A. Richard Green

The Pharmacology and Clinical Pharmacology of 3,4-Methylenedioxymethamphetamine (MDMA, “Ecstasy”)

The amphetamine derivative (±)-3,4-methylenedioxymethamphetamine (MDMA, ecstasy) is a popular recreational drug among young people, particularly those involved in the dance culture. MDMA produces an acute, rapid enhancement in the release of both serotonin (5-HT) and dopamine from nerve endings in the brains of experimental animals. It produces increased locomotor activity and the serotonin behavioral syndrome in rats. Crucially, it produces dose-dependent hyperthermia that is potentially fatal in rodents, primates, and humans. Some recovery of 5-HT stores can be seen within 24 h of MDMA administration. However, cerebral 5-HT concentrations then decline due to specific neurotoxic damage to 5-HT nerve endings in the forebrain. This neurodegeneration, which has been demonstrated both biochemically and histologically, lasts for months in rats and years in primates. In general, other neurotransmitters appear unaffected. In contrast, MDMA produces a selective long-term loss of dopamine nerve endings in mice. Studies on the mechanisms involved in the neurotoxicity in both rats and mice implicate the formation of tissue-damaging free radicals. Increased free radical formation may result from the further breakdown of MDMA metabolic products. Evidence for the occurrence of MDMA-induced neurotoxic damage in human users remains equivocal, although some biochemical and functional data suggest that damage may occur in the brains of heavy users. There is also some evidence for long-term physiological and psychological changes occurring in human recreational users. However, such evidence is complicated by the lack of knowledge of doses ingested and the fact that many subjects studied are or have been poly-drug users.

Studies, using in vivo microdialysis, on the effect of the dopamine uptake inhibitor GBR 12909 on 3,4-methylenedioxymethamphetamine ('ecstasy')-induced dopamine release and free radical formation in the mouse striatum

The present study examined the mechanisms by which 3,4‐methylenedioxymethamphetamine (MDMA) produces long‐term neurotoxicity of striatal dopamine neurones in mice and the protective action of the dopamine uptake inhibitor GBR 12909. MDMA (30 mg/kg, i.p.), given three times at 3‐h intervals, produced a rapid increase in striatal dopamine release measured by in vivo microdialysis (maximum increase to 380 ± 64% of baseline). This increase was enhanced to 576 ± 109% of baseline by GBR 12909 (10 mg/kg, i.p.) administered 30 min before each dose of MDMA, supporting the contention that MDMA enters the terminal by diffusion and not via the dopamine uptake site. This, in addition to the fact that perfusion of the probe with a low Ca2+ medium inhibited the MDMA‐induced increase in extracellular dopamine, indicates that the neurotransmitter may be released by a Ca2+‐dependent mechanism not related to the dopamine transporter. MDMA (30 mg/kg × 3) increased the formation of 2,3‐dihydroxybenzoic acid (2,3‐DHBA) from salicylic acid perfused through a probe implanted in the striatum, indicating that MDMA increased free radical formation. GBR 12909 pre‐treatment attenuated the MDMA‐induced increase in 2,3‐DHBA formation by approximately 50%, but had no significant intrinsic radical trapping activity. MDMA administration increased lipid peroxidation in striatal synaptosomes, an effect reduced by approximately 60% by GBR 12909 pre‐treatment. GBR 12909 did not modify the MDMA‐induced changes in body temperature. These data suggest that MDMA‐induced toxicity of dopamine neurones in mice results from free radical formation which in turn induces an oxidative stress process. The data also indicate that the free radical formation is probably not associated with the MDMA‐induced dopamine release and that MDMA does not induce dopamine release via an action at the dopamine transporter.

3,4‐Methylenedioxymethamphetamine increases interleukin‐1β levels and activates microglia in rat brain: studies on the relationship with acute hyperthermia and 5‐HT depletion

3,4‐Methylenedioxymethamphetamine (MDMA) administration to rats produces acute hyperthermia and 5‐HT release. Interleukin‐1β (IL‐1β) is a pro‐inflammatory pyrogen produced by activated microglia in the brain. We examined the effect of a neurotoxic dose of MDMA on IL‐1β concentration and glial activation and their relationship with acute hyperthermia and 5‐HT depletion. MDMA, given to rats housed at 22°C, increased IL‐1β levels in hypothalamus and cortex from 1 to 6 h and [3H]‐(1‐(2‐chlorophenyl)‐N‐methyl‐N‐(1‐methylpropyl)3‐isoquinolinecarboxamide) binding between 3 and 48 h. Increased immunoreactivity to OX‐42 was also detected. Rats became hyperthermic immediately after MDMA and up to at least 12 h later. The IL‐1 receptor antagonist did not modify MDMA‐induced hyperthermia indicating that IL‐1β release is a consequence, not the cause, of the rise in body temperature. When MDMA was given to rats housed at 4°C, hyperthermia was abolished and the IL‐1β increase significantly reduced. The MDMA‐induced acute 5‐HT depletion was prevented by fluoxetine coadministration but the IL‐1β increase and hyperthermia were unaffected. Therefore, the rise in IL‐1β is not related to the acute 5‐HT release but is linked to the hyperthermia. Contrary to IL‐1β levels, microglial activation is not significantly modified when hyperthermia is prevented, suggesting that it might be a process not dependent on the hyperthermic response induced by MDMA.

Elevation of Ambient Room Temperature has Differential Effects on MDMA-Induced 5-HT and Dopamine Release in Striatum and Nucleus Accumbens of Rats

3,4-Methylenedioxymethamphetamine (MDMA) produces acute dopamine and 5-HT release in rat brain and a hyperthermic response, which is dependent on the ambient room temperature in which the animal is housed. We examined the effect of ambient room temperature (20 and 30°C) on MDMA-induced dopamine and 5-HT efflux in the striatum and shell of nucleus accumbens (NAc) of freely moving rats by using microdialysis. Locomotor activity and rectal temperature were also evaluated. In the NAc, MDMA (2.5 or 5 mg/kg, i.p.) produced a substantial increase in extracellular dopamine, which was more marked at 30°C. 5-HT release was also increased by MDMA given at 30°C. In contrast, MDMA-induced extracellular dopamine and 5-HT increases in the striatum were unaffected by ambient temperature. At 20°C room temperature, MDMA did not modify the rectal temperature but at 30°C it produced a rapid and sustained hyperthermia. MDMA at 20°C room temperature produced a two-fold increase in activity compared with saline-treated controls. The MDMA-induced increase in locomotor activity was more marked at 30°C due to a decrease in the activity of the saline-treated controls at this high ambient temperature. These results show that high ambient temperature enhances MDMA-induced locomotor activity and monoamine release in the shell of NAc, a region involved in the incentive motivational properties of drugs of abuse, and suggest that the rewarding effects of MDMA may be more pronounced at high ambient temperature.

Effect of repeated ('binge') dosing of MDMA to rats housed at normal and high temperature on neurotoxic damage to cerebral 5-HT and dopamine neurones.

The technique of ‘binge’ dosing (several doses in one session) by recreational users of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) requires evaluation in terms of its consequences on the acute hyperthermic response and long-term neurotoxicity. We examined the neurotoxic effects of this dosing schedule on 5-HT and dopamineneurones in the rat brain. When repeated (three) doses of MDMA (2, 4 and 6 mg/kg i.p.) were given 3 h apart to rats housed at 19 °C, a dose-dependent acute hyperthermia and long-term loss of 5-HT was observed in several brain regions (hippocampus, cortex and striatum), with an approximate 50% loss following 3 × 4mg/kg and 65% decrease following 3 × 6mg/kg. No decrease in striatal dopamine content was detected. When MDMA (4 mg/kg i.p.) was given repeatedly to rats housed at 30 °C, a larger acute hyperthermic response than that observed in rats treated at 19 °C environment was seen (maximum response 2.6 ± 0.1 °C versus 1.3 ± 0.2 °C). A long-term cerebral 5-HT loss of approximately 65% was also detected in both the cortex and hippocampus, but no loss in striatal dopamine content occurred. These data emphasize the increased acute hyperthermic response and neurotoxicity which occurs when MDMA is administered in a hot room environment compared to normal room temperature conditions, and support the view that MDMA is a selective 5-HT neurotoxin, even when a binge dosing schedule is employed and the rats are present in a hot environment.

Behavioural and neurochemical comparison of chronic intermittent cathinone, mephedrone and MDMA administration to the rat

The synthetic cathinone derivative, mephedrone, is a controlled substance across Europe. Its effects have been compared by users to 3,4-methylenedioxymethamphetamine (MDMA), but little data exist on its pharmacological properties. This study compared the behavioural and neurochemical effects of mephedrone with cathinone and MDMA in rats. Young-adult male Lister hooded rats received i.p. cathinone (1 or 4 mg/kg), mephedrone (1, 4 or 10mg/kg) or MDMA (10mg/kg) on two consecutive days weekly for 3 weeks or as a single acute injection (for neurochemical analysis). Locomotor activity (LMA), novel object discrimination (NOD), conditioned emotional response (CER) and prepulse inhibition of the acoustic startle response (PPI) were measured following intermittent drug administration. Dopamine, 5-hydroxytryptamine (5-HT) and their major metabolites were measured in striatum, frontal cortex and hippocampus by high performance liquid chromatography 7 days after intermittent dosing and 2h after acute injection. Cathinone (1, 4 mg/kg), mephedrone (10mg/kg) and MDMA (10mg/kg) induced hyperactivity following the first and sixth injections and sensitization to cathinone and mephedrone occurred with chronic dosing. All drugs impaired NOD and mephedrone (10mg/kg) reduced freezing in response to contextual re-exposure during the CER retention trial. Acute MDMA reduced hippocampal 5-HT and 5-HIAA but the only significant effect on dopamine, 5-HT and their metabolites following chronic dosing was altered hippocampal 3,4-dihydroxyphenylacetic acid (DOPAC), following mephedrone (4, 10mg/kg) and MDMA. At the doses examined, mephedrone, cathinone, and MDMA induced similar effects on behaviour and failed to induce neurotoxic damage when administered intermittently over 3 weeks.

MDMA: On the translation from rodent to human dosing

The recent paper in this journal by Goni-Allo et al. (2008) was a welcome addition to the literature on the effects of MDMA in rodents because it examined functional changes and related them to the systemic exposure (e.g., plasma concentrations) of the drug. Such pharmacodynamic– pharmacokinetic (or quantitative pharmacology) studies are vital if we are to attempt to relate preclinical findings to the possible acute and long-term consequences of human ingestion of MDMA. The debate on whether preclinical findings on the serotonergic neurotoxicity induced by MDMA in the rodent brain can be extrapolated to human recreational usage has engaged scientists’ minds for around 20 years. Concerns have been raised as to whether the administered dose of MDMA typically used to cause neurotoxicity in rats allows any translational projections to be made as to the doses required to produce similar damage in the brains of humans following recreational use of the drug. These concerns are discussed in this short article. In order to extrapolate doses used in animal studies to those in man it has been suggested by some (McCann and Ricaurte 2001) that the technique of interspecies scaling (Mordenti and Chappell 1989) should be used. Based on similar exposure (AUC, Css) to MDMA in rats and humans, this proposes that using the equation Dhuman1⁄4Danimal Whuman=Wanimal ð Þ (where D is dose in milligram and W is weight in kilogram) allows calculation of equivalent doses in animals and humans. Accordingly, the dose of 20 mg/kg in rats becomes equivalent to a human dose of 280 mg (4 mg/kg) or somewhat over three ecstasy tablets. Other investigators (Sessa and Nutt 2008) have intimated that a dose of (for example) 20 mg/kg given by intraperitoneal injection to rats can be directly extrapolated and therefore proposed that a similar oral dose is required by human users to achieve a similar effect. So, a 20 mg/kg dose in rats is deemed “equivalent” to a 1,400 mg dose (20 mg/kg× 70 kg body weight) which is around 20 ecstasy tablets. Of course, neither of these approaches has been shown to be valid for MDMA. Furthermore, the common practice of relating the pharmacological response directly to the administered dose is basically flawed. In examining the pharmacodynamics of a specific compound, factors like bioavailability, active metabolites, plasma protein binding differences, and pattern of systemic exposure can all play a major role in determining the onset, intensity, and duration of final effect. Since the exposure patterns of MDMA and active metabolite(s) can vary markedly between species, they confound any simple interpretation on a drug effect at any specific dose in one species producing a quantitatively similar effect in another, since it is impossible for all of the administered substance (at any stated dose) to be responsible for the observed pharmacological effect. For intelligent interpretation of any data collected, it is important at the very least to have a measurement of “exposure” of parent and potentially active metabolites and by that we mean the AUC or average concentration within a dosing interval or the peak plasma concentration that occurs following drug administration. Ideally, this means the unbound plasma concentration. This still fails to take into account the half-life of the drug, plasma protein binding (which can even change with plasma drug concentration in the same species), and the pharmacological action of active Psychopharmacology (2009) 204:375–378 DOI 10.1007/s00213-008-1453-8

Repeated administration of subconvulsant doses of GABA antagonist drugs

Repeated subconvulsant doses of the GABA antagonist drugs picrotoxin (5 mg/kg), pentylenetetrazol (PTZ) (30 mg/kg), and bicuculline (3.5 mg/kg), were given IP once daily to rats. Picrotoxin produced rapid kindling to full seizures in about 5 days, PTZ produced sporadic myoclonic seizures after 17 days whereas bicuculline only produced occasional mild jerking. Following these treatments, seizure thresholds to these drugs were measured by an IV infusion method to minimise problems of systemic uptake and metabolism of the drugs. Picrotoxin-and PTZ- treated animals showed no alteration in seizure threshold. However, following bicuculline pretreatment, seizure threshold was raised. Methodological problems in the interpretation of pharmacological kindling are discussed.

5-Hydroxytryptamine and Other Indoles in the Central Nervous System

Although this review concentrates on the biochemistry and pharmacology of 5-hydroxytryptamine (5-HT) and related indolealkylamines in the central nervous system, any physiological role which these substances have occurs within the constraints of the gross anatomy, microanatomy, and ultrastructural organization of neuronal tissue. Consideration of their synthesis, compartmentation, release, inactivation, metabolism, and pharmacological action should when possible always take into account these structural constraints. As stated elsewhere, “The brain is not a plastic bag filled with a solution of diffusing neurohumors” (Grahame-Smith, 19736). The early studies of Amin et al. (1954) and Bogdanski et al. (1957) showed that 5-HT was unevenly distributed in the brain, and a great deal of work has been done to plot this regional distribution (see Erspamer, 1966; Dahlstrom et al., 1973).

MDMA: fact and fallacy, and the need to increase knowledge in both the scientific and popular press

The recreational drug 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) has elicited substantial interest over the last few years not only from preclinical and clinical scientists, but also from major parts of the media such as newspapers and television. Scientific interest can be gauged simply by examining the number of papers appearing in journals. In 1986, 1 year after the US Drug Enforcement Administration classified MDMA as a Schedule 1 drug due to its high abuse potential and lack of clinical application, a total of eight papers appeared in the literature. Around 30 papers per year were published through most of the 1990s, but the number then climbed rapidly and is currently around 100 per year (Fig. 1). The studies reported cover many aspects of the medical science, including molecular neurochemistry, experimental psychology, neuropharmacology, clinical psychopharmacology and toxicology. However, there is still much we do not know about the drug, despite the very active research efforts of many scientists and this is partly due to its complex pharmacological actions. The interest of the media and the public is perhaps harder to understand. In the UK there are around 12–15 deaths a year in persons who have taken MDMA. Given the fact that around 500,000 young persons ingest the drug in a very uncontrolled way every week in this country, these figures do not indicate MDMA to be a particularly toxic compound, regrettable though the death of every young person is. In contrast, around 100 deaths a year occur from solvent abuse, and again these deaths involve young people. Why does 10 times the number of deaths per year from solvent abuse receive so much less interest than those involving MDMA? One wonders whether it is because MDMA use is generally associated with happy young people enjoying themselves in the social situation of dance clubs or “raves”, which contrasts markedly with solvent abuse, an activity often envisaged as taking place in small isolated groups of users when they are present in unattractive surroundings. The reason I raise this point is that the media interest has resulted in much nonsense about MDMA being presented. It has been stated that it is a very dangerous drug when taken acutely (which, as noted above, is not supported by the evidence). It is sometimes even proposed that any dose is potentially fatal, thereby ignoring the fact that MDMA, like all other drugs, obeys normal pharmacological principles. Acute adverse effects (including death) are related to the dose ingested, the frequency of dosing, the ingestion of other drugs and perhaps the environmental conditions in which the user was present. The last point is of some importance, given the evidence that hot, crowded conditions increase acute amphetamine-induced lethality in animals, an observation first made over 60 years ago (Gunn and Gurd 1940) and MDMA-induced lethality is also increased by grouping animals or housing them in hot conditions (Malberg and Seiden 1998; Fantegrossi et al. 2003). Nevertheless, it is true that the acute response to the drug in humans can be somewhat unpredictable. This is probably because