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Mephedrone in the Rat: Mechanisms of Action
and Adverse Consequences
_______________________________
Craig Motbey (BSc)
A thesis submitted in fulfilment
of the requirements for the degree of
Doctor of Philosophy
School of Psychology
University of Sydney
November, 2012
i
Table of Contents
List of Figures ............................................................................................................................... vi
List of Tables ................................................................................................................................vii
Peer-reviewed Papers.................................................................................................................. viii
Conference Presentations............................................................................................................ viii
General Readership Publications................................................................................................. viii
Selected Mass Media Coverage and Interviews ........................................................................... ix
Abbreviations................................................................................................................................. xi
Acknowledgements...................................................................................................................... xii
Abstract ....................................................................................................................................... xiii
1. Introduction............................................................................................................................ 1-1
1.1. Origin of Mephedrone (MMC)............................................................................................. 1-2
1.2. Spread of MMC.................................................................................................................... 1-3
1.3. Consequences of MMC ....................................................................................................... 1-6
1.4. Structure of MMC.............................................................................................................. 1-11
1.5. Experimental Approach...................................................................................................... 1-12
1.6. References.......................................................................................................................... 1-16
2. Patterns of Brain Activation .................................................................................................. 2-1
Mephedrone (4-methylmethcathinone,‘meow’): acute behavioural effects and distribution of
Fos expression in adolescent rats.............................................................................................. 2-2
2.1. Abstract ................................................................................................................................ 2-4
2.2. Introduction.......................................................................................................................... 2-4
2.3. Experimental Procedures...................................................................................................... 2-5
Subjects ................................................................................................................................. 2-5
Drugs .................................................................................................................................... 2-5
Locomotor Activity............................................................................................................... 2-6
Social Preference................................................................................................................... 2-6
Immunohistochemistry.......................................................................................................... 2-6
Counting of Labelled Cells.................................................................................................... 2-6
ii
Data Analysis......................................................................................................................... 2-6
2.4. Results.................................................................................................................................. 2-7
Locomotor Activity............................................................................................................... 2-7
Social Preference................................................................................................................... 2-7
c-Fos Immunoreactivity....................................................................................................... 2-10
2.5. Discussion .......................................................................................................................... 2-11
Effects of MMC on Locomotor Activity and Social Behaviour......................................... 2-11
Effects of MMC on c-Fos Expression................................................................................. 2-13
Dose and Developmental Considerations............................................................................ 2-14
Conclusions......................................................................................................................... 2-14
2.6. References.......................................................................................................................... 2-15
3. Influence on Neurotransmitter Systems ............................................................................... 3-1
Mephedrone in Adolescent Rats: Residual Memory Impairment and Acute but Not Lasting 5-
HT Depletion............................................................................................................................... 3-2
3.1. Abstract ................................................................................................................................ 3-4
3.2. Introduction.......................................................................................................................... 3-4
3.3 Experimental Procedures....................................................................................................... 3-5
Subjects ................................................................................................................................. 3-5
Drugs and Doses.................................................................................................................... 3-5
Experiment 1: Effects of Acute and Repeated METH and MMC on Locomotor Activity, DA
and 5-HT........................................................................................................................... 3-6
Procedure.......................................................................................................................... 3-6
Apparatus ......................................................................................................................... 3-6
Neurotransmitter and Metabolite Analysis....................................................................... 3-7
Autoradiography .............................................................................................................. 3-7
Experiment 2: Long-term Residual Effects of MMC............................................................ 3-7
Behavioural Tests.............................................................................................................. 3-7
Elevated Plus Maze........................................................................... .......................... 3-7
Social Preference.......................................................................................................... 3-7
Novel Object Recognition............................................................................................ 3-7
iii
Neurotransmitter and Metabolite Analysis....................................................................... 3-7
Data Analysis.................................................................................................. ...................... 3-7
3.4. Results ................................................................................................................................. 3-7
Drug Analysis........................................................................................................................ 3-7
Experiment 1.......................................................................................................................... 3-7
Weight Gain..................................................................................................................... 3-7
Locomotor Activity......................................................................................................... 3-7
Neurotransmitter Levels.................................................................................................. 3-7
Autoradiography Results................................................................................................. 3-8
Experiment 2.......................................................................................................................... 3-9
Long Term Residual Effects of MMC............................................................................. 3-9
Neurotransmitter Levels.................................................................................................. 3-9
3.5. Discussion ............................................................................................................................ 3-9
3.6. References.......................................................................................................................... 3-12
4. Potential for Addiction ......................................................................................................... 4-1
High Levels of Intravenous Mephedrone (4-Methylmethcathinone) Self-Administration In
Rats: Neural Consequences and Comparison with Methamphetamine................................... 4-2
4.1. Abstract ................................................................................................................................ 4-4
4.2. Introduction.......................................................................................................................... 4-4
4.3. Experimental Procedures...................................................................................................... 4-5
Subjects.................................................................................................................................. 4-5
Drugs..................................................................................................................................... 4-5
Surgery .................................................................................................................................. 4-5
Post-Operative Procedures.................................................................................................... 4-6
Apparatus............................................................................................................................... 4-6
Procedure............................................................................................................................... 4-6
Autoradiography.................................................................................................................... 4-7
Neurotransmitter and Metabolite Analysis............................................................................ 4-7
Data Analysis......................................................................................................................... 4-7
4.4. Results ................................................................................................................................. 4-8
iv
Drug Analysis........................................................................................................................ 4-8
Behavioural Data................................................................................................................... 4-8
Initial Training.................................................................................................................. 4-8
Active Pokes................................................................................................................. 4-8
Inactive Pokes............................................................................................................... 4-8
Infusions....................................................................................................................... 4-8
Activity......................................................................................................................... 4-8
Body Weight ................................................................................................................ 4-8
Dose Response FR1.......................................................................................................... 4-9
Active Pokes................................................................................................................. 4-9
Inactive Pokes............................................................................................................. 4-10
Infusions..................................................................................................................... 4-10
Activity....................................................................................................................... 4-11
Body Weight .............................................................................................................. 4-11
Dose Response Under PR Schedule................................................................................ 4-11
Active Pokes............................................................................................................... 4-11
Inactive Pokes............................................................................................................. 4-11
Infusions..................................................................................................................... 4-11
Activity....................................................................................................................... 4-11
Body Weight .............................................................................................................. 4-13
Maximal Dosing Phase................................................................................................... 4-13
Autoradiography............................................................................................................. 4-13
Neurotransmitter and Metabolite Analysis..................................................................... 4-13
4.5. Discussion .......................................................................................................................... 4-13
4.6. References ......................................................................................................................... 4-15
5. General Discussion ................................................................................................................ 5-1
5.1. Understanding the Mechanisms of MMC............................................................................ 5-2
5.2. Responding to MMC............................................................................................................ 5-8
5.3. Dangerous Possibilities: The Example of MPTP............................................................... 5-11
5.4. Conclusions........................................................................................................................ 5-13
v
5.5. References.................................... ..................................................................................... 5-14
6. Appendix: Statements from Co-Authors ............................................................................... 6-1
vi
List of Figures
Chapter 1
Figure 1. Life-threatening Necrotizing Fasciitis Due to ‘Bath Salts’ Injection.......................... 1-8
Figure 2. Structural Similarities Between Amphetamines and Cathinones.............................. 1-13
Chapter 2
Figure 1. Schematic Diagram of Brain Regions Counted.......................................................... 2-8
Figure 2. Locomotor and Stationary Activity During Locomotor Test...................................... 2-9
Figure 3. Representative Locomotor Paths During Locomotor Test.......................................... 2-9
Figure 4. Social Interaction Time and Locomotor Activity During Social Preference Test...... 2-9
Figure 5. Representative Images of c-Fos Immunoreactivity in the Medial Striatum.............. 2-11
Figure 6. Representative Images of c-Fos Immunoreactivity in the Nucleus Accumbens....... 2-12
Chapter 3
Figure 1. Weight Gain Over the Dosing Period.......................................................................... 3-7
Figure 2. Locomotor Activity on First and Last Days of Dosing............................................... 3-8
Figure 3. Residual Effects on Elevated Plus Maze and Social Preference............................... 3-10
Figure 4. Residual Effects on Novel Object Recognition......................................................... 3-10
Chapter 4
Figure 1. Self-administration Behaviour During the Initial Training Period............................ 4-10
Figure 2. Self-administration Behaviour During Fixed Ratio Dose Response......................... 4-11
Figure 3. Self-administration Behaviour During Progressive Ratio Dose Response................ 4-13
Chapter 5
Figure 1. Survivors of the MPTP epidemic.............................................................................. 5-12
vii
List of Tables
Chapter 2
Table 1. Counts of Fos-positive Cells in Regions Significantly Different From Vehicle......... 2-10
Table 2. Counts of Fos-positive Cells in Regions Not Significantly Different From Vehicle.. 2-12
Chapter 3
Table 1. Test Sequence Employed in Experiment 2.................................................................... 3-7
Table 2. Acute Effects on Neurotransmitter and Metabolite Levels........................................... 3-8
Table 3. Autoradiographic Measures........................................................................................... 3-9
Table 4. Residual Effects on Neurotransmitter and Metabolite Levels..................................... 3-11
Chapter 4
Table 1. Mean (SEM) Autoradiography Results........................................................................4-14
Table 2. Mean (SEM) Striatal Neurotransmitter and Metabolite Levels…….......................... 4-14
viii
Peer-reviewed Papers
Motbey CP, Hunt GE, Bowen MT, Artiss S, Mcgregor IS (2011) Mephedrone (4-
methylmethcathinone, ‘meow’): acute behavioural effects and distribution of Fos
expression in adolescent rats. Addiction Biology 17(2): 409-422.
Motbey CP, Karanges E, Li KM, Wilkinson S, Winstock AR, Ramsay J, Hicks C, Kendig MD,
Wyatt N, Callaghan PD, McGregor IS (2012) Mephedrone in adolescent rats: residual
memory impairment and acute but not lasting 5-HT depletion. PLoS ONE 7(9): e45473.
Motbey CP, Clemens KJ, Apetz N, Winstock AR, Ramsey J, Li KM, Wyatt N, Callaghan PD,
Bowen MT, Cornish J, McGregor IS (2013). High levels of intravenous mephedrone (4-
methylmethcathinone) self-administration in rats: neural consequences and comparison
with methamphetamine. Journal of Psychopharmacology. Epub ahead of print June 5,
2013, doi: 10.1177/0269881113490325
Conference Presentations
Motbey CP, Hunt GE, Karanges E, Apetz N, Kendig M, Callaghan PD, Clemens K, Cornish J,
McGregor IS (2011) Behavioural and neural characteristics of acute and chronic
mephedrone (4- methylmethcathinone, meow) treatment in adolescent rats. Poster
presented at the 2011 International Behavioural Neuroscience Society meeting.
General Readership Publications
Motbey CP (2012) Mephedrone: what doesn’t kill you might still mess you up. The
Conversation. Available at http://theconversation.edu.au/mephedrone-what-doesnt-kill-
you-might-still-mess-you-up-6238
Motbey CP (2012) Party’s over: mephedrone causes memory impairment. The Conversation.
Available at http://theconversation.edu.au/partys-over-mephedrone-causes-memory-
impairment-9672
ix
Selected Mass Media Coverage and Interviews
Mephedrone an “ecstasy, meth” combo. Published online at
http://www.nzdoctor.co.nz/news/2011/october-2011/28/mephedrone-an-%27ecstasy,-
meth%27-combo.aspx on 28 October 2011.
Meow meow has rats firing. Published online at http://www.theage.com.au/national/meow-
meow-has-rats-firing-20111027-1mm64.html on 28 October 2011.
Party drug lights up rodent brains. Published online at
http://www.australasianscience.com.au/article/issue-januaryfebruary-2012/party-drug-
lights-rodent-brains.html in January 2012.
Mephedrone may lead to permanent brain damage. Published online at
http://www.doctortipster.com/11287-mephedrone-may-lead-to-permanent-brain-
damage.html on 19 September 2012.
Party drug users risking memory loss. Published online at
http://www.scoop.co.nz/stories/GE1209/S00078/party-drug-users-risking-memory-
loss.htm on 19 September 2012.
Drug study suggests lasting brain damage. Published online at
http://www.nzdoctor.co.nz/news/2012/september-2012/19/drug-study-suggests-lasting-
brain-damage.aspx on 19 September 2012.
Party drug damaging to memory? Published online at
http://articles.timesofindia.indiatimes.com/2012-09-20/health/33976464_1_party-drug-
mephedrone-high-doses on 20 September 2012.
Party drug causes memory impairment. Published online at
http://www.indiatvnews.com/news/world/party-drug-causes-memory-impairment-
8919.html on 20 September 2012.
x
Dangerous synthetic drugs becoming more widespread. Published online at
http://www.reportageonline.com/2012/10/dangerous-synthetic-drugs-becoming-more-
widespread/ on 22 September 2012.
“Killer” party pill becoming popular. Published online at
http://www.odt.co.nz/news/national/226954/killer-party-pill-becoming-popular on 22
September 2012.
New revellers’ designer drug alarms authorities. Published online at
http://www.bayofplentytimes.co.nz/news/new-revellers-designer-drug-alarms-
authorities/1560749/ on 27 September 2012.
Mephedrone and memory loss: an interview with Craig Motbey. Published online at
http://www.news-medical.net/news/20121016/Mephedrone-and-memory-loss-an-
interview-with-Craig-Motbey.aspx on 16 October 2012.
The hamster wheel: health in the media. Available online at
http://www.youtube.com/watch?v=bXr12OzX62Y. First broadcast on ABC TV Australia
on 31 October 2012.
Popular party drug no good for memory. Available online at
http://www.theleader.com.au/story/748365/popular-party-drug-no-good-for-memory/ on
10 November 2012.
Street drug innovation leaves scientists in wake. Published online at
http://newsciencejournalism.com/01/2013/street-drug-innovation-leaves-scientists-in-
wake/ on 7 January 2013.
xi
Abbreviations
5-HT 5-hydroxytryptamine (serotonin)
DA Dopamine
DAT Dopamine Transporter
FR Fixed Ratio
HPLC High-Performance Liquid Chromatography
IHC Immunohistochemistry
IP Intraperitoneal
IV Intravenous
MDMA 3,4-methylenedioxymethamphetamine (Ecstasy)
MDPV Methylenedioxypyrovalerone
METH Methamphetamine
MMC 4-methylmethcathinone (Mephedrone)
MPPP 1-methyl-4-phenyl-4-propionoxypiperidine
MPTP 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine
NE Norepinephrine
NET Norepinephrine Transporter
PR Progressive Ratio
REU Regular Ecstasy Users
SERT 5-HT Transporter
xii
Acknowledgements
Jim, because I said I would.
xiii
Abstract
Mephedrone (4-methylmethcathinone, MMC) is a novel recreational drug that first made a
significant appearance in Europe in the closing years of the first decade of the 21st century. Users
reported MMC as having the stimulant-like properties of methamphetamine (METH) or cocaine,
combined with prosocial, entactogenic effects suggestive of 3,4-
methylenedioxymethamphetamine (MDMA, “Ecstasy”). Anecdotal reports also suggested that
MMC was a drug with a very high addictive potential, prone to inducing uncontrolled bingeing.
The spread of MMC was accompanied by a growing corpus of emergency room case reports
suggesting toxicity in overdose. However, at the beginning of the MMC boom there was almost
no published research available as to the acute or lasting effects, mechanism of action or
potential for toxicity. Although the research program presented in this thesis and the work of the
various competing laboratories around the world represent an effort to correct this deficiency, it
remains the case that our understanding of MMC is still relatively meagre. As such, the fact that
very large numbers of people have been consuming substantial amounts of MMC over a period
of years is of great concern.
The research presented in this thesis consisted of three laboratory studies. In the first study, we
examined the behavioural and neural effects of two different MMC doses (15 and 30 mg/kg,
intraperitoneal (IP)) relative to the well-known stimulant METH (2 mg/kg IP) in male adolescent
Wistar rats. Rats were injected, assessed for locomotor activity for 60 minutes and then tested in
a 10-minute social preference test (measuring time spent in close proximity to a real rat versus a
dummy rat). Their brains were then processed using Fos immunohistochemistry to determine
patterns of brain activation. Results showed that MMC caused profound locomotor hyperactivity
at both dose levels while tending to reduce social preference. Patterns of Fos expression with
MMC resembled a combination of those observed with METH and MDMA, with particularly
strong Fos expression in the cortex, dorsal and ventral striatum, ventral tegmental area (all
typical of both MDMA and METH) and supraoptic nucleus (typical of MDMA). These results
demonstrated for the first time the powerful stimulant effects of MMC in animal models and its
capacity to activate mesolimbic regions. These results also provided some empirical basis to user
reports that MMC subjectively resembles a MDMA/METH hybrid.
xiv
The second study consisted of two experiments. In Experiment 1 male adolescent Wistar rats
received single or repeated (once a day for 10 days) injections of MMC (30 mg/kg) or the
comparator drug METH (2.5 mg/kg). Both MMC and METH caused robust hyperactivity in the
1 h following injection, and this effect did not tend to sensitize with repeated treatment. Striatal
dopamine (DA) levels were increased 1 h following either METH or MMC while striatal and
hippocampal serotonin (5-HT) levels were decreased 1 h following MMC but not METH. MMC
caused greater increases in 5-HT metabolism and greater reductions in DA metabolism in rats
that had been previously exposed to MMC. Autoradiographic analysis showed no signs of
neuroinflammation ([125
I]CLINDE ligand used as a marker for translocator protein (TSPO)
expression) with repeated exposure to either MMC or METH. In Experiment 2, rats received
repeated MMC (7.5, 15 or 30 mg/kg once a day for 10 days) and were examined for residual
behavioral effects following treatment. Repeated high (30 mg/kg) dose MMC produced impaired
novel object recognition 5 weeks after drug treatment. However, no residual changes in 5-HT or
DA tissue levels were observed at 7 weeks post-treatment. This lasting memory dysfunction was
the first demonstration of MMC-induced cognitive impairment in the rat and provided mutual
support to a contemporaneous finding suggesting memory damage in human MMC users.
The third study explored the characteristics of intravenous MMC self-administration in the rat,
with METH again used as a comparator drug. Male Sprague-Dawley rats were trained to nose
poke for intravenous MMC or METH in daily 2 h sessions over a 10 day acquisition period
before dose-response functions were established under fixed- and progressive-ratio (FR and PR,
3 days at each of 4 doses) schedules, followed by a 3 day period of maximal dosing. Brains were
analysed for striatal 5-HT and DA levels while autoradiography assessed changes in the density
of 5-HT and DA transporters (SERT, DAT) as well as induction of the inflammation marker
Translocator Protein (TSPO) in various brain regions. Under a FR1 schedule, peak responding
for MMC was obtained at 0.1 mg/kg/infusion, versus 0.01 mg/kg/infusion for METH. Break
points under a PR schedule peaked at 1 mg/kg/infusion MMC versus 0.3 mg/kg/infusion for
METH. Final intakes of MMC were 31.3 mg/kg/day compared to 4 mg/kg/day for METH.
METH, but not MMC, self-administration caused elevated TSPO receptor density in the nucleus
accumbens and hippocampus. MMC, but not METH, self-administration caused decreased 5-
xv
HIAA levels. MMC supported high levels of self-administration, matching or exceeding those
previously reported with any other drug of abuse. This finding combined with evidence from a
variety of other sources confirms the anecdotal reports of MMC’s potential for addiction.
Overall, the evidence presented in this thesis suggests that MMC may induce lasting cognitive
impairment and carries a high risk of addiction. MMC use involves a significant risk of
morbidity and a relatively minor risk of mortality. While not amongst the most dangerous of
abused drugs, MMC does appear to be substantially more harmful than the drug it partially
displaced (MDMA). As such, the case of MMC consists of not just another example of the
failure of prohibition to prevent the harms associated with illicit drug use, but also provides a
case study for the accelerating dangers of the novel psychoactive market.
1-1
1. Introduction
_______________________________
1-2
1.1 Origin of Mephedrone (MMC)
Throughout much of the modern era, recreational and addicted drug use has been
focussed upon a few traditionally utilised substances along with a limited set of familiar
pharmaceuticals. While the consequences of this use of alcohol, cannabis, opiates,
amphetamines, barbiturates and psychedelics were sometimes tragic, they were at least
predictable. The situation, while regrettable, was relatively stable. This is no longer the case.
Rising to prominence in the early 1980’s, the phenomenon of “designer drugs” appears to
have begun with the illicit production of established medicinal drugs. The effects of many
existing drugs can be greatly enhanced or altered by slightly modifying their molecular
structures, and the tools and techniques developed by modern chemists to pursue these
possibilities eventually spread into the world of illicit drug development. Although the initial
motivation to develop novel psychoactives was probably largely driven by a desire to evade
narrowly targeted drug laws, it did not take long for the recreational drug industry to embrace the
opportunity to diversify and enhance their products (Langston and Palfreman, 1995).
Most prominent amongst the recently emerging drugs of abuse is mephedrone (MMC1).
As of early 2010, Europol reported MMC detection in twenty European member states
(European Monitoring Centre for Drugs and Drug Addiction (EMCDDA), 2010). MMC use is
also well established in Australia (Bruno et al., 2012) and the U.S.A. (US Department of Justice
Drug Enforcement Agency, 2011), and has been detected in Canada (Maheux et al., 2010) and
New Zealand (Winstock and Wilkins, 2011).
First synthesized in 1929 (de Burnaga Sanchez, 1929; EMCDDA, 2010), MMC remained
little more than a chemical footnote until it developed a following amongst recreational drug
users in the late 2000s. MMC is a substituted cathinone, with a degree of divergence from the
basic cathinone molecule similar to that between 3,4-methylenedioxymethamphetamine
1 AKA 4’-methylmethcathinone or β-keto-(4,N-dimethylamphetamine) or 4,N-dimethylcathinone or p-methyl-methcathinone or 2-
aminomethyl-1-tolyl-propan-1-one or (±)-1-(4-methylphenyl)-2-methylaminopropan-1-one or N-methylephedrone or (RS)-2-methylamino-1-(4-
methylphenyl)propan-1-one; molecular formula C11H15NO, molecular weight 177.242.
1-3
(MDMA) and amphetamine. The human use of cathinones for their psychoactive properties
traces back to prehistory. Known as khat (or some close variation thereof) or miraa in eastern
and southern Africa, the leaves and twigs of the Catha edulis shrub have traditionally been
chewed in order to experience the effects of the natural cathinone contained within (Manghi et
al., 2009). While not entirely devoid of negative consequences (Corkery et al., 2010; Manghi et
al., 2009), traditionally managed cathinone consumption nevertheless lies at the milder end of the
spectrum of stimulants of concern (Klein and Metaal, 2010).
In its most common manifestation, MMC is a white or off-white powder. This
hydrochloride salt form of MMC is both stable and water soluble. Most of the MMC so far seen
has been of extremely high purity, and is apparently the product of industrial-scale production in
Asia (Brunt et al., 2010; Dargan et al., 2011; EMCDDA, 2010; Nutt, 2011; Whalen, 2010; Wood
et al., 2010b). In addition to being sold undisguised as itself, MMC is one of a number of
ingredients that appear in drug mixes sold as “plant food” or “bath salts”. While both of these
terms apparently gained popularity as a means of evading regulations applying to products sold
for human consumption, it appears that the term “plant food” may be due to MMC’s brief use as
an experimental insecticide in Israel (Nutt, 2012).
It appears that the most common non-MMC ingredient of “bath salts” is the related
synthetic stimulant methylenedioxypyrovalerone (MDPV). While the European experience of
novel cathinones has been focussed upon MMC with a relatively minor contribution from
MDPV, the North American situation appears to be the reverse (Schneir et al., 2012). Evidence
suggests that MDPV may carry a higher risk of toxicity and lethality than MMC (Baumann et al.,
2012), and that differences in their mechanisms of action may render them particularly
dangerous in combination (Cameron et al., 2012). Like MMC, a subset of MDPV users consume
the drug by injection, and rates of IV MDPV use appear to be rising rapidly (Csák et al., 2013).
1.2 Spread of MMC
MMC developed a substantial presence on the drug markets of the industrialised world at
the close of the first decade of the 21st century. The first official detection of MMC appears to
1-4
have happened in early 2008, in the analysis of a suspect powder contained in capsules seized by
Finnish customs in late 2007 and early 2008 (EMCDDA, 2010). User reports involving MMC
also began appearing on the online “psychonaut” resource Erowid in early 2008 (Erowid, 2011).
During 2009, there was a steep increase in the frequency with which the U.K. Forensic Science
Service identified MMC and other cathinones in drug seizures, and by March 2010, these
outnumbered MDMA and piperazine seizures combined (Dargan and Wood, 2010).
The popularity of MMC grew rapidly, and was accompanied by an increase in MMC-
related hospital presentations during 2009 (James et al., 2011; Wood and Dargan, 2010). By
2010 the annual Mixmag drug survey (measuring a self-selected sample drawn from a U.K.
based nightclubbing-focused population) stated that 41.7% of their respondents reported prior
use of MMC and 33.6% had consumed the drug in the previous month. A 2010 survey of 1006
college and high school students in Scotland found that 205 (20.3%) reported consuming MMC
at least once previously (Dargan et al., 2010). As well as this intentional consumption of MMC,
there is also evidence that MMC was being substituted for MDMA: 11.5% of the “Ecstasy”
surveyed in the Netherlands in 2009 contained MMC (Brunt et al., 2010).
It has been repeatedly suggested in both the academic and popular press that China is the
root source of MMC (Brunt et al., 2010; Dargan et al., 2011; Nutt, 2011; Whalen, 2010; Wood et
al., 2010b), although the truth of this statement remains to be conclusively demonstrated
(Vardakou et al., 2010). Chinese origin was not uncommon in MMC detections to date (Brunt et
al., 2010; EMCDDA, 2010; Wood et al., 2010b), and several drug interdiction agencies have
identified China as a key source of MMC (EMCDDA, 2010; Vardakou et al., 2010; US
Department of Justice Drug Enforcement Agency, 2011). However, as the location of synthetic
drug production is geographically flexible, the potential rise of indigenous MMC-producing
industries in MMC-using countries cannot be ruled out.
It appears that a substantial driver of the use of MMC was user dissatisfaction with the
purity and availability of MDMA and cocaine, possibly created by an increase in the
effectiveness of prohibition enforcement (Advisory Council on the Misuse of Drugs, 2010;
Measham et al., 2010; Winstock and Wilkins, 2011). A variety of sources confirm that the
worldwide MDMA supply had seen a sharp decline in quality and availability (Brunt et al., 2010;
Dick and Torrance, 2010; Winstock and Wilkins, 2011). Between 2006 and 2010 MDMA
seizures by the U.K. customs service dropped from roughly 1,100 per quarter to less than one
1-5
hundred. Concurrent with this reduction in MDMA confiscation, customs seizures of MMC and
other cathinones rose from a base of essentially zero to a steeply-climbing 600 in early 2010 (just
prior to the introduction of legal controls on the possession of MMC in the U.K. during April of
that year) (Dargan and Wood, 2010).
Although the focus of MMC consumption appears to be concentrated in Europe and the
U.K., MMC has begun developing a presence worldwide. MMC use is reported to be substantial
and increasing in the U.S.A. (US Department of Justice Drug Enforcement Agency, 2011),
although it appears that their novel psychoactive market has a greater focus upon MDPV than is
the case in Europe (Baumann et al., 2012). MMC use is well established in Ireland (Van Hout
and Brennan, 2010; Van Hout and Brennan, 2011b; Van Hout and Brennan, 2011c; Van Hout
and Bingham, 2012) and across central Europe (EMCDDA, 2010; Pharris et al., 2011).
Geographic isolation appears to be little defence, with MMC detected in “ecstasy” tablets seized
in New Zealand (Winstock and Wilkins, 2011). A nationwide survey of Australian regular
ecstasy users (REU) in 2010 found that 17% reported MMC use in the previous six months. The
same survey found that rates of prior six months reported MMC use amongst REU in Tasmania
climbed from zero in 2007 to near 50% in 2010 (Bruno et al., 2012).
Media and political reaction to the rise of MMC has tended to follow the classical
progression from prurient curiosity through hyperbolic panic to knee-jerk response (Davey et al.,
2010; Forsyth, 2012; Lancaster et al., 2011; Measham et al., 2011; Sare, 2011; Schifano et al.,
2011; Silverman, 2010; Van Hout and Bingham, 2012). The U.K. tabloid campaign was notable
for the frequent referencing of fatalities that, on more thorough examination, proved to be
unrelated to MMC (Davey et al., 2010; Lancaster et al., 2011; Sare, 2011; Winterton, 2010). The
subsequent process leading to prohibition was marked by the resignation of several members of
the U.K. Advisory Council on the Misuse of Drugs (still recovering from the recent and
controversial dismissal of David Nutt (Nutt, 2009b; Nutt, 2010)) in protest at political
interference in the activity of the council (Sare, 2011). However, the role of the internet in both
the distribution of and media response to MMC differentiates this case from previous drug panics
(Forsyth, 2012; Measham et al., 2010; Schifano et al., 2011). Two factors in particular stand out.
Firstly, it is commonly observed that initial media reports on an emerging drug of abuse
effectively serve as advertisements for the use of that drug (Forsyth, 2012; Measham et al.,
2011). In the case of MMC, this phenomenon acquired a newly literal meaning, as context-
1-6
sensitive advertising servers accompanied the internet editions of early newspaper reports on
MMC with advertisements for online MMC suppliers (Forsyth, 2012; Schifano et al., 2011).
Secondly, the interactive nature of internet media appears to have somewhat altered the
dynamics of the classical drug scare campaign. The rise of drug-focussed (“psychonaut”) internet
discussion groups and the spread of commenting options on mainstream media reporting provide
a powerful platform for competing narratives to reach large-scale audiences. While it remains too
early to accurately predict what effect this may have on the MMC phenomenon, it appears likely
that this new voice may substantially modify the effect of the traditional media presentation
(Forsyth, 2012).
A significant feature of the early years of the MMC pandemic was the fact that MMC
remained legal in the U.K. and several other European countries until mid to late 2010.
Compared to the subterranean word-of-mouth spread of previous novel drugs, the ready
availability and active marketing of online and main street “research chemical” and “headshop”
suppliers gave MMC a significant boost in establishing its presence. MMC is now controlled
across the European Union (Dargan et al., 2011). In countries with broadly drawn analogue laws
such as Australia, MMC was already arguably illegal when use first surfaced (Duff, 2010).
While MMC regulation in the U.S.A. is complicated by the diversity of state legislation, by mid
2011 a majority of the states had enacted laws controlling MMC and other cathinones (US
Department of Justice Drug Enforcement Agency, 2011) and in July 2012 President Obama
signed the Synthetic Drug Abuse Prevention Act forbidding the sale of MMC, MDPV and a
number of other drugs nationwide (Gershman and Fass, 2012).
1.3 Consequences of MMC use
The reported health consequences of MMC use range from the surprisingly mild to the
alarmingly gruesome. The typical picture of “MMC gone wrong” involves a distressed MMC
consumer presenting at a hospital emergency room with a broad spectrum of sympathomimetic
symptoms (Wood et al., 2010a; Wood et al., 2010b; Wood et al., 2010c; Wood and Dargan,
2010). Clinical reports suggest users occasionally suffer from agitation, combative behaviour,
psychosis, tachycardia, hyperthermia, hypertension and seizures (Bajaj et al., 2010; Baumann et
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al., 2012; McGaw and Kankam, 2010; Wood et al., 2010a; Wood et al., 2010b; Wood et al.,
2010c). However, in the majority of cases, patients were discharged without apparent lasting
harm after a few hours of sedation and symptomatic treatment.
While the possibility of fatal MMC-linked hyperthermia has been raised in one animal
study (McGregor et al., 2011), the lack of any reported hyperthermia or lethality in similar
studies (Miller et al., 2012; Wright et al., 2012) and the scarcity of reports of fatal hyperthermia
in human users (Wood et al., 2010a) argues against over-interpreting this result. Although a
number of deaths have been linked to MMC (Dickson et al., 2010; Gustavsson and Escher, 2009;
Lusthof et al., 2011; Maskell et al., 2011; Schifano et al., 2012; Torrance and Cooper, 2011;
Wood et al., 2010a), there are few in which a causal role for MMC alone has been unequivocal
(Winstock et al., 2011). While it appears certain that MMC can be fatal to the sufficiently
unlucky or foolish, the overall impact of MMC on fatality seems remarkably slight given the
scale of use.
The most serious immediate harms of MMC are focused upon the minority of users who
consume the drug intravenously. MMC is mildly acidic in solution, which may at least partially
explain the common reports of burning sensations on insufflation (Dargan et al., 2010; Erowid,
2011) or injection (Van Hout and Bingham, 2012). However, this acidity does not appear
sufficient to explain the dramatically negative outcomes (Figure 1) seen in studies of MMC
injectors (Dorairaj et al., 2011; Russo et al., 2012; Van Hout and Bingham, 2012). One study
reported “vein blockages, […] skin erosion, localised infections, blisters, spots, cold sores,
abscesses, scabs, lumps, gangrenous tissue, blood clots and large holes at overused injecting
sites”. To quote one of their respondents: “it clots up your veins twice as fast as anything
else…that’s one thing I did learn fast from it, say you used one vein on it and got two hits out of
it, you wouldn’t get the vein again, you’d have to move onto a different vein to get another hit
out of it, that’s why everyone started using their groin” (Van Hout and Bingham, 2012: p 191). A
case series from a single hospital reported multiple cases of MMC induced injuries serious
enough to require major surgical debridement and added that “extravasation of these substances
either intentionally or accidentally appear to result in local cutaneous reactions similar to that
seen with cytotoxic drug use” (Dorairaj et al., 2011: p e39).
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Figure 1. Life-threatening Necrotizing Fasciitis Due to ‘Bath Salts’ Injection (adapted from Russo et al.
(2012)). 1: Patient at time of operative incision. 2: Photograph during forearm debridement.
The exact cause of the dramatic impact of MMC on injecting users remains to be
identified. However, the behavioural characteristics of MMC consumption appear likely to
significantly contribute to the negative outcomes for injectors. Early anecdotal reports of MMC
frequently emphasized its binge-inducing properties (Erowid, 2011). A survey of Dutch MMC
users found that 61 of 70 reported “craving for the drug” as an acute effect (Brunt et al., 2010).
An observational test of MMC intoxication in humans found that ratings on a 1-7 Likert scale of
“want MMC” rose from below 4 to approximately 6 after drug consumption (and were just
above 2 on a drug-free comparison test a week later) (Freeman et al., 2012). A report on injecting
MMC users states that “all participants described intense cravings to re-inject compulsively
within each injecting episode” (Van Hout and Bingham, 2012).
From the point of view of safer injecting practices, the hyper-confidence and risk
insensitivity associated with stimulant intoxication already provides a high-risk setting for
injecting drug use. However, unlike other drugs associated with compulsive re-dosing (e.g.:
cocaine), MMC is not a pure stimulant. The additional sensorimotor distortion induced by the
entactogenic and psychedelic aspects of MMC, when combined with the prospect of multiple
intoxicated injection events, provides an unparalleled opportunity for infection and other
injection-related harms. This is particularly the case when the drug, like MMC, is observed to be
prone to causing direct damage to injection sites (Dorairaj et al., 2011; Van Hout and Bingham,
2012). The first signs of MMC’s impact on the injecting community may already be apparent,
1-9
with a rise in central European rates of I.V. MMC use accompanied by a rise in H.I.V. detections
(Pharris et al., 2011).
Although it is apparently not the typical method, an alternative MMC synthesis route can
potentially result in manganese-contaminated product (Dargan et al., 2011). Manganese toxicity
is known to be related to movement disorders suggestive of Parkinson’s, and such toxicity has
been observed in users of other cathinones (Sikk et al., 2007; Sikk et al., 2010; Stepens et al.,
2008). While the possibility of manganese toxicity in MMC consumers has been considered and
dismissed by other authors (Dargan et al., 2011), this dismissal was based largely upon the lack
of reports of Parkinsonian symptoms amongst MMC users. Given the recent reports of
Parkinsonian symptoms in I.V. MMC users (Brennan and Van Hout, 2012; Van Hout and
Bingham, 2012), the possibility of manganese contamination in the MMC supply may need to be
reconsidered.
Behavioural evidence from human studies suggests that MMC use may induce cognitive
impairment, as human MMC users displayed substantially impaired performance on a prose
recall task even while not acutely intoxicated (Freeman et al., 2012). In this study, regular recent
MMC users (reporting more than two instances of use per month) were compared to a control
group of participants who had consumed MMC on a single occasion more than six months ago.
Psychometric testing was conducted on two consecutive weekends, with participants in the
MMC group consuming participant-provided drug during the first testing session. MMC group
participants displayed elevated depression and schizotypy scores relative to controls when drug-
free. Measures of impulsivity correlated positively with reported length of MMC-using sessions.
User reports subjective effects showed substantial increases in “self-confidence”, “buzzing”,
“dizziness” and “impaired concentration and memory” after MMC consumption.
In addition to direct neurological or other physical damage, there is also the social and
psychological toll of addiction and uncontrolled use to consider. MMC is subjectively perceived
by human users to be intensely prone to inducing compulsive use (Brunt et al., 2010; Erowid,
2011; Freeman et al., 2012; Van Hout and Bingham, 2012). A highly common feature of early
reports of MMC use was that drug consumption continued until the supply was exhausted, even
when this supply was intended to last for an extended period (Erowid, 2011). Clinical and
anecdotal reports of sustained addiction to MMC are not uncommon (Bajaj et al., 2010; Dargan
et al., 2010; Erowid, 2010; Simu et al., 2010; Van Hout and Bingham, 2012). MMC has been
1-10
shown to induce a conditioned place preference in rats, mice and flatworms (Lisek et al., 2012;
Ramoz et al., 2012) and promotes self-administration behaviour in rats (Hadlock et al., 2011,
Aarde et al., 2013; Motbey et al., 2013).
The behavioural consequences of MMC use extend beyond addiction alone. MMC use
has been associated with both violent and accidental death (Prosser and Nelson, 2011) as well as
a wide assortment of high-risk sexual behaviours (Van Hout and Brennan, 2011a). Significantly
enhanced confidence and euphoria, when combined with dizziness, disturbed concentration,
general confusion and a distorted time sense (Freeman et al., 2012), are features that can be
expected to seriously impair the risk-assessment abilities of MMC users (Borlik, 2012; Van Hout
and Bingham, 2012; Van Hout and Brennan, 2011a). Also of concern is the apparent connection
between MMC and suicide, with self-inflicted deaths making up 29% of a recent sample of
MMC-linked fatalities (Schifano et al., 2012) and elevated levels of depression reported amongst
MMC users (Freeman, 2012).
The speculation as to links between MMC and the "Miami Cannibal" case, in which a
man brutally attacked a randomly chosen victim, were based upon nothing more than the
unsupported comment of one police officer (Helberg, 2012). Media coverage of the event far
outpaced toxicological analysis, which ultimately confirmed that MMC had no role in the attack
(Hiaasen et al., 2012). However, events such as these are not altogether without precedent in the
MMC literature. One of the early MMC fatality reports was the case of a man in the Netherlands
who, under the influence of MMC and a variety of other drugs, died after smashing a number of
windows while in a “rage of fury” (Lusthof et al., 2011). As with the Miami case, the subject of
this report was also found to have stripped himself naked. Indeed, public nakedness appears to be
a fairly common theme in media reports of “bath salts” intoxication (Borlik, 2012; Shepard,
2012; Warren, 2012). MMC-induced hyperthermia likely plays a role in these events, along with
the entactogenic, disinhibiting and aphrodisiac effects of the drug. More alarmingly, it appears
that face-biting is also a recurring feature (Campbell, 2012; McCorquodale, 2012). However, the
unfounded speculation around the Miami case has likely introduced a strong selection bias into
the media coverage of MMC and the causative role of “bath salts” (MMC or not) in some of
these events is rather questionable.
1-11
1.4 Structure of MMC
The cathinone family of molecules consists of a nearly exact replication of the
amphetamines (Figure 2), with the sole exception of the addition of a ketone group at the β
position of the carbon chain (which is why the cathinones are sometimes referred to as “beta
keto-” or “BK-amphetamines”). In all of the examples studied so far (Dal Cason et al., 1997;
Patel, 2009), the psychoactive properties of the cathinones have closely resembled those of their
amphetamine “twins”, albeit with slightly reduced potency by mass (most likely due to an
impaired ability to cross the blood-brain barrier, as a result of the increased hydrophilicity
induced by the extra ketone group (Gibbons and Zloh, 2010)).
Cathinone functions similarly to amphetamine (Griffiths et al., 2010), affecting the
dopamine (DA), serotonin (5-HT) and norepinephrine (NE) systems through the stimulation of
neurotransmitter release and the modulation of reuptake. Cathinone also slows neurotransmitter
metabolism by inhibiting the action of monoamine oxidase (MAO), with a preferential action
biased more towards MAO-B than MAO-A (Osorio-Olivares et al., 2004). As MAO-B is the
primary enzyme responsible for DA breakdown, inhibition results in an increase in the synaptic
availability of DA. Chronic administration of cathinone to rats creates DA depletions similar to
those seen after chronic amphetamine or cocaine (Ellison, 2002). The cathinones have also been
shown to affect NA systems through the influence of both release (Cleary et al., 2003) and
reuptake (Cozzi et al., 1999).
The existence of well-characterized amphetamine analogues allows one to predict the
likely behavioural and neural effects of many of the cathinones. However, while its production is
not entirely unknown (Davis et al., 2012), the lack of research on the effects of 4’-
methylmethamphetamine (the amphetamine equivalent of MMC) leaves a broad scope of
investigation for researchers of MMC. While the picture has now become somewhat clearer,
there is still considerable work to be done.
1-12
1.5 Experimental Approach
Much of the information now available regarding the neural mechanisms and impacts of
MMC was provided over the last few years by a variety of research groups utilising animal
models of MMC consumption. At the initiation of the research underlying this thesis, no
previous findings had been published exploring the effects of MMC in animal models. Virtually
nothing was known about the mechanisms or dangers of this drug. For the sake of coherence and
clarity, all of the information derived from research with animal models is discussed in Chapter
5, where it is integrated with the findings of the research presented in the body of this thesis. The
literature reviews in the previously published research papers (Chapters 2-4) demonstrate the
development of MMC-related knowledge over the past few years.
The use of animal models provides a significant methodology for the study of illicit
drugs. Given the invasive nature of many neuroscientific research methods and the high risk of
neurotoxic effects with these drugs, both ethical and practical considerations limit the ability of
researchers to investigate these topics in human subjects. Animal models have long been utilised
by many researchers to explore the behavioural impact (e.g.: Clemens et al., 2006; Cornish et al.,
2008; Cornish et al., 2012; Frohmader et al., 2010; Hadlock et al., 2011; Krasnova et al., 2010;
Lisek et al., 2012; Ramoz et al., 2012) as well as the mechanisms of action (e.g.: Kehr et al.,
2012; Martinez-Clemente et al., 2011; Patel, 2009) and potential toxicity (e.g.: McGregor et al.,
2011; Miller et al., 2012; Wright et al., 2012) of illicit drug use. Although the differences
between human and non-human physiology create the need for caution when interpreting the
results of such investigations, these studies provide an essential starting point for further human-
based research.
The experimental program forming the core of this thesis consisted of three rat-based
studies, which were intended to address three basic questions. Firstly, how does MMC influence
the brain? Secondly, does MMC use damage the brain? Thirdly, is MMC addictive, and if so,
how much so?
1-13
Figure 2. Structural similarities between amphetamines and cathinones (adapted from Baumann et al.
(2012)).
The first of these studies (Chapter 2) used c-Fos immunohistochemistry to display the
pattern of brain activation induced by MMC intoxication. As Fos protein is transiently expressed
by neurons in response to strong activation, visualising the distribution of this protein allows the
equivalent of a cellular-resolution timelapse brainscan. Researchers from the University of
Sydney Psychopharmacology group have previously reported the brain-wide distribution of Fos
expression with a diverse range of party drugs including MDMA (Stephenson et al. 1999;
Hargreaves et al. 2007), cannabis (Arnold et al. 2001), methamphetamine (Carson et al. 2010)
and g-hydroxybutyric acid (GHB) (Van Nieuwenhuijzen, McGregor & Hunt 2009a).
A notable benefit of using c-Fos immunohistochemistry is that the protein requires
approximately half an hour to express. Because of this feature, any neural activity in the last half
hour of life will not be recorded by c-Fos expression. As well as removing the influence of the
perfusion process, this also allows for the possibility of excluding portions of the experimental
program from the c-Fos signal. This was done in the experiment presented in Chapter 2; although
the behavioral data from a social preference test is presented alongside the Fos results, the study
1-14
was arranged so that the experience of this test would be excluded from the c-Fos data. This
allowed for the presentation of the clearest possible picture of the basic neural effects of MMC
dosage.
The second study (Chapter 3) aimed to further explore the acute mechanisms of MMC as
well as investigating the potential for neurotoxicity. Autoradiographic tracking of the
inflammation marker TSPO (Translocator Protein) was used to investigate the possibility of an
acute inflammatory response, while High Performance Liquid Chromatography (HPLC) allowed
measurement of the influence of MMC upon DA and 5-HT systems. Acute inflammatory
responses (Pascual et al., 2007) and lasting alterations in neurotransmitter metabolism (Wagner
et al., 1980) are both known consequences of some psychoactive drug use. This analysis was
conducted on brain tissue collected shortly after dosing as well as tissue harvested from animals
given an extended (47 day) washout period. This allowed the examination of both the acute
impact of the drug as well as the possiblity of lasting neuroadaptation or neurotoxicity.
A variety of behavioural measures were also employed during the washout period,
investigating the potential lasting influence of MMC treatment upon anxiety, social behaviour
and memory. Evidence from human studies (Freeman, 2012) had raised concern regarding the
potentially harmful influence of MMC use on memory and other cognitive functions. A broad
range of pilot testing of behavioural measures was undertaken, only some of which is reported in
Chapter 3. These tests were intended to function as the preliminary screen for neurotoxicity and
other health concerns. This screen was performed in conjunction with related work by other
researchers from the University of Sydney Psychopharmacology lab and used a wide variety of
testing paradigms well established in behavioural neuroscience research.
The third study (Chapter 4) focussed upon the addictive potential of MMC use via an
extended self-administration paradigm which compared the properties of MMC to the well-
characterised stimulant METH. A substantial number of previous studies have examined the
impact of self-administration of stimulants on the brain using a variety of indices (Krasnova et
al., 2010; Letchworth et al., 2001; Norwood et al., 2003; Sim-Selley et al., 2000; Turchan et al.,
1998; Wilson and Kish, 1996; Wise et al., 1995). It has long been known that variations in
reinforcement schedule play a key role in such studies (Ferster and Skinner, 1957). Due to this
fact, both fixed and progressive ratio schedules were utilized here in the testing of MMC. Fixed
1-15
ratio schedules allow the measurement of willingness to consume: do the rats like it? In contrast,
progressive ratio schedules measure commitment to gaining reward: how much do the rats want
it? Although subtle, the exploration of differences such as these helps form a thorough
understanding of the addictive potential of a drug.
In this study, dose-response curves were generated for METH and MMC under both
fixed- and progressive-ratio reward schedules. Autoradiographic techniques were also employed
to investigate potential changes in levels of DA and 5-HT transporters as well as the
inflammation marker TSPO. HPLC analysis of striatal tissue was conducted once again in order
to explore the possible differences between the relatively brief experimenter-administered drug
treatment employed in Chapter 3 and the more sustained subject-administered drug treatment of
Chapter 4.
The conclusion of this thesis aims to place these findings into the broader context of
emerging psychoactives and explores some potential responses to this rapidly changing situation.
It will be argued that the example of MMC provides a case study for the flaws of conventional
approaches to drug control in the context of the modern psychoactive drug market.
1-16
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2-1
2. Patterns of Brain Activation
_______________________________
2-2
Mephedrone (4-methylmethcathinone,‘meow’): acute
behavioural effects and distribution of Fos expression
in adolescent rats
Craig P. Motbey, Glenn E. Hunt, Michael T. Bowen, Suzanne Artiss and Iain S. McGregor.
Addiction Biology, 17: 409-422
2-3
Co-Author Contribution
McGregor, I.S.
Provided technical assistance, contributed to the research design and manuscript editing.... 15%
Hunt, G.E.
Provided technical assistance .....................................................................................................8%
Bowen, M.T.
Provided technical assistance .....................................................................................................8%
Artiss, S.
Provided technical assistance .....................................................................................................3%
TOTAL........................................................................................................................34%
2-4
2-5
2-6
2-7
2-8
2-9
2-10
2-11
2-12
2-13
2-14
2-15
2-16
2-17
3-1
3. Influence on Neurotransmitter Systems
_______________________________
3-2
Mephedrone in Adolescent Rats: Residual Memory
Impairment and Acute but Not Lasting 5-HT Depletion
Craig P. Motbey, Emily Karanges, Kong M. Li, Shane Wilkinson, Adam R. Winstock, John
Ramsay, Callum Hicks, Michael D. Kendig, Naomi Wyatt, Paul D. Callaghan and Iain S.
McGregor.
PLoS ONE, 7(9): e45473. doi:10.1371/journal.pone.0045473
3-3
Co-Author Contribution
McGregor, I.S.
Provided technical assistance, contributed to the research design and manuscript editing.... 10%
Callaghan, P.D.
Provided technical assistance..................................................................................................... 4%
Wyatt, N.
Provided technical assistance..................................................................................................... 1%
Kendig, M.D.
Provided technical assistance..................................................................................................... 4%
Hicks, C.
Provided technical assistance..................................................................................................... 4%
Ramsay, J.
Provided technical assistance..................................................................................................... 1%
Winstock, A.R.
Provided technical assistance..................................................................................................... 1%
Wilkinson, S.
Provided technical assistance..................................................................................................... 4%
Li, K.M.
Provided technical assistance..................................................................................................... 4%
Karanges, E.
Provided technical assistance..................................................................................................... 4%
TOTAL................................................................................................................37%
3-4
3-5
3-6
3-7
3-8
3-9
3-10
3-11
3-12
3-13
4-1
4. Potential for Addiction
_______________________________
4-2
High Levels of Intravenous Mephedrone (4-
Methylmethcathinone) Self-Administration In Rats: Neural
Consequences and Comparison with Methamphetamine
Craig P. Motbey1, Kelly Clemens
2, Nadine Apetz
1, Adam R. Winstock
3, John Ramsey
4, Kong M. Li
5, Naomi
Wyatt6, Paul D. Callaghan
6, Michael T. Bowen
1, Jennifer Cornish
2, Iain S. McGregor
1
1School of Psychology, University of Sydney, NSW 2006, Australia
2School of Psychology, Macquarie University, NSW 2109, Australia
3Institute of Psychiatry, Kings College, University of London, UK
4TICTAC Communications Ltd., St George’s College, University of London, UK
5Department of Pharmacology, University of Sydney, NSW 2006, Australia
6Australian Nuclear Science and Technology Organisation, Sydney, NSW 2234, Australia
Manuscript submitted to “Journal of Psychopharmacology”
4-3
Co-Author Contribution
Cornish, J.
Provided technical assistance ...................................................................................................10%
McGregor, I.S.
Provided technical assistance, contributed to the research design and
manuscript editing .....................................................................................................................10%
Clemens, K.
Provided technical assistance .....................................................................................................8%
Bowen, M.T.
Assisted with statistical analyses................................................................................................7%
Callaghan, P.D.
Provided technical assistance .....................................................................................................2%
Li, K.M.
Provided technical assistance .....................................................................................................2%
Apetz, N.
Provided technical assistance .....................................................................................................1%
Wyatt, N.
Provided technical assistance .....................................................................................................1%
Ramsey, J.
Provided technical assistance .....................................................................................................1%
Winstock, A.R.
Provided technical assistance .....................................................................................................1%
TOTAL...................................................................................................................................... 43%
4-4
4-5
4-6
4-7
4-8
4-9
4-10
4-11
4-12
4-13
4-14
4-15
4-16
4-17
5-1
5. General Discussion
_______________________________
5-2
5.1 Understanding the Mechanisms of MMC
Overall, the findings presented here and elsewhere have been largely in accord with what
would be predicted based upon the anecdotal reports of MMC consumers regarding the euphoric,
entactogenic and stimulant properties of the drug. Users frequently described MMC using
analogies to MDMA and METH (Erowid, 2012). Both MDMA and METH induce significant c-
Fos expression in the prelimbic cortex, caudate-putamen, Islands of Calleja, nucleus accumbens
core, ventral tegmental area and dorsal raphe. METH additionally activates the ventral orbital
cortex, lateral orbital cortex, barrel fields 6a/6b, and primary motor cortex, while MDMA
activates the nucleus accumbens shell, supraoptic nucleus, anterior paraventricular thalamic
nucleus, central and basolateral amygdala, Edinger-Westphal nucleus and lateral parabrachial
nucleus. MMC induces significant levels of c-Fos expression in all of these areas.
The pattern of brain activation demonstrated by c-Fos expression in MMC-dosed rats
resembles an MDMA/METH hybrid, in accord with early descriptions of the drug (Erowid,
2012). Unsurprisingly therefore, MMC intoxication has a strong acute impact upon NE, DA and
5-HT systems (Hadlock et al., 2011; Kehr et al., 2011; Motbey et al., 2012; Shortall et al., 2013).
Acute MMC induces a rapid and substantial efflux of all three of these neurotransmitters
(Baumann et al., 2012; Baumann et al., 2012; Cameron et al., 2012; Kehr et al., 2011; Wright et
al., 2012), and in all three cases this release is mediated at least partially by the reversal of
transporter function (Baumann et al., 2012). However, it appears that both the speed of release
and the pace of metabolism may substantially differ between these neurotransmitter systems
(Kehr et al., 2011; Motbey et al., 2011). MMC effects upon DAT function appear to be similar to
those of METH (Cameron et al., 2012), in that both drugs induce a depolarizing current in DAT,
thereby acting as an excitatory ligand and driving DA release. In contrast, the related synthetic
cathinone MDPV (like MMC, a common component of “bath salts” and “plant food” drugs)
produces a hyperpolarizing current in DAT (an effect similar to that seen with cocaine),
inhibiting DA reuptake (Cameron et al, 2012; Baumann et al., 2013). However, the effects of
MMC and MDPV on DA levels are synergistically enhanced when these two drugs are given in
combination, due to the differing temporal pattern of each drug’s effects (Cameron et al., 2013).
5-3
MMC appears to carry a substantial potential for addiction and uncontrolled bingeing
(Hadlock et al., 2011; Lisek et al., 2012; Motbey et al., 2013; Ramoz et al., 2012). User reports
frequently refer to the addictive potential of the drug (eg: Erowid, 2009; Erowid, 2010) and a
diverse range of animal models of addictive behaviour have displayed positive results with
MMC (Hadlock et al, 2011; Lisek et al., 2012; Motbey et al., 2013; Ramoz et al., 2012;
Robinson et al., 2012). Accumulating evidence from multiple sources strongly suggests that
MMC consumption may lead to lasting cognitive impairment (den Hollander et al., 2012;
Freeman et al., 2012; Motbey et al., 2012). However, no researcher has yet provided an
explanation as to the mechanism of this cognitive impairment or any direct evidence of
significant MMC-induced neurotoxicity. We also do not yet possess any evidence to explain the
differing temporal patterns of MMC’s expression or metabolism.
Behavioural observations in the first experiment of this thesis (Chapter 2; Motbey et al.,
2011) suggested thigmotaxic effects, which may relate to the entactogenic reports from human
consumers. This suggestion was further supported by the strong c-Fos expression found in the
somatosensory barrel fields of MMC-dosed rats, as primary sensory areas such as these could be
expected to be stimulated by entactogenic or thigmotaxic intoxication. MMC in humans appears
to combine entactogenic with pro-social effects (Erowid, 2012). As with MDMA, it may be the
case than any prosocial effects of MMC are mediated by the oxytocin system (Green et al.,
2003). However, no study has yet directly addressed the question of MMC-related oxytocin and
vasopressin expression or their influence on MMC-driven behaviour. Pursuing this question
provides an obvious future target for MMC research.
The c-Fos expression demonstrated in the supraoptic nucleus (SON) of rats, an area rich
in both oxytocin and vasopressin (Kohno et al., 2008), also suggests a potential interaction of
MMC with these systems. Activation in areas associated with oxytocin receptors (such as the
central amygdala) also provides support. However, the levels of SON activation found in MMC-
dosed rats were relatively mild compared to those typically found with MDMA or GHB
(Hargreaves et al., 2007; van Nieuwenhuijzen et al., 2010), suggesting that the interaction of
MMC with these systems may be of a lower intensity than that seen with these other drugs.
The extremely strong c-Fos expression demonstrated in the striatum of MMC-dosed rats
suggested a 5-HT/dopamine mode of action. This suggestion was later confirmed by the research
5-4
described in Chapter 3 of this thesis (Motbey et al., 2012). Neurochemical analysis of brains
extracted from MMC-dosed rats 1 h post injection revealed contrasting effects on DA and 5-HT.
In both the striatum and hippocampus, 5-HT levels were substantially lowered (by approximately
30%) while 5-HIAA/5-HT ratios (a measure of 5-HT metabolic turnover) were increased. In
contrast, striatal DA levels were significantly increased while measures of DA metabolism
((DOPAC+HVA)/DA ratios) were decreased.
These findings suggested that MMC may induce a very rapid efflux of 5-HT (largely
metabolized by the 1 h post dosing point) and a more gradual and sustained increase in DA,
possibly by inhibition of reuptake or metabolism. Such a hypothesis is consistent with other
evidence that MMC acts preferentially as a 5-HT releaser, but also induces a vigorous efflux of
DA (Baumann et al., 2011, Baumann et al., 2012). Evidence from a number of studies has
demonstrated that MMC blocks the uptake of NE, DA and 5-HT (Hadlock et al., 2011; Lopez-
Arnau et al., 2012; Martinez-Clemente et al., 2012; Simmler et al., 2012), and a recent release-
assay study (Baumann et al., 2012) showed that MMC functions as a non-selective transporter
substrate, actively stimulating the release of NE, DA and 5-HT. Given these facts, it follows that
there must be some alteration in metabolic rates in order to explain the contrasting pictures of
DA and 5-HT levels from the brains of rats examined 1 hr after MMC dosing (Motbey et al.,
2012; Chapter 3). However, the close investigation of the mechanisms required to clarify exactly
what mechanism is driving these differences remains to be done.
One plausible mechanism for the reduction in DA metabolism would be a selective
inhibition of monoamine oxidase B (MAO-B). Indeed, research suggests that the basic cathinone
molecule does act in this manner (Kelly, 2011). However, an investigation of the interaction of
monoamine oxidases with a wide variety of cathinones revealed that MMC does not substantially
impact MAO-A or B (Osorio-Olivares et al., 2004). Therefore, any MMC-induced inhibition of
DA metabolism must utilise an alternative mechanism, possibly via the next step in the
metabolic process, aldehyde dehydrogenase (ALDH). This potential for MMC-induced
inhibition of ALDH raises the possibility that MMC and alcohol may produce a synergistic
neurotoxicity similar to that seen with MDMA (Izco et al., 2007; Upreti et al., 2009). There is
also some suggestion that the combination of MMC and alcohol may induce cardiac impairment
(McGaw and Kankam, 2010), although there is no current evidence to suggest that this is more
5-5
than a reversible acute effect of the drugs. Due to the influence of MMC on cardiac function
(Nicholson et al., 2010; Vardakou et al., 2010; Wood et al., 2010a), an investigation has been
made into the potential for MMC-induced cardiac injury in the rat (Meng et al., 2011). While this
study confirmed the strong acute effects of MMC on cardiac function, it did not find any
evidence of lasting harm.
Several authors have investigated the possibility of lasting neurotoxicity or
neuroadaptation in response to both acute and chronic MMC consumption (Hadlock et al., 2011;
Angoa-Pérez et al., 2012). In agreement with the results presented here in Chapter 3 (Motbey et
al., 2012), a group of mice given a binge-style treatment of 4 x 40mg/kg I.P. MMC (doses at 2
hour intervals) failed to display any signs of toxicity, inflammation or any lasting alteration in
neurotransmitter systems 7 days afterwards (Angoa-Pérez et al., 2012). However, a study in
which rats were dosed with 4 x 25 mg/kg S.C. MMC (also at 2 hour intervals) while maintained
at substantially elevated ambient temperatures (≥ 27° C) did find lasting decreases in
hippocampal 5-HT uptake when tested 7 days after treatment (Hadlock et al., 2011).
While this research may present an important lead towards the discovery of MMC-
induced neurotoxicity, the complicating variable of temperature and contrasting results of other
studies (Angoa-Pérez et al., 2012; Motbey et al., 2012; Simmler et al., 2012) constrains the
breadth of conclusions to be drawn from it. The interaction of MMC with temperature is
particularly problematic, as studies have shown that the effect of MMC on thermoregulation can
be substantially altered by variations in dosage and environmental conditions. While singly-
housed rats given one relatively low dose of MMC have demonstrated a hypothermic response
(Shortall et al., 2013), studies utilising binge-dosing protocols with group-housed animals have
shown hyperthermic responses at both normal and elevated ambient temperatures (Baumann et
al., 2011; Hadlock et al., 2011). In addition, elevated ambient temperatures and group housing
have both been shown to abolish the hypothermic response even in singly-dosed animals (Miller
et al., 2012; Shortall et al., 2013). Recent work has also revealed strain-based differences in
thermal response (Aarde et al., 2013, Wright et al., 2012). As variations in ambient temperature
and thermoregulatory impairments have been shown to have a significant influence upon
neurotoxicity with other stimulants (Malberg and Seiden, 1998), careful consideration of this
variable is necessary when investigating the toxicity of such drugs.
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Of the research in this thesis, the results presented in chapters 2 and 3 derive from Wistar
rats while those of chapter 4 involved Sprague-Dawley animals. Although later work has shown
that MMC-based self-administration characteristics do not sharply differ between Wistar and
Sprague-Dawley rats (Aarde et al., 2013), the strain-based diversity of MMC responses shown in
thermoregulatory and other measures (Aarde et al., 2013, Wright et al., 2012) suggests that this is
a factor that deserves consideration during further investigation.
In the rat, MMC binds to the 5-HT, NE and DA transporters (SERT, NET and DAT) with
a selectivity and potency similar to MDMA, reversing the action of these transporters and
driving neurotransmitter release (Baumann et al., 2011). Similar results have been found with
transporter-transfected human embryonic kidney cells (Simmler et al., 2012) and human DAT-
expressing Xenopus laevis oocytes (Cameron et al., 2012). The influence of MMC on 5-HT, NE
and DA has been demonstrated by an increasing variety of studies (Baumann ete al., 2011;
Hadlock et al., 2011; Kehr et al., 2011; Motbey et al., 2012, 2013; Shortall et al., 2013). Acute
MMC treatment evokes dose-related increases in 5-HT efflux in the nucleus accumbens, again
similar to MDMA on a per-mg basis. DA efflux in the nucleus accumbens after MMC treatment
appears to exceed that found with MDMA, albeit not to the extent seen with methamphetamine
(METH) (Baumann et al., 2011; Kehr et al., 2011; Wright et al., 2012).
Although MMC has a profound acute influence on 5-HT and DA systems, no lasting
alteration could be detected in the levels of these neurotransmitters or their metabolites 47 days
post-treatment (Chapter 3, Motbey et al., 2012). This finding was somewhat surprising given the
evidence of persistent effects upon neurotransmitter systems with similar drugs such as MDMA
(Beaumann et al., 2007). While this result is in agreement with the findings of Angoa-Pérez et al.
(2012) and Simmler et al. (2012), it does present a contrast with the decreases in rat hippocampal
5-HT levels 7 days post-treatment discovered by Hadlock et al. (2011). The recent work of den
Hollander et al. (2012) further complicates the pictures, with their finding of an MMC-induced
reduction in HVA two weeks after a four-day binge treatment in mice. However, a group of rats
given similar treatment in the same study showed no significant neurochemical changes, in
agreement with the findings presented in Chapter 3 of this thesis (Motbey et al., 2012). Although
it provides useful early evidence of possible species differences in the MMC response, the
reduction of HVA in den Hollander’s (2012) mice does little to clarify the picture regarding
MMC neurotoxicity.
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However, the possibility of at least short-term MMC-induced alteration in
neurotransmitter levels was supported by the results presented in Chapter 4 (Motbey et al.,
2013). Striatal tissue collected 3 days post-treatment from rats given 44 days of access to self-
administered MMC was found to have significantly reduced levels of 5-HIAA. However, the
finding of a lack of lasting alteration in 5-HIAA levels amongst rats given brief chronic MMC
(Motbey et al., 2012) suggests that this 5-HIAA reduction may have been destined to fade if
given more than 3 days of recovery. Although it appears that any lasting influence of MMC
consumption on neurotransmitter systems is likely to be subtle, the possibility of such an effect
should not be ruled out. Further research is obviously required to clarify this point.
Regardless of whether or not the mechanism is as straightforward as a simple alteration in
neurotransmitter levels, there is significant behavioural evidence that MMC use can induce
cognitive impairment. Rats given 10 days of 30 mg/kg I.P. MMC show a clear deficit in
recognition memory when tested 35 days post-dosing (Motbey et al., 2012; Chapter 3). This
finding is in agreement with the prose recall impairment displayed by human MMC users
(Freeman et al., 2012), and has since been further supported by the finding of an MMC-induced
impairment of working memory in mice when tested by a T-maze spontaneous alternation task
two weeks after a four day binge treatment (den Hollander et al., 2012). These findings in
combination present the clearest evidence to date regarding MMC neurotoxicity or adverse
neuroadaptation. Although the direct physical cause has yet to be located, it appears clear that
MMC consumption can induce cognitive impairment, and that this impairment may be persistent.
Identifying the mechanisms behind this cognitive impairment is a clear priority for further
research.
The evidence from animal studies also supports the anecdotal reports of MMC’s
addictiveness (eg: Erowid, 2011; 2012). MMC has been shown to induce a conditioned place
preference in rats, mice and flatworms (Lisek et al., 2012; Ramoz et al., 2012; Motbey et al.,
2013) and maintain self-administration behaviour in rats (Hadlock et al., 2011; Aarde et al.,
2013). Levels of MMC-seeking responses in rats (Motbey et al., 2013; Chapter 4) given
extended access to 2 h day MMC nosepoke-triggered self administration dramatically exceeded
those seen in methamphetamine-dosed comparison animals. This response was achieved with no
pretraining and may represent the most vigorous self-administration response ever found with
any drug. Under progressive ratio testing conditions, rats continued to work for MMC infusions
5-8
even when response/reward ratios rose into the hundreds (Motbey et al., 2012). MMC induces
high levels of c-Fos activation across the mesolimbic dopamine pathway (Motbey et al., 2011)
and stimulates a rapid and substantial release of DA and 5-HT into the nucleus accumbens in rats
(Kehr et al., 2011). Overall, the evidence suggests that MMC is a drug with an extremely high
risk of uncontrolled consumption.
Intravenous MMC carries a high danger of extremely serious physical harm (Dorairaj et
al., 2011; Pharris et al., 2011; Russo et al., 2012; Van Hout and Bingham, 2012). The exact cause
of these harms are as yet unclear, but they appear to be due to a combination of the behavioural
characteristics of MMC use (i.e.: repeated dosing episodes while intoxicated, leading to poor
injection hygeine) and an innate toxicity of the drug (Dorairaj et al., 2011).
5.2 Responding to MMC
The evidence as to the impact of legal restrictions on MMC use is mixed. Although
prohibition in the U.K. has driven an increase in the price of MMC (Winstock et al., 2010a), a
survey of London club-goers in mid 2010 (3 months after prohibition) found that 27% reported
use or intention to use on the night of the survey (Measham et al., 2011). A follow-up survey on
the same site one year later reported that this figure had risen to 41% (Wood et al., 2012). In both
cases, MMC was the most consumed drug reported on the night. Participants in the second
survey were also questioned as to their favourite recreational drug; MMC was the leading
contender, taking 20.4% of the vote (ahead of cocaine at 14.9%) (Wood et al., 2012). It appears
likely that MMC and other synthetic cathinones will remain part of the contemporary drug
culture for some time to come (Brennan and Van Hout, 2012).
While it may have ended its phase of rapid growth, MMC still maintains a substantial
presence amongst UK drug users (Archer et al., 2013). Australian rates of MMC consumption
appear to have remained relatively slight (Sindicich and Burns, 2012) except amongst some
small samples drawn from a population of regular drug users (Bruno et al., 2012). However,
reports from around the world show that MMC use continues to be popular in many areas
regardless of legality (Archer et al., 2012; Brennan and Van Hout, 2012; Bruno et al., 2012;
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Brunt et al., 2010; US Department of Justice Drug Enforcement Agency, 2011; Wood et al.,
2012).
Some recent data suggests that MMC use in the UK has ceased increasing and may have
begun to decline (Measham et al., 2012). However, this study was deliberately targeted upon a
sample of “mainstream” nightclubs rather than the dance-culture venues typical of much earlier
research. It is unclear how much of the apparent decline is due to this factor, as lower rates of
novel psychoactive use are not unexpected amongst this subject group relative to the drug early-
adopters of the dance music community. If there is a genuine decline, it is also unclear what
proportion relates to increasing legal restriction versus an increased awareness within the drug
using community of the undesirable effects of MMC consumption or competitive displacement
by newly-arriving novel psychoactives.
The reports of injection-related harms occuring in human populations (Dorairaj et al.,
2011; Pharris et al., 2011; Russo et al., 2012; Van Hout and Bingham, 2012) suggest that
responses to MMC should include a strong emphasis on discouraging intravenous use. While
non-injecting use of MMC is not without danger, the dramatic impact of MMC on injecting users
justifies a focus upon harm minimization and diversion to less damaging methods of
consumption.
There may also be worthwhile harm minimization strategies to be pursued with non-
injecting users. For example, the tendency of MMC to induce uncontrolled bingeing suggests
that minimizing on-hand supplies of the drug may be a useful strategy. However, while MMC
bingeing may be reduced if users acquire their drugs via relatively smaller (although possibly
more frequent) purchases, such an approach increases the exposure of users to the legal and
physical risks of illicit drug purchasing transactions. The effect of prohibition is to encourage
fewer, larger purchasing transactions and promote stockpiling, which in turn provides the setting
for increased binge consumption.
Although it has been suggested that the rise of novel psychoactive drugs provides an
opportunity to “consider the trial of alternative policy and legislative approaches to drug control”
(Winstock and Wilkins, 2011; Hughes and Winstock, 2012), it appears more likely that the
response to MMC will follow the traditional patterns of simplistic prohibition that are so widely
seen to be counterproductive (United Nations Office on Drugs and Crime, 2009; Winstock et al.,
2010b; Winstock and Wilkins, 2011; Australia21, 2012; Hughes and Winstock, 2012). Legal
5-10
prohibition and the interdiction of precursor chemicals may eventually prove effective in limiting
supply, as appears to have been the case with MDMA (Winstock and Wilkins, 2011). However,
even if the prohibition of MMC does prove to be successful in reducing use, such an approach
provides a temporary respite at best.
A large number of easily manufactured cathinone variants exist, most of which are likely
to be psychoactive to some degree. The flexibility of modern chemistry allows for an almost
inexhaustible supply of alternative molecules. In 2010 alone, European authorities identified 41
new psychoactive drugs (EMCDDA, 2011). Just the contents of the phenethylamine family
(Shulgin, 1991) are enough to occupy the global research community for decades, to say nothing
of the cathinones, the tryptamines and so on.
To date, the main reaction has been to reflexively ban each new molecule as it appears on
the market, frequently before any adequate evidence has been presented as to whether the
potential harms of the substance justify such a response (Hughes and Winstock, 2012; Measham
et al., 2010). This approach creates the perverse risk of driving users towards an endless stream
of novel drugs, many of which may be more harmful than their already-established alternatives.
Although evidence as to relative risk is still being collected, it appears that this is exactly what
has happened with the substitution of MMC for MDMA consumption. While there is still no
conclusive evidence of MMC-driven neurotoxicity, it appears clear that MMC induces persistent
cognitive impairment and carries an extremely high risk of addiction and excessive consumption.
While MDMA use is not without dangers of its own (Burgess et al., 2000), it appears clear that it
is a lower-risk option than MMC.
There is no reason to believe that prohibition is likely to be any more successful with the
continuously expanding list of new drugs than it has been with the drugs of the 20th
century. An
effective response to the fast-moving realities of the modern drug market is likely to require a
radical departure from business as usual. The development of systems to facilitate the rapid
assessment and provisional classification of emerging drugs appears to be a clear priority
(Winstock and Ramsey, 2010; Hughes and Winstock, 2012). However, any system of provisional
classification followed by later refinement necessarily implies the occasional lessening of
restrictions on drugs found to be relatively less harmful. The reality of the current drug debate
renders such an action politically unpalatable to governments in many countries. Therefore, it is
essential that such classification bodies be rigorously independent of political influence if they
5-11
are to function as intended (Nutt, 2009a; Nutt, 2009b; Reuter, 2011; Winstock and Wilkins,
2011). The recent series of controversies involving the U.K. Advisory Council on the Misuse of
Drugs (Nutt, 2009b; Nutt, 2010; Sare, 2011) provide a stark example of the failure of such
systems if this requirement is not met.
5.3 Dangerous Possibilities: The Example of MPTP
The history of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) suggests a chilling
possibility that argues for urgent action in response to this situation. In 1982, the accidental
production of MPTP by illicit chemists in the U.S.A. led to a micro-epidemic of drug-induced
Parkinsonism (Ballard et al., 1985; Langston and Palfreman, 1995). Closely related to the
synthetic heroin substitute MPPP (1-methyl-4-phenyl-4-propionoxypiperidine, an “enhanced”
variant of pethidine which was the intended product of most of the illicit chemists responsible for
this incident), MPTP is a highly potent and selective dopaminergic neurotoxin that is commonly
used today to create animal models of Parkinson’s disease (Jackson-Lewis et al., 2012).
However, the dangers of MPTP were almost entirely unknown to science in 1982 (Langston and
Palfreman, 1995), much as is the case with many emerging drugs of abuse today. The first
warning of the MPTP incident came when a wave of young heroin addicts appeared in a variety
of hospitals with mysteriously rapid and premature symptoms of Parkinson’s disease (Figure 1).
5-12
Figure 1. Survivors of the MPTP epidemic. This photo was taken in 1991, 9 years after the poisoning
incident. George Carillo (standing left) and Juanita Lopez (standing right) had achieved a substantial
degree of recovery following fetal-tissue transplants. Bill Silvey (seated left), Connie Sainz (seated
centre) and David Silvey (standing right) were still profoundly impaired by their illness. Image from
Langston and Palfreman (1995, p150).
5-13
While the history of MPTP was tragic for those directly affected, from the point of view
of the wider community the incident could be seen as a fortunate escape from calamity. The
small scale of MPTP production and the rapid action of MPTP toxicity allowed the situation to
be detected and contained before it affected more than a relatively small number of people
(Langston and Palfreman, 1995). If we continue with the revolving door of reflexively banning
each new psychoactive drug as it appears, thereby encouraging further exploration with novel
pharmaceuticals, it is only a matter of time before the recurrence of a situation analogous to that
of MPTP. However, next time we may not be so lucky.
Two factors combine to exacerbate the danger. The rapid and worldwide spread of novel
psychoactive drugs (as seen with MMC) in advance of any substantial research into toxicity is
the first factor. The second issue is the subtle and progressive nature of many neurological
disorders, with substantial and lasting neural damage often preceding overt behavioural
symptoms and “incubation” periods varying from instantaneous to life-long. The combination of
these factors provides for the alarming possibility of a global pandemic of drug-induced
neurological injury. A hypothetical novel psychoactive drug which induced a slowly-appearing
neurotoxic impairment, when combined with market penetration of speed and depth similar to
MMC, has the potential to cause an immense amount of harm before the danger of such a drug is
even detected by science.
5.4 Conclusion
Overall, it would appear that the acute risks of MMC consumption, while real, are
nonetheless relatively mild, perhaps somewhere towards the midpoint between MDMA and
methamphetamine. However, although there is no conclusive evidence of direct MMC
neurotoxicity, several studies strongly suggests that MMC consumption may induce lasting
cognitive impairment (den Hollander et al., 2012; Freeman et al., 2012; Motbey et al., 2012). In
addition, the behavioural risks of MMC intoxication and addiction present a substantial harm in
themselves (Motbey, 2012; Van Hout and Bingham, 2012; Van Hout and Brennan, 2010; Van
Hout and Brennan, 2011a; Van Hout and Brennan, 2011b), and the physical damage to the
minority of MMC users consuming the drug by injection is extreme (Dorairaj et al., 2011; Van
Hout and Bingham, 2012).
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Looking beyond the direct impact of MMC use, the example of MMC is also useful as a
case study for the wider problem of novel psychoactive drug consumption. The increasing pace
and diversity of recreational drug markets worldwide has altered the situation to such an extent
that the already-flawed conventional approach to drug control has become actively dangerous. At
the very least, the situation calls for a considerable increase in the resources allocated to the
investigation of the toxicity of emerging drugs of abuse. However, although essential, such a
response represents at best a band-aid reaction to the new reality.
The rise of MMC was arguably a consequence of the prohibition of MDMA. An
increased focus upon suppression of MMC is most likely to shift the market towards a greater
reliance on the apparently more dangerous MDPV. Eliminating MDPV will create a market for
the next new drug, and the next, and the next. Each iteration of this process carries the risk of
another MPTP, and the iterations are accelerated by our prohibitionist responses. The novel
psychoactive market is a hydra that grows more dangerous with each head we remove.
An increased focus on toxicity research is a necessary but not sufficient step towards
addressing the underlying problem, which is rooted in the lack of concordance between the
relative risks of drug harms and varying degrees of legal restriction (Hughes and Winstock,
2012; Nutt et al., 2007; Nutt et al., 2010; Winstock and Ramsey, 2010). Even with greatly
enhanced resources, the psychopharmacology community cannot possibly keep ahead of the
ever-expanding modern drug market. Effects that take years to manifest require years to
research, and the current situation is such that hundreds of thousands of people could potentially
be severely affected before any evidence of danger came to the attention of science. Eventually,
society may have to face the difficult choice of deciding to openly permit a limited set of
tolerably-dangerous drugs that are sufficient to satisfy at least most of the apparently inflexible
demand. If we wish to avert the substantial risks associated with the emergence of novel
psychoactive drugs, scientific research must be accompanied by a substantial reform of the way
in which governments worldwide seek to control the use of these drugs.
5-15
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6. Appendix: Statements from Co-Authors
_______________________________
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Callum Hicks B.Sc PhD Candidate, Psychopharmacology Laboratory
School Of Psychology, A18 , Australia, NSW 2006 Email:[email protected]
18 March 2013
To whom it may concern,
Craig was the primary author on the original manuscript entitled “Mephedrone in Adolescent Rats:
Residual Memory Impairment and Acute but Not Lasting 5-HT Depletion” published in the journal PLoS
ONE, 7(9): e45473. doi:10.1371/journal.pone.0045473.
I was a co-author on this manuscript and helped with the behavioural tests.
Yours Sincerely,
Callum Hicks (Co-author)
PhD Candidate, Psychopharmacology Laboratory
School of Psychology
The University of Sydney
6-3
6-4
I was co-author on one manuscript appearing in Addiction Biology (vol 17, 409-422). Craig was the
primary author, and I contributed to the Fos staining, counting and data analysis. .
Signed,
March 18, 2013.
Glenn Hunt, Discipline of psychiatry, University of Sydney
6-5
6-6
To Whom It May Concern,
This written statement is to attest to the fact that Craig P. Motbey was the primary author on the papers
listed below on which I am a co-author. My contribution to Paper 1 was performing the transcardial
perfusions of the rats, the extraction of their brains, and the preparation of the brains for slicing. My
contribution to Paper 2 was statistical analysis of the results for the self-administration data and writing
the related sections of the methods and results with the primary author.
Paper 1
Mephedrone (4-methylmethcathinone,‘meow’): acute behavioural effects and distribution of Fos
expression in adolescent rats
Craig P. Motbey, Glenn E. Hunt, Michael T. Bowen, Suzanne Artiss and Iain S. McGregor.
Addiction Biology, 17: 409-422
Paper 2
High Levels of Intravenous Mephedrone (4-Methylmethcathinone) Self-Administration In Rats: Neural
Consequences and Comparison with Methamphetamine
Craig P. Motbey, Kelly Clemens, Nadine Apetz, Adam R. Winstock, John Ramsey, Kong M. Li, Naomi
Wyatt, Paul D. Callaghan, Michael Bowen, Jennifer Cornish, Iain S. McGregor
Submitted to the Journal of Psychopharmacology
Kind regards,
Michael T. Bowen
6-7
6-8
To whom it may concern,
Craig Motbey was the primary author of: Mephedrone (4-methylmethcathinone,‘meow’): acutebehavioural effects and distribution of Fos expressionin adolescent rats. Craig P. Motbey, Glenn E. Hunt, Michael T. Bowen, Suzanne Artiss and Iain S. McGregor. I contributed to the acquisition of animal data and, along with the other co-authors, reviewed the content of the paper and approved final version for publication. Suzanne Artiss [email protected] 0433 615 178
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Statement of Contribution
I, Nadine Apetz, herewith assure that Craig C. Motbey was the primary author of the article "High Levels
of Intravenous Mephedrone (4-Methylmethcathinone) Self-Administration In Rats: Neural
Consequences and Comparison with Methamphetamine".
My contribution to this project consisted of assistance in the conduction of the self-administration
experiments.
6-10
6-11
6-12
6-13
I, Dr Kong M. Li, of the Sydney University is to confirm that Craig Motbey was the primary
author and I was co-author for the publication of following manuscripts:
Mephedrone in Adolescent Rats: Residual Memory Impairment and Acute but Not Lasting 5-HT
Depletion
Craig P. Motbey, Emily Karanges, Kong M. Li, Shane Wilkinson, Adam R. Winstock, John
Ramsay, Callum Hicks, Michael D. Kendig, Naomi Wyatt, Paul D. Callaghan and Iain S.
McGregor.
PLoS ONE, 7(9): e45473. doi:10.1371/journal.pone.0045473
High Levels of Intravenous Mephedrone (4-Methylmethcathinone) Self-Administration In Rats:
Neural Consequences and Comparison with Methamphetamine
Craig P. Motbey, Kelly Clemens, Nadine Apetz, Adam R. Winstock, John Ramsey, Kong M. Li,
Naomi Wyatt, Paul D. Callaghan, Michael Bowen, Jennifer Cornish, Iain S. McGregor
Submitted to the Journal of Psychopharmacology
6-14
I, Emily Karanges, certify that Craig Motbey was the primary author on the publication entitled
“Mephedrone in Adolescent Rats: Residual Memory
Impairment and Acute but Not Lasting 5-HT Depletion” (PLoS ONE, 7(9): e45473.
doi:10.1371/journal.pone.0045473). I provided practical assistance with the running and analysis of the
HPLC data (neurotransmitter and metabolite analysis).
Emily Karanges
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I was co-author on the manuscripts featured in Chapters 3 and 4. Craig Motbey was the primary author,
and I contributed to the provision and characterization of the test compound (mephedrone).
Signed March 27. 2013. Adam Winstock Kings College London
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Iain S. McGregor MA (Oxon) PhD (Sydney)
Professor of Psychopharmacology
Tel +61 2 9351 3571 Fax +61 2 9351 8023
School Of Psychology, A18 , Australia, NSW 2006 Email: [email protected]
28 December 2013
I was the primary supervisor of Craig Motbey’s PhD thesis and the senior (last) author on the two
published papers and one submitted paper that appear in his thesis.
Craig was the first and primary author and performed most of the hands-on research and the writing of
the manuscripts.
I contributed towards the conception of the studies, experimental design, the forming of collaborative
networks, the writing of the manuscripts and the funding of the research via ARC, NHMRC and AINSE
grants.
Signed
1st April 2013
Professor Iain McGregor, School of Psychology, University of Sydney
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