other drug or the impurities are causing problems. I’ve taken many doses of MDMA in many forms from aniseed smelling crystal to odorless product of very high purity. The smelly crystal crap is most likely neurotoxic to some degree, but the pure clean stuff felt almost benign to me. But maybe thats just how it is for me.
    Not sure about G.Ricaurte’s work given his retracted article on MDMA when Methamphetamine was used instead. Also the DEA connection is of concern.
    Hell maybe I’ve taken too much and I’m bias!
    If you know of any definitive studies on neurotoxicity then please met me know.
    - Seth Fitzgerald
      I’m noting it’s not really an opposing view because the TDC page and video are essentially literature reviews/analyses and relatively little opinion is expressed, so the article isn’t an opposing review of the literature, for example.
      Perhaps you have taken the content as having more of an opinion than it is meant to? I do not take a hard stance on the neurotoxicity topic, I just explain the literature appears to show there is some toxic potential that poses relatively little clear risk when used moderately. That’s about as far as I would go in giving a view on the topic, so the bulk of the video/page are just a non-opinion review of a large portion of the available studies so that people can make up their own opinion on MDMA in an informed way.
      - Adam
        My statement of ‘two opposing points of view’ was directed purely at the content contained within the article, not ‘an’ opposing point of view of this page, I see what you were getting at now but that was not what I intended to convey. Just thought it was interesting and worth a read. I understand the context of this page and its reporting on evidence rather than giving a definitive opinion.
        If you are aware of Ricaurte’s connection to the DEA it may be wise to question his motivation. The Portman Group for example funded the Leah Betts poster campaign, their motivation was to curb MDMA usage as it was depleting alcohol sales at the time.
        We really need cognition studies of people given pure lab grade MDMA on a frequent basis (imagine the size of the application list for that!), hopefully MAPS can help. My experience with very pure MDMA on a once a week basis was just a disappearance of effects over time (almost to the degree of a built in fail-safe of sorts), no problems with cognition, sleep, anxiety etc. But my experience with weekly use of ‘street grade’ MDMA produced some problems for sure, albeit quite mild. Again maybe all just unique to me.
        There may be some neurotoxic potential over time, but I’m not sure any studies have definitively concluded this only suggested it. It might be too good to be true that a drug that takes all the best bits of other drugs and puts them in one place is also not neurotoxic, but lets hope so!
        Thanks for taking the time to reply.