in vitro characterization of psychoactive substances at...
TRANSCRIPT
1521-0103/357/1/134–144$25.00 http://dx.doi.org/10.1124/jpet.115.229765THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS J Pharmacol Exp Ther 357:134–144, April 2016Copyright ª 2016 by The American Society for Pharmacology and Experimental Therapeutics
In Vitro Characterization of Psychoactive Substances at Rat,Mouse, and Human Trace Amine-Associated Receptor 1s
Linda D. Simmler, Danièle Buchy, Sylvie Chaboz, Marius C. Hoener, and Matthias E. LiechtiDivision of Clinical Pharmacology and Toxicology, Department of Biomedicine, University Hospital Basel, University of Basel,Basel, Switzerland (L.D.S., M.E.L.); and Neuroscience Research, Pharma Research and Early Development, Roche InnovationCenter Basel, F. Hoffmann-La Roche Ltd., Basel, Switzerland (D.B., S.C., M.C.H)
Received October 5, 2015; accepted January 19, 2016
ABSTRACTTrace amine-associated receptor 1 (TAAR1) has been implicatedin the behavioral effects of amphetamine-type stimulant drugs inrodents. TAAR1 has also been suggested as a target for novelmedications to treat psychostimulant addiction. We previouslyreported that binding affinities at TAAR1 can differ betweenstructural analogs of psychostimulants, and species differenceshave been observed. In this study, we complement our previousfindings with additional substances and the determination offunctional activation potencies. In summary, we present herepharmacological in vitro profiles of 101 psychoactive substancesat human, rat, andmouseTAAR1.p-Tyramine,b-phenylethylamine,and tryptamine were included as endogenous comparatorcompounds. Functional cAMP measurements and radioliganddisplacement assays were conducted with human embryonickidney 293 cells that expressed human, rat, or mouse TAAR1.
Most amphetamines, phenethylamine, and aminoindanesexhibited potentially physiologically relevant rat and mouseTAAR1 activation (EC50 , 5 mM) and showed full or partial(Emax , 80%) agonist properties. Cathinone derivatives,including mephedrone and methylenedioxypyrovalerone,exhibited weak (EC50 5 5–10 mM) to negligible (EC50 . 10 mM)binding properties at TAAR1. Pipradrols, including methylphe-nidate, exhibited no affinity for TAAR1. We found considerablespecies differences in activity at TAAR1 among the highly activeligands, with a rank order of rat . mouse . human. Thischaracterization provides information about the pharmacologi-cal profile of psychoactive substances. The species differencesemphasize the relevance of clinical studies to translationallycomplement rodent studies on the role of TAAR1 activity forpsychoactive substances.
IntroductionTrace amine-associated receptor 1 (TAAR1) is a relatively
recently discovered G protein–coupled receptor (Borowskyet al., 2001; Bunzow et al., 2001), which is expressed inmonoaminergic brain regions and throughout the limbicsystem (Borowsky et al., 2001; Lindemann et al., 2008;Espinoza et al., 2015). TAAR1 is thought to play a role inregulating the limbic network, reward circuits, cognitiveprocesses, and mood states and has been proposed as apharmacological target for the treatment of mental disorders(Wolinsky et al., 2007; Lindemann et al., 2008; Miller, 2011;Revel et al., 2013) and psychostimulant dependence (Di Caraet al., 2011; Pei et al., 2014; Cotter et al., 2015; Jing and Li,2015). TAAR1 is stimulated by endogenous ligands, includingb-phenylethylamine (b-PEA), p-tyramine, tryptamine, and3-iodothyronamine (Scanlan et al., 2004; Zucchi et al., 2006).Many psychoactive compounds, including amphetamine and
phenethylamine derivatives, also bind to TAAR1 (Bunzowet al., 2001; Wainscott et al., 2007; Simmler et al., 2013; Reeseet al., 2014). The activation of TAAR1 results in elevations inintracellular cAMP (Bunzow et al., 2001; Xie and Miller,2007).Amphetamines have structural similarity to the endoge-
nous ligand b-PEA and were initially identified as TAAR1ligands (Bunzow et al., 2001). We previously reported thatmany novel psychoactive substances are also ligands of ratandmouse TAAR1 (Simmler et al., 2013, 2014a,b; Rickli et al.,2015a,b,c). However, several novel psychoactive substances donot bind to TAAR1, and little has been reported on theactivation of human TAAR1. The pharmacological and toxi-cological actions of novel psychoactive substances are also ofinterest because of the emergence of hundreds of thesesubstances, referred to as “legal highs” or “research chem-icals.” These chemical compounds are recreationally used buthave poorly known pharmacological properties.TAAR1 is implicated in the control of neuronal firing
frequency and is thus likely to contribute to psychoactiveand abuse-related drug effects. Ex vivo electrophysiologyexperiments that used slices from TAAR1 knockout (KO)mice (Lindemann et al., 2008) or pharmacological TAAR1blockade (Bradaia et al., 2009) suggest that TAAR1 is
This research was supported by the Federal Office of Public Health [Grant13.006497] and F. Hoffmann-La Roche Ltd. and the University of Basel[Translational Medicine Hub Innovation Fund].
dx.doi.org/10.1124/jpet.115.229765.s This article has supplemental material available at jpet.aspetjournals.org.
ABBREVIATIONS: b-PEA, b-phenylethylamine; DA, dopamine; 5-HT, 5-hydroxytryptamine (serotonin); KO, knockout; MDMA,3,4-methylenedioxymethamphetamine; PBS, phosphate-buffered saline; RO5166017, (S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine; RO5203648, (S)-4-(3,4-dichlorophenyl)-4,5-dihydrooxazol-2-amine dihydrochloride; TAAR1, trace amine-associated receptor 1; WT, wild type.
134
http://jpet.aspetjournals.org/content/suppl/2016/01/20/jpet.115.229765.DC1Supplemental material to this article can be found at:
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
constitutively active to control dopamine (DA) and serotonin[5-hydroxytryptamine (5-HT)] tone.Compared with wild-type (WT) mice, TAAR1 KO mice
were shown to consume more ethanol and be more suscepti-ble to its sedating effects (Lynch et al., 2013). The TAAR1partial agonist RO5203648 [(S)-4-(3,4-dichlorophenyl)-4,5-dihydrooxazol-2-amine dihydrochloride] reduced cocaine self-administration and cocaine-induced hyperlocomotion in rats(Revel et al., 2012b). Both selective TAAR1 partial agonistsand selective TAAR1 full agonists reduced cocaine self-administration and the reinstatement of drug-seeking behaviorin rats (Pei et al., 2014, 2015) and decreased cocaine-mediatedintracranial self-stimulation (Pei et al., 2015). Reductions ofhyperlocomotion, self-administration, and reinstatement byTAAR1 partial agonism have also been reported for metham-phetamine (Cotter et al., 2015; Jing andLi, 2015). These studiesestablished TAAR1 as a promising target for therapeutics totreat substance use disorders, regardless of the TAAR1binding properties of the abused substances themselves. Bydirectly interacting with TAAR1, psychoactive substancesmay also modulate their own pharmacological effects. Forexample, amphetamine induces markedly more striatalmonoamine release in TAAR1 KO mice than in WT mice(Lindemann et al., 2008). Methamphetamine and amphet-amine increase locomotor activity to a greater extent inTAAR1 KO mice compared with WT mice (Achat-Mendeset al., 2012). TAAR1 also plays a role in contingent oralmethamphetamine intake (Harkness et al., 2015). Similar toamphetamine and methamphetamine, 3,4-methylenedioxy-methamphetamine (MDMA) significantly increased extra-cellular striatal DA and 5-HT levels to a greater extent inTAAR1 KO mice compared with WT mice (Di Cara et al.,2011). TAAR1 KO mice are hypersensitive to psychoactivesubstances that are also TAAR1 ligands. By contrast, TAAR1overexpression in mice reduced locomotor activity inresponse to amphetamine (Revel et al., 2012a). BecauseMDMA, methamphetamine, and amphetamine are TAAR1ligands, they possibly autoinhibit their own effects on neuro-transmitter release. Di Cara et al. (2011) supported theconcept of the autoregulation of TAAR1-activating psychostim-ulants, showing that the TAAR1 ligand o-phenyl-3-iodotyraminedecreased the DA release response to p-chloroamphetamine,which is a psychostimulant that is inactive at TAAR1, in WTmice but not in TAAR KO mice.Because TAAR1might be significantly involved in the mode
of action of many psychoactive drugs, we determined theTAAR1 binding and activation properties of a series of mostlynovel substances and found considerable differences inTAAR1 binding properties within and between substanceclasses. Our data set provides evidence of significant speciesdifferences in ligand/receptor interactions between rodent andhuman TAAR1.
Materials and MethodsChemicals. The compounds were purchased from Lipomed
(Arlesheim, Switzerland) or Cayman Chemicals (Ann Arbor, MI)as racemic mixtures, with the exception of D-amphetamine,D-methamphetamine, (1)-ephedrine, and (2)-ephedrine. A list ofgeneric or full chemical names is provided in (Supplemental Table 1).5-EAPB, diclophensine, diphenidine, ethylphenidate,methoxphenidine,and N-methyl-2-AI were obtained from the Forensic Institute Zürich
(Zürich, Switzerland). Naphyrone and MDAI were synthesized inour laboratory as reported previously (Simmler et al., 2013, 2014b).Radiochemicals (3H-isotopes) were purchased from PerkinElmer(Schwerzenbach, Switzerland), with the exception of [3H]RO5166017[(S)-4-[(ethyl-phenyl-amino)-methyl]-4,5-dihydro-oxazol-2-ylamine],which was synthesized at Roche (Basel, Switzerland).
Cell Culture and Membrane Preparation. Human embryonickidney 293 cells that stably expressed human, rat, or mouse TAAR1were used as described previously (Revel et al., 2011). All of the celllines were maintained at 37°C and 5% CO2 in high-glucose Dulbecco’smodified Eagle’s medium that contained 10% fetal calf serum (heat-inactivated for 30 minutes at 56°C), 1% penicillin/streptomycin, and375 mg/ml Geneticin (Gibco, Zug, Switzerland). For membranepreparation, the cells were released from culture flasks usingtrypsin/EDTA, harvested, washed twice with ice-cold phosphate-buffered saline (PBS; without Ca21 andMg21), pelleted at 1000� g for5 minutes at 4°C, frozen, and stored at 280°C. Frozen pellets weresuspended in buffer A [20 ml HEPES-NaOH (20 mM, pH 7.4) thatcontained 10 mM EDTA] and homogenized with a Polytron (PT 6000;Kinematica, Luzern, Switzerland) at 14,000 rpm for 20 seconds. Thehomogenate was centrifuged for 30 minutes at 48,000 � g at 4°C. Thesupernatant was removed and discarded, and the pellet was resus-pended in buffer A using the Polytron (20 seconds at 14,000 rpm). Thecentrifugation and removal of the supernatant was repeated, andthe final pellet was resuspended in buffer A and homogenized usingthe Polytron. Typically, 2-ml aliquots of membrane portions werestored at 280°C. With each new membrane batch, the dissociationconstant (Kd) was determined by a saturation curve.
Radioligand Binding Assay. For the competitive binding assays,the TAAR1 agonist [3H]RO5166017 was used as a TAAR1 radioligandat a concentration equal toKd values, which was usually around 0.7 nM(mouse TAAR1) and 2.3 nM (rat TAAR1). Nonspecific binding wasdefined as the amount of radioligand bound in the presence of 10 mMRO5166017. Compoundswere tested at a broad range of concentrations(10 pM to 10mM) in duplicate. Compounds (20ml/well) were transferredto a 96-deep-well plate (TreffLab, Degersheim, Switzerland), and 180mlbinding buffer (20mMHEPES-NaOH, 10mMMgCl2, and 2mMCaCl2,pH 7.4), 300 ml radioligand, and 500 ml membranes (resuspended at60 mg protein/ml) were added. The plates were incubated at 4°C for90 minutes. Incubations were terminated by rapid filtration throughUnifilter-96 plates (Packard Instrument Company, PerkinElmer) andglass filtersGF/C (PerkinElmer) presoaked for 1 hour in polyethylenimine(0.3%) and washed three times with 1 ml cold binding buffer. After theaddition of 45 ml Microscint 40 (PerkinElmer), the Unifilter-96 platewas sealed. After 1 hour, radioactivity was counted using a TopCountMicroplate Scintillation Counter (Packard Instrument Company).IC50 values were determined by calculating nonlinear regressioncurves for a one-site model using at least three independent 10-pointconcentration-response curves, run in duplicate, for each compound.Ki (affinity) values, which correspond to the dissociation constants,were determined using the Cheng–Prusoff equation. Ki values arepresented as means 6 S.D. (in micromoles). For reasons of integrity,Table 1 includes severalKi values that we have previously publishedas indicated by the references in the table.
Functional TAAR1 Activity. Substances were tested for bindingaffinity at rat and mouse TAAR1 as described above. If relevantbinding was observed, then we also determined potencies for receptoractivation and maximal efficacy at rat, mouse, and human TAAR1 tocharacterize the compounds as full or partial agonists (Emax , 80%).The endogenous TAAR1 ligands b-PEA, p-tyramine, and tryptamineserved as reference substances for comparisons of affinity values andfunctional potency and efficacy. cAMPmeasurements were performedas described previously (Revel et al., 2011). In brief, cells that expressedrat or mouse TAAR1 were plated on 96-well plates (BIOCOAT 6640;Becton Dickinson, Allschwil, Switzerland) and incubated for 20 hoursat 37°C. Prior to stimulation of the cells with a broad concentra-tion range of agonists for 30 minutes at 37°C, the cells werewashed with PBS and preincubated with PBS that contained 1 mM
Psychoactive Substances and TAAR1 135
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
TABLE
1Bindingaffinities,activa
tion
potenc
ies,
andefficacy
ofps
ychoa
ctivesu
bstanc
esan
den
doge
nous
compa
ratorcompo
unds
atrat,mou
se,a
ndhu
man
TAAR1
Value
saregivenas
mea
ns6
S.D.K
iva
lues
ofhu
man
TAAR1wereno
tde
term
ined
becaus
eof
thelack
ofarelia
bleradioligan
dne
eded
forthein
vitroassay.
EC50
was
determ
ined
forsu
bstanc
eswithreleva
ntbind
ing(K
iva
lue,
10mM).
Kiva
lues
from
ourpr
eviouspu
blicationsareinclude
dan
dreferenced.
Thege
neric
orfullch
emical
nam
esforab
brev
iatedsu
bstancesarelisted
inSupp
lemen
talTab
le1.
Subs
tance
Rat
TAAR1
Mou
seTAAR1
Human
TAAR1
Recep
tor
BindingKi
Activation
Poten
cyEC50
Emax
Recep
tor
BindingKi
ActivationPoten
cyEC50
Emax
Activation
Poten
cyEC50
Emax
mM
%mM
%mM
%
Endo
genou
sliga
nds
b-PEA
0.24
60.12
0.11
60.08
1006
110.31
60.15
0.20
60.09
1026
80.26
60.09
1046
10p-Tyram
ine
0.05
96
0.02
00.03
60.02
194
611
0.38
60.13
0.28
60.17
886
90.99
60.29
916
8Tryptam
ine
0.13
60.05
0.41
60.15
916
81.46
0.4
2.76
0.5
1176
221
613
736
16Phen
ethylam
ines
25B-N
B2O
Me
0.28
60.00
2a1.26
0.4
376
194.56
1.7a
6.16
2.5
476
4.10
25C-N
B2O
Me
0.52
60.10
a1.66
0.4
296
515
62a
6.76
1.6
486
6.30
25D-N
B2O
Me
0.81
60.10
a1.56
0.4
346
413
64a
4.06
0.6
676
12.30
25E-N
B2O
Me
0.26
60.03
a0.65
60.39
376
151.16
0.3a
1.86
0.3
466
10.10
25H-N
B2O
Me
1.56
0.2a
3.06
1.4
376
11.20
a6.16
2.6
536
10.10
25I-NB2O
Me
0.44
60.07
a1.86
1.0
326
173.46
0.9a
5.26
2.2
176
9.10
25N-N
B2O
Me
2.26
0.14
a1.56
0.3
346
9.20
a.30
.10
25P-N
B2O
Me
0.05
56
0.00
4a0.51
60.20
346
170.24
60.03
a1.36
0.2
406
26.10
25T2-NB2O
Me
0.35
60.02
a0.93
60.38
246
144.26
0.6a
2.96
0.6
306
23.10
25T4-NB2O
Me
0.12
60.02
a1.16
0.6
316
121.56
0.4a
4.76
2.1
336
12.10
25T7-NB2O
Me
0.08
86
0.03
2a0.55
60.17
526
231.06
0.2a
2.16
0.5
686
23.10
2C-B
0.07
96
0.00
8a0.24
60.16
576
162.26
0.3a
2.36
0.4
696
133.36
0.9
106
22C
-B-Fly
0.02
96
0.00
8b0.27
60.16
486
10.71
60.23
b1.86
0.7
496
6.30
2C-C
0.11
60.02
a0.34
60.15
516
144.16
0.3a
2.36
1.5
576
23.10
2C-D
0.15
60.03
a0.49
60.14
556
103.56
0.1a
2.06
0.2
616
19.10
2C-E
0.06
66
0.00
9a0.18
60.14
726
131.26
0.1a
1.16
0.2
646
23.10
2C-H
0.96
0.16
a1.56
0.7
806
711
62a
7.56
3.3
566
146.56
0.7
536
52C
-I0.12
60.02
a0.19
60.11
506
193.36
0.1a
2.46
0.8
516
21.10
2C-N
0.34
60.02
a0.25
60.12
596
16.20
a15
612
286
13.10
2C-P
0.02
06
0.00
5a0.03
06
0.02
284
68
0.28
60.03
a0.56
60.23
916
274.26
0.5
726
112C
-T2
0.04
36
0.00
6a0.09
66
0.05
186
617
2.26
0.6a
4.36
2.8
546
14.10
2C-T4
0.05
36
0.00
8a0.08
36
0.05
067
611
4.56
0.9a
3.76
2.2
516
21.10
2C-T7
0.03
36
0.00
5a0.07
96
0.03
483
611
0.56
60.12
a0.91
60.67
676
4.10
Mescaline
3.36
0.5a
3.76
1.8
376
1811
64a
4.86
3.7
256
20.10
Mescaline-NB2O
Me
136
6a.30
.20
a.30
.10
Amph
etam
ines
Amph
etam
ine
0.23
60.18
c0.66
60.61
916
200.08
96
0.05
9c0.53
60.67
906
302.86
0.8
916
154-APB
0.11
60.02
b0.16
60.09
756
92.16
0.1b
0.85
60.78
726
124.16
2.1
506
215-APB
0.04
26
0.00
6b0.06
76
0.04
088
612
0.11
60.00
2b0.13
60.07
676
116.16
2.3
436
166-APB
0.05
26
0.01
7b0.04
26
0.02
990
614
0.05
66
0.01
5b0.06
76
0.03
993
613
7.26
0.3
476
77-APB
0.06
66
0.00
6b0.05
86
0.03
310
96
130.13
60.02
b0.11
60.07
956
170.63
60.13
896
45-APDB
0.49
60.05
b1.46
0.7
936
200.77
60.06
b1.56
1.2
646
14.10
6-APDB
1.06
0.04
b1.06
0.97
836
170.21
60.04
b0.51
60.27
956
19.10
5-EAPB
0.81
60.08
b1.16
0.6
396
10.15
b13
67
266
11.10
N-E
thylam
phetam
ine
2.56
1.4d
0.88
60.05
626
10.10
d.10
.10
4-Fluoroa
mph
etam
ine
0.08
16
0.04
1e0.06
96
0.00
478
612
0.32
60.1e
0.13
60.02
776
123.56
0.6
676
94-Fluorom
etham
phetam
ine
0.24
60.09
e0.16
60.02
766
111.76
0.9e
0.46
60.05
696
66.26
2.2
446
115-IT
0.15
60.02
0.20
60.01
666
40.36
60.15
0.34
60.07
636
5.30
5-MAPDB
0.67
60.09
b1.16
0.9
656
113.56
0.1b
4.36
3.0
596
11.10
MBDB
1.26
0.1c
1.76
1.2
756
83.66
1.1c
4.16
1.1
346
28.30
MDA
0.25
60.04
b0.74
60.16
866
50.16
60.01
b0.58
60.08
1026
113.66
0.4
116
4MDEA
2.76
1.0c
1.56
0.9
666
246.66
3.1c
6.26
3.4
356
33.30
MDMA
0.37
60.12
c1.06
0.7
566
102.46
1.1c
4.06
1.0
716
1635
621
266
8
(con
tinu
ed)
136 Simmler et al.
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
TABLE
1—Con
tinued
Subs
tance
Rat
TAAR1
Mou
seTAAR1
Human
TAAR1
Recep
tor
BindingKi
Activation
Poten
cyEC50
Emax
Recep
tor
BindingKi
ActivationPoten
cyEC50
Emax
Activation
Poten
cyEC50
Emax
Metha
mph
etam
ine
0.35
60.12
c0.85
60.38
736
100.55
60.24
c0.73
60.47
786
75.36
2.3
706
214-Methy
lamph
etam
ine
0.10
60.01
0.11
60.02
936
170.15
60.07
0.07
16
0.01
394
68
.30
4-MTA
0.28
60.04
d0.26
60.03
566
100.04
36
0.00
7d0.08
96
0.01
679
619
.10
PMA
0.66
60.02
d0.34
60.13
836
80.14
60.10
d0.24
60.14
916
5.30
PMMA
1.36
0.2d
0.63
60.29
756
150.26
60.11
d1.06
0.1
826
13.30
Cathinon
es4-Bromom
ethcathinon
e1.86
0.1e
5.26
2.5
286
1213
63e
156
750
612
.30
Buph
edrone
.10
d.10
d
Bupr
opion
.20
.20
Butylone
.20
c.20
c
Cathino
ne
2.26
0.7c
1.26
0.3
286
82.16
0.7c
1.26
0.3
666
316.96
3.2
536
16N,N
-Dim
ethy
lcathino
ne
.10
d.10
d
Ethcathinon
e.10
d.10
d
4-Ethylmethcathinon
e.20
e.20
e
Ethylon
e.20
c.20
c
Fleph
edrone
5.46
1.7c
156
346
68
.10
c.20
.30
3-Fluorom
ethcathinon
e.10
d.10
d
MDPBP
.20
e.50
e
MDPPP
166
7e.20
e
MDPV
7.26
1.1c
5.96
2.7
626
14.10
c.30
.30
Mep
hedr
one
4.36
2c9.06
4.3
526
3.10
c20
67
876
16.30
Methcathinon
e4.16
1.2c
8.26
2.5
416
10.10
c6.86
2.7
646
16.30
Methe
dron
e18
64d
.20
d
Methy
lened
ioxy
cath
inon
e4.86
0.9b
5.76
0.9
536
206.56
2.8b
7.86
2.4
576
2.30
3-Methy
lmethc
athinon
e5.76
1.4
.10
116
13.86
0.04
256
7.10
4-Methy
lethcathinon
e.20
d.20
d
Methylon
e.13
c.10
c
Nap
hyron
e.20
c.20
c
Pen
tedr
one
.10
d.10
d
Pen
tylone
.10
d.10
d
a-PVP
166
6e.20
e
Pyrov
aleron
e.13
c.10
c
Eph
edrine
s(2
)-Eph
edrine
3.76
0.9
2.56
0.7
426
5.15
146
231
67
.10
(+)-Eph
edrine
5.26
1.7
106
629
69
.15
196
1021
616
.10
4-Fluoroeph
edrine
2.66
1.2e
2.26
0.8
406
1818
68e
236
810
06
10.30
Tryptam
ines
5-MeO
-aMT
1.16
0.2
2.36
0.2
386
64.86
0.9
3.76
1.6
556
5.10
4-HO-D
iPT
.15
.15
4-HO-M
ET
3.16
0.2
2.16
0.3
716
912
63
2.56
1.1
786
4.10
5-MeO
-MiPT
.15
.15
DiPT
.15
.15
N,N
-DMT
2.26
0.2
1.56
0.1
816
153.36
0.4
1.26
0.2
736
1.10
Psilocin
1.46
0.2
0.92
60.58
856
717
62
2.76
2.5
806
9.30
Aminoinda
nes
2-AI
0.31
60.09
5f0.11
60.04
906
52.16
0.4f
0.33
60.06
546
151.56
0.1
1106
5N-M
ethy
l-2-AI
0.53
60.04
0.37
60.21
636
52.66
0.1
0.94
60.09
1086
143.36
0.2
546
85-IA
I0.03
06
0.00
7f0.03
36
0.01
396
624
1.16
0.4f
0.41
60.00
236
610
3.26
0.8
336
5MDAI
0.57
60.19
f0.22
60.13
956
151.86
0.1f
0.52
60.24
996
144.16
0.5
306
7Piperaz
ines
Ben
zylpiperaz
ine
.20
f.20
f
m-C
PP
0.05
46
0.01
0f0.15
60.11
606
136.66
1.1f
3.26
1.2
406
20.30
(con
tinu
ed)
Psychoactive Substances and TAAR1 137
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
3-isobutyl-1-methylxanthine for 10 minutes at 37°C and 5% CO2.Stimulation with 0.2% dimethylsulfoxide was set as the basal level,and the effect of 30 mM b-PEA was set as the maximal response.Subsequently, the cells were lysed, and cAMP assays were performedaccording to the manufacturer’s instructions (cAMP kit; Upstate/Millipore, Schaffhausen, Switzerland). Finally, the plates were readwith a luminometer (1420 Multilabel Counter; PerkinElmer), and theamount of cAMP was calculated. The results were obtained from atleast three independent experiments. Experiments were run induplicate or triplicate. EC50 values are presented as means 6 S.D.(in micromoles). The Emax value for the functional activity data atTAAR1 describes the degree of functional activity compared with100% for the endogenous ligand and full agonist b-PEA.
ResultsThe binding affinity values (Ki), receptor activation poten-
cies (EC50), and maximal efficacy (Emax) of 104 substances aresummarized in Table 1. These substances represent thecollection of compounds in our laboratory that have been usedto characterize the in vitro pharmacology of novel designerdrugs (for review, see Liechti, 2015). The substances for thesestudies were chosen based on the availability of pure chemicalcompounds and the reported abuse of these substances. For asubset of compounds, we have previously published rat andmouse TAAR1 affinities as indicated by references in Table 1,but no human, rat, and mouse activity data. All substanceswere grouped according to their basic chemical structure(Fig. 1) as phenethylamines, amphetamines, cathinones,ephedrines, tryptamines, aminoindanes, pipradrols, andpiperazines. A few psychoactive substances, such as cocaineand lysergic acid diethylamide, were added but not classi-fied because of the lack of common basic structures.TAAR1 Binding and Functional TAAR1 Activation.
We found marked differences in affinities at TAAR1 andthe functional activation of TAAR1 among the variouspsychoactive substances. The ligand properties varied con-siderably within substance classes, with the exception ofcathinones and pipradrols, which generally exhibited noneto very weak binding to TAAR1. We also observed speciesdifferences in TAAR1 activation. At the human TAAR1, only19 substances had EC50 values that indicated functionalactivation (,10 mM), whereas the EC50 values were , 10 mMfor 52 and 68 substances at the mouse and rat TAAR1,respectively. Therefore, below we present the properties ofthe psychoactive compounds compared with the endogenousTAAR1 ligands b-PEA, p-tyramine, and tryptamine sepa-rately for each species.Human TAAR1. b-PEA and p-tyramine activated the
human TAAR1 with EC50 values of 0.26 and 0.99 mM,respectively, and showed full agonistic properties (Emax 5104% and 91%, respectively). Amphetamine, 7-APB, and 2-AIhad potency (EC50 5 0.6–2.8 mM) and agonist efficacy (Emax .89%) that were comparable to b-PEA and p-tyramine athuman TAAR1. Methamphetamine exhibited an EC50 of5.3 mM and 70% efficacy. The endogenous ligand tryptamineexhibited weak activation of human TAAR1, with an EC50 of21 mM (Emax 5 73%). Generally, most of the psychoactivecompounds that were tested were weak human TAAR1ligands, and none of them were more potent than theendogenous ligand b-PEA.Rat TAAR1. At the rat TAAR1, b-PEA had a Ki of 0.24 mM
and showed full agonism, with an EC50 of 0.11 mM, whereasTABLE
1—Con
tinued
Subs
tance
Rat
TAAR1
Mou
seTAAR1
Human
TAAR1
Recep
tor
BindingKi
Activation
Poten
cyEC50
Emax
Recep
tor
BindingKi
ActivationPoten
cyEC50
Emax
Activation
Poten
cyEC50
Emax
TFMPP
0.38
60.06
f0.75
60.18
596
162.36
0.6f
3.86
0.2
446
4.30
Pipradr
ols
2-DPMP
.10
f.10
f
D2P
M.10
f.10
f
Ethylph
enidate
.15
.15
Methox
phen
idine
.15
.15
Methylph
enidate
.15
f15
610
586
9.30
f29
63
446
15.30
Others
Cocaine
.10
c.10
c
Diclofensine
1.36
0.1
116
0.1
306
16.96
0.7
.30
.10
Diphen
idine
.15
.15
LSD
0.45
60.05
a1.46
0.4
296
810
63a
9.76
3.5
136
4.20
Methylhex
anam
ine
.15
.15
Mod
afinil
.15
.30
.15
.30
.30
aKiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inRickliet
al.(20
15c).
bKiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inRickliet
al.(20
15b).
c Kiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inSim
mleret
al.(201
3).
dKiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inSim
mleret
al.(201
4a).
e Kiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inRickliet
al.(201
5a).
f Kiva
lues
forratan
dmou
seTAAR1pr
eviouslypu
blished
inSim
mleret
al.(201
4b).
138 Simmler et al.
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
p-tyramine was more potent, with a Ki of 0.06 mM and EC50 of0.03 mM, with full agonist properties (Emax 5 94%). The thirdendogenous ligand tested, tryptamine, showed similar affinity(Ki 5 0.13 mM) to b-PEA and p-tyramine and slightly lowerfunctional activity (EC50 5 0.41 mM). Several of the screenedamphetamines (5-APB, 6-APB, 7-APB, and4-fluoroamphetamine)and phenethylamines (2C-P, 2C-T2, 2C-T4, and 2C-T7) andthe aminoindane 5-IAI had affinities and EC50 values thatwere comparable to p-tyramine, the most potent of the threeendogenous ligands at rat TAAR1, but none of these had amore potent EC50 than p-tyramine. The majority of thesepotent novel psychoactive substances exhibited full agonistproperties (Emax . 80%) at rat TAAR1. The Emax values of4-fluoroamphetamine and 2C-T4 were 78% and 67%, suggestingpartial agonism.The piperazine m-CPP and several amphetamines (4-APB,
4-fluoromethamphetamine, 5-IT, 4-methylamphetamine, and4-MTA), phenethylamines (2C-B, 2C-B-Fly, 2C-C, 2C-E, 2C-I,and 2C-N), and aminoindanes (2-AI, N-methyl-2-AI, andMDAI) were comparable to the less potent endogenous ligandsb-PEA and tryptamine in their affinities and functionalpotencies at rat TAAR1. The majority of these compoundswere partial agonists. Amphetamine and the well knownamphetamine derivatives methamphetamine, MDMA, andMDA were slightly less active than the structurally relatedendogenous ligand b-PEA.Mouse TAAR1. b-PEA and p-tyramine had equal affini-
ties at mouse TAAR1, with Ki values of 0.31 and 0.38 mM,respectively, and full agonist properties, with EC50 values of0.2 and 0.28 mM, respectively. Various amphetamines (am-phetamine, 6-APDB, 4-fluoroamphetamine, 5-IT, MDA, andPMA) and one phenethylamine (2C-P) showed similar bindingaffinities and functional potencies to these endogenousTAAR1 ligands, whereas some amphetamines were evenmorepotent (5-APB, 6-APB, 7-APB, 4-methylamphetamine, and 4-MTA), with mostly full agonist properties. The endogenousligand tryptamine was slightly weaker than b-PEA and
p-tyramine, and many phenethylamines (25E-NB2OMe,25P-NB2OMe, 25T7-NB2OMe, 2C-E, and 2C-T7), amphet-amines (4-APB, 4-fluoroamphetamine, 5-IT, methamphet-amine, and PMMA), and aminoindanes (2-AI, N-methyl-2-AI,and MDAI) were similarly active as b-PEA and p-tyramine,although all of them were partial agonists (Emax 5 36%–78%),with the exception of PMMA and MDAI (Emax 5 82% and99%, respectively). Interestingly, binding affinity did notalways predict functional potency, such as with the amino-indanes, which showed functional activities similar to am-phetamine but exhibited much lower binding affinities thanamphetamine.Differences in Activation Potencies at Human versus
Rat and Mouse TAAR1. Our results suggest significantspecies differences in TAAR1 affinities and activationpotencies for most of the substances with relevant bindingproperties in the rat. Importantly, although the endogenousligands p-tyramine and tryptamine activated TAAR1 with apotency rank order of rat . mouse . human, like manypsychostimulant compounds, b-PEA had similar EC50
values across the three species. This is relevant because acomparison of activation potencies across species with invitro assays could be biased by assay-specific variables, suchas expression levels of the transporters in the respective celllines. However, b-PEA can serve as a reference compoundfor species comparisons. Wainscott et al. (2007) also report-ed comparable EC50 values for b-PEA at rat and humanTAAR1 and species differences for other compounds. Toquantify the extent of species differences in TAAR1 activa-tion potencies, we calculated human/rat EC50 ratios andhuman/mouse EC50 ratios for substances with low tosubmicromolar (,10 mM) potencies for human TAAR1activation. The human/rat ratios ranged from 171 to 2.4,demonstrating the lower activity of the compounds at hu-man versus rat TAAR1 (Table 2). This broad range of ratiosshowed that the extent of species differences varied sub-stantially between compounds. The endogenous ligand
Fig. 1. Chemical structures of basic compounds (in bold) and location for derivatization (indicated by “R”) for compounds included in the data set.Residues for each compound are specified in Fig. 2 and Supplemental Table 1. Structural analogs were grouped according to the basic chemicalstructures for presentation of the data in tables and heat maps (Fig. 2). The generic or full chemical names for abbreviated substances are listed inSupplemental Table 1.
Psychoactive Substances and TAAR1 139
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
tryptamine exhibited a high human/rat ratio (51). Metham-phetamine and amphetamine presented relatively lowhuman/rat ratios of 6.2 and 4.2, respectively, whereas thehuman/rat ratio for MDMA was significantly higher (35). Thehuman/mouse ratios were lower than their respective human/rat ratios for all substances, with human/rat ratios $ 8.9.Calculations of ratios for substances that were inactive athuman TAAR1 (EC50 . 10 mM) were not meaningful, butsubstantial differences between human/rat and human/mouseratios were observed among the substances that were active atrat and mouse TAAR1.Species differences and differences across substances in
TAAR1 activation potencies are presented as a heat map inFig. 2, in which the substances were sorted according totheir EC50 values for the activation of rat TAAR1. Clearly,there was an overall rank order of rat . mouse . humanacross the psychoactive substances. Figure 2 also shows thatTAAR1 binding and activation was greater for certain sub-stance classes than for others. Amphetamines, cathinones,and phenethylamines represented the three largest substanceclasses in our screen. Amphetamines and phenethylamineswere well represented among the potent TAAR1 ligands,with EC50 values within a range that could be physiologi-cally relevant (30 nM to 5 mM), whereas the activity ofcathinone derivatives was low (EC50 . 5 mM, except forcathinone). None of the pipradrols exhibited significantbinding properties. Interestingly, tryptamine derivativeswere weak agonists or did not bind to TAAR1 at all, althoughtryptamine itself is an endogenous rat and mouse TAAR1ligand, with activation potency that is comparable to b-PEAand full efficacy.
DiscussionFor this in vitro study, we determined the binding affinities
and activation potencies of a large set of psychoactivecompounds at the human, rat, and mouse TAAR1 in heterol-ogous expression systems. We also characterized the ligandsas full or partial agonists. None of the active compounds hadfull antagonist properties. As indicated by our previousstudies (Simmler et al., 2013, 2014a), cathinone derivativesstood out as poor TAAR1 ligands. Most of the other psychoac-tive compounds were potent to moderate rat and mouseTAAR1 agonists but exhibited generally weak or no activityat human TAAR1. The active compounds showed full orpartial TAAR1 agonist properties, with generally no distinctpatterns related to their chemical structure.To our knowledge, our screening is the most extensive
published data set to date, which included 101 psychoactivesubstances and 3 endogenous ligands as comparator com-pounds. The in vitro pharmacology of comparator com-pounds and some psychoactive substances (amphetamine,methamphetamine, MDMA) were previously determined byus and others (Borowsky et al., 2001; Bunzow et al., 2001;Reese et al., 2007; Wainscott et al., 2007; Lindemann et al.,2008; Lewin et al., 2011). The replication of those datafor this study was an important validation of our assays.Furthermore, because we determined binding affinities forsome novel psychoactive substances in earlier studies, weincluded these data in this study. As a result, all datadetermined by our laboratory are conveniently summarizedhere in Table 1.Species differences in TAAR1 activation between rodent
and human receptors have been reported previously forphenethylamine analogs (Wainscott et al., 2007), p-tyramine,and methamphetamine (Reese et al., 2007). These compoundshave consistently exhibited lower potencies for human TAAR1activation than for rodent TAAR1 activation. Structure-activity correlations for b-PEA derivatives with regard tohuman TAAR1 activation have shown that bulky residues onthe amine or phenyl ring reduced ligand potency (Lewin et al.,2008). Reduced human TAAR1 activity could be expected fornovel psychoactive substances for which substantial derivati-zation is typical. Together with previous reports on speciesdifferences, the data provide evidence that many psychoactivesubstances are considerably less potent at human TAAR1than at mouse or rat TAAR1. In rodents, psychoactivecompounds could reduce neuronal firing via TAAR1 activation,comparable to the endogenous TAAR1 ligand p-tyramine,which has been shown to reduce the DA neuron firingrate (Bradaia et al., 2009). Consequently, the psychoactiveTAAR1 ligands likely exert autoregulatory effects on theirTAAR1-independent effects, such as reducing drug-inducedDA release in the striatum (Di Cara et al., 2011). Given thatstudies in rodents have reported autoregulatory effects of thepsychostimulant TAAR1 ligands amphetamine, methamphet-amine, and MDMA (Lindemann et al., 2008; Di Cara et al.,2011; Achat-Mendes et al., 2012; Harkness et al., 2015), thesespecies differences at TAAR1 could be relevant to the trans-lational validity of preclinical studies. Particularly for novelpsychoactive substances with large TAAR1 species differ-ences, the abuse liability that is evaluated in rodent modelsmay actually underestimate the risk for addiction that isposed by the drugs in humans.
TABLE 2EC50 ratios calculated for all substances with EC50 values , 10 mM forthe human TAAR1Ratios are ranked according to the human/rat ratio. Values . 1 indicate lowerpotency at the human TAAR1 versus the rat or mouse TAAR1.
Substance
EC50 Ratio
Structure ClassHuman/Rata
Human/Mouse
6-APB 171 107 Amphetamines2C-P 140 7.5 Phenethylamines5-IAI 97 7.8 Aminoindanes5-APB 91 47 AmphetaminesTryptamine 51 7.8 Endogenous
ligandsb
4-Fluoroamphetamine 51 27 Amphetamines4-Fluoromethamphetamine 39 13 AmphetaminesMDMA 35 8.6 Amphetaminesp-Tyramine 33 3.5 Endogenous
ligandsc
4-APB 26 4.8 AmphetaminesMDAI 19 7.9 Aminoindanes2C-B 14 1.4 Phenethylamines2-AI 14 4.5 Aminoindanes7-APB 11 5.7 AmphetaminesN-Methyl-2-AI 8.9 3.5 AminoindanesMethamphetamine 6.2 7.3 AmphetaminesCathinone 5.8 5.8 CathinonesMDA 4.9 6.2 Amphetamines2C-H 4.3 0.9 PhenethylaminesAmphetamine 4.2 5.3 Amphetaminesb-PEA 2.4 1.3 Endogenous
ligandsc
aSubstances are sorted according to human/rat ratios.bStructure class: tryptamines.cStructure class: phenethylamines.
140 Simmler et al.
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
Species differences in TAAR1/ligand interactions have beenpredicted from sequence alignment studies that compared thecritical residues for the binding of b-PEA, showing that aminoacids that correspond to the critical residues differ betweenrat, mouse, and human TAAR1 (Kratochwil et al., 2011). Site-directed mutagenesis studies have identified two locations inTAAR1 transmembrane domains 6 and 7, where amino acidsubstitutions markedly reduce or increase methamphetamineTAAR1 activation potencies in the rat and mouse TAAR1(Reese et al., 2014). Docking studies with a homologymodel forthe human TAAR1 (Cichero et al., 2013, 2014) could serve tofurther elucidate the essential amino acids that are requiredfor ligand binding and discover structural determinants forthe TAAR1 activity or inactivity of psychoactive substances.
Importantly, TAAR1 is a promising target for therapeuticdrugs for the treatment of substance use disorders, regardlessof species differences in the direct TAAR1 agonism propertiesof psychoactive substances. TAAR1 agonists that have beenreported in the literature are similarly potent at both humanand rat TAAR1. Furthermore, the efficacy that has beenreported in animal models is comparable to the efficacy thathas been reported in in vitro expression systems, which mayprovide a basis for predicting effective doses in humans. TheTAAR1 partial agonist RO5203648 effectively reduced cocaineself-administration and relapse to drug-seeking behavior inrats (Revel et al., 2012b; Pei et al., 2014), although cocaineis not a TAAR1 ligand itself. TAAR1 partial agonism mark-edly reduced cocaine-induced DA overflow in the nucleus
Fig. 2. Heat map illustrating the diversity of TAAR1 activity between individual substances and between human, rat, and mouse TAAR1. Thesubstances are sorted according to their rat TAAR1 activity (EC50 values). The compounds were split into the 52more active (A) and the 51 less active (B)rat TAAR1 ligands. Next to the substance names, the respective substance classes are specified by color code and basic chemical structures are definedby numbers. The residues R2–R11 refer to the chemical structures presented in Fig. 1. Underlined residues indicate ring structures between twolocations for derivatization. The generic or full chemical names for abbreviated substances are listed in Supplemental Table 1.
Psychoactive Substances and TAAR1 141
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
accumbens (Pei et al., 2014). Because TAAR1 is involved in theconstitutive regulation of neuronal firing (Bradaia et al.,2009), pharmacological TAAR1 activation with a therapeuticdrug may regulate neuronal firing and result in hyposensi-tivity to drugs, similar to the overexpression of TAAR1 in atransgenic mouse model (Revel et al., 2012a). Moreover, theefficacy of these potentially therapeutic compounds could beeven greater in humans than in rodents. In rodents, but not orless so in humans, TAAR1-mediated negative feedback thatis induced by the abused substances could be present andattenuate the extent of therapeutic drug effects.Ex vivo electrophysiology experiments in the ventral teg-
mental area and dorsal raphe nuclei showed that both partialagonists and antagonists enhanced DA and 5-HT neuron
firing rates in WT mice (Bradaia et al., 2009; Revel et al.,2012b), whereas full agonists like p-tyramine decreased firingrates (Revel et al., 2011, 2012b). However, both full and partialagonists have been shown to be protective against the re-warding and reinforcing effects of the psychostimulant cocaine(Pei et al., 2015), but the opposing effects of full and partialagonists on firing rates suggest that psychoactive substancesthat are full agonists would exert effects that are differentfrom partial agonists. Data on the full or partial agonistproperties of TAAR1 ligands are thus important. Whereas fullagonists such as amphetamine induce negative feedback toblunt their own effect on DA and 5-HT systems (Lindemannet al., 2008; Di Cara et al., 2011), partial agonists mightincrease their effect on neuronal signal transmission by
Fig. 2. Continued.
142 Simmler et al.
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
increasing firing rates via TAAR1. This is an assumption thatwould be based on findings with selective TAAR1 ligands andrequires further investigation.In our data set and based on data reported previously by two
different laboratories (Reese et al., 2007; Wainscott et al.,2007), the activation potencies of b-PEA at rat, human, andmouse TAAR1 exhibited similar EC50 values between species,whereas p-tyramine was more potent at rat TAAR1, followedby mouse and human TAAR1. Given that these data weregenerated in independent laboratories that used differentassay conditions and expression systems, the similarities ofthe pharmacological profiles suggest good consistency of thedata and support the validity of the comparisons betweenspecies.In this study, we simply determined activity at specific
targets, which is common with interpretations of in vitro data,and we did not take into account that the processes that allowa substance to interact with TAAR1 in vivo depend on morevariables than solely substance/receptor interactions. Thelocation of TAAR1 expression is mostly intracellular inneurons (Miller, 2011) and also in glial cells (Cisneros andGhorpade, 2014). Because the substances need to reach thelocation of expression of TAAR1 to bind to the receptor, theintracellular availability of the ligands is also relevant.Certain psychoactive substances, such as amphetamine de-rivatives, are substrates of monoaminergic transporters andcarried into the cell (Zaczek et al., 1991). These substrate-typesubstances, therefore, might be more likely available tointracellular TAAR1 than substances that are not trans-porter substrates, including, for example, cocaine, MDPV, otherpyrovalerone cathinones, methylphenidate, and other pipra-drols (Simmler et al., 2013, 2014b). One limitation of our studyis that we did not consider stereoselectivity of the compoundsby screening racemic mixtures for most substances. As withactivity at other psychostimulant targets, such as monoamin-ergic reuptake transporters, TAAR1 has a stereoselective bind-ing site, and the assessment of racemates could underestimatethe activity of the more active isomer (Lewin et al., 2011).In conclusion, we provide an extensive data set on the ligand
properties of psychoactive substances at TAAR1. With differ-ences between activity at rodent and human TAAR1, weprovide evidence of significant species differences in interac-tions between TAAR1 and psychoactive drugs, which could berelevant to the translational validity of preclinical studies toclinical applications.
Acknowledgments
The authors thank Lipomed for providing the 2C andNBOMedrugsat no cost, Michael Arends for text editing, and Roger Norcross forhelpful discussions.
Authorship Contributions
Participated in research design: Hoener, Liechti.Conducted experiments: Buchy, Chaboz.Performed data analysis: Simmler, Buchy, Chaboz, Hoener.Wrote or contributed to the writing of the manuscript: Simmler,
Hoener, Liechti.
References
Achat-Mendes C, Lynch LJ, Sullivan KA, Vallender EJ, and Miller GM (2012)Augmentation of methamphetamine-induced behaviors in transgenic mice lackingthe trace amine-associated receptor 1. Pharmacol Biochem Behav 101:201–207.
Borowsky B, Adham N, Jones KA, Raddatz R, Artymyshyn R, Ogozalek KL, DurkinMM, Lakhlani PP, Bonini JA, and Pathirana S, et al. (2001) Trace amines:
identification of a family of mammalian G protein-coupled receptors. Proc NatlAcad Sci USA 98:8966–8971.
Bradaia A, Trube G, Stalder H, Norcross RD, Ozmen L, Wettstein JG, Pinard A,Buchy D, Gassmann M, and Hoener MC, et al. (2009) The selective antagonistEPPTB reveals TAAR1-mediated regulatory mechanisms in dopaminergic neuronsof the mesolimbic system. Proc Natl Acad Sci USA 106:20081–20086.
Bunzow JR, Sonders MS, Arttamangkul S, Harrison LM, Zhang G, Quigley DI,Darland T, Suchland KL, Pasumamula S, and Kennedy JL, et al. (2001) Am-phetamine, 3,4-methylenedioxymethamphetamine, lysergic acid diethylamide, andmetabolites of the catecholamine neurotransmitters are agonists of a rat traceamine receptor. Mol Pharmacol 60:1181–1188.
Cichero E, Espinoza S, Franchini S, Guariento S, Brasili L, Gainetdinov RR,and Fossa P (2014) Further insights into the pharmacology of the human traceamine-associated receptors: discovery of novel ligands for TAAR1 by a virtualscreening approach. Chem Biol Drug Des 84:712–720.
Cichero E, Espinoza S, Gainetdinov RR, Brasili L, and Fossa P (2013) Insights intothe structure and pharmacology of the human trace amine-associated receptor 1(hTAAR1): homology modelling and docking studies. Chem Biol Drug Des 81:509–516.
Cisneros IE and Ghorpade A (2014) Methamphetamine and HIV-1-induced neuro-toxicity: role of trace amine associated receptor 1 cAMP signaling in astrocytes.Neuropharmacology 85:499–507.
Cotter R, Pei Y, Mus L, Harmeier A, Gainetdinov RR, Hoener MC, and Canales JJ(2015) The trace amine-associated receptor 1 modulates methamphetamine’sneurochemical and behavioral effects. Front Neurosci 9:39.
Di Cara B, Maggio R, Aloisi G, Rivet JM, Lundius EG, Yoshitake T, Svenningsson P,Brocco M, Gobert A, and De Groote L, et al. (2011) Genetic deletion of trace amine 1receptors reveals their role in auto-inhibiting the actions of ecstasy (MDMA).J Neurosci 31:16928–16940.
Espinoza S, Lignani G, Caffino L, Maggi S, Sukhanov I, Leo D, Mus L, Emanuele M,Ronzitti G, and Harmeier A, et al. (2015) TAAR1 modulates cortical glutamateNMDA receptor function. Neuropsychopharmacology 40:2217–2227.
Harkness JH, Shi X, Janowsky A, and Phillips TJ (2015) Trace amine-associatedreceptor 1 regulation of methamphetamine intake and related traits. Neuro-psychopharmacology 40:2175–2184.
Jing L and Li JX (2015) Trace amine-associated receptor 1: a promising target for thetreatment of psychostimulant addiction. Eur J Pharmacol 761:345–352.
Kratochwil NA, Gatti-McArthur S, Hoener MC, Lindemann L, Christ AD, Green LG,Guba W, Martin RE, Malherbe P, and Porter RH, et al. (2011) G protein-coupledreceptor transmembrane binding pockets and their applications in GPCR researchand drug discovery: a survey. Curr Top Med Chem 11:1902–1924.
Lewin AH, Miller GM, and Gilmour B (2011) Trace amine-associated receptor 1 is astereoselective binding site for compounds in the amphetamine class. Bioorg MedChem 19:7044–7048.
Lewin AH, Navarro HA, and Mascarella SW (2008) Structure-activity correlations forbeta-phenethylamines at human trace amine receptor 1. Bioorg Med Chem 16:7415–7423.
Liechti M (2015) Novel psychoactive substances (designer drugs): overview andpharmacology of modulators of monoamine signaling. Swiss Med Wkly 145:w14043.
Lindemann L, Meyer CA, Jeanneau K, Bradaia A, Ozmen L, Bluethmann H, BettlerB, Wettstein JG, Borroni E, and Moreau JL, et al. (2008) Trace amine-associatedreceptor 1 modulates dopaminergic activity. J Pharmacol Exp Ther 324:948–956.
Lynch LJ, Sullivan KA, Vallender EJ, Rowlett JK, Platt DM, and Miller GM (2013)Trace amine associated receptor 1 modulates behavioral effects of ethanol. SubstAbuse 7:117–126.
Miller GM (2011) The emerging role of trace amine-associated receptor 1 in thefunctional regulation of monoamine transporters and dopaminergic activity.J Neurochem 116:164–176.
Pei Y, Lee J, Leo D, Gainetdinov RR, Hoener MC, and Canales JJ (2014) Activation ofthe trace amine-associated receptor 1 prevents relapse to cocaine seeking. Neuro-psychopharmacology 39:2299–2308.
Pei Y, Mortas P, Hoener MC, and Canales JJ (2015) Selective activation of the traceamine-associated receptor 1 decreases cocaine’s reinforcing efficacy and preventscocaine-induced changes in brain reward thresholds. Prog NeuropsychopharmacolBiol Psychiatry 63:70–75.
Reese EA, Bunzow JR, Arttamangkul S, Sonders MS, and Grandy DK (2007) Traceamine-associated receptor 1 displays species-dependent stereoselectivity for iso-mers of methamphetamine, amphetamine, and para-hydroxyamphetamine.J Pharmacol Exp Ther 321:178–186.
Reese EA, Norimatsu Y, Grandy MS, Suchland KL, Bunzow JR, and Grandy DK(2014) Exploring the determinants of trace amine-associated receptor 1’s functionalselectivity for the stereoisomers of amphetamine and methamphetamine. J MedChem 57:378–390.
Revel FG, Meyer CA, Bradaia A, Jeanneau K, Calcagno E, André CB, Haenggi M,Miss MT, Galley G, and Norcross RD, et al. (2012a) Brain-specific overexpression oftrace amine-associated receptor 1 alters monoaminergic neurotransmission anddecreases sensitivity to amphetamine. Neuropsychopharmacology 37:2580–2592.
Revel FG, Moreau JL, Gainetdinov RR, Bradaia A, Sotnikova TD, Mory R, Durkin S,Zbinden KG, Norcross R, and Meyer CA, et al. (2011) TAAR1 activation modulatesmonoaminergic neurotransmission, preventing hyperdopaminergic and hypo-glutamatergic activity. Proc Natl Acad Sci USA 108:8485–8490.
Revel FG, Moreau JL, Gainetdinov RR, Ferragud A, Velázquez-Sánchez C, SotnikovaTD, Morairty SR, Harmeier A, Groebke Zbinden K, and Norcross RD, et al. (2012b)Trace amine-associated receptor 1 partial agonism reveals novel paradigm forneuropsychiatric therapeutics. Biol Psychiatry 72:934–942.
Revel FG, Moreau JL, Pouzet B, Mory R, Bradaia A, Buchy D, Metzler V, Chaboz S,Groebke Zbinden K, and Galley G, et al. (2013) A new perspective for schizophrenia:TAAR1 agonists reveal antipsychotic- and antidepressant-like activity, improvecognition and control body weight. Mol Psychiatry 18:543–556.
Psychoactive Substances and TAAR1 143
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from
Rickli A, Hoener MC, and Liechti ME (2015a) Monoamine transporter and receptorinteraction profiles of novel psychoactive substances: para-halogenated amphet-amines and pyrovalerone cathinones. Eur Neuropsychopharmacol 25:365–376.
Rickli A, Kopf S, Hoener MC, and Liechti ME (2015b) Pharmacological profile ofnovel psychoactive benzofurans. Br J Pharmacol 172:3412–3425.
Rickli A, Luethi D, Reinisch J, Buchy D, Hoener MC, and Liechti ME (2015c) Receptorinteraction profiles of novel N-2-methoxybenzyl (NBOMe) derivatives of 2,5-dimethoxy-substituted phenethylamines (2C drugs). Neuropharmacology 99:546–553.
Scanlan TS, Suchland KL, Hart ME, Chiellini G, Huang Y, Kruzich PJ, Frascarelli S,Crossley DA, Bunzow JR, and Ronca-Testoni S, et al. (2004) 3-Iodothyronamine isan endogenous and rapid-acting derivative of thyroid hormone.Nat Med 10:638–642.
Simmler LD, Buser TA, Donzelli M, Schramm Y, Dieu LH, Huwyler J, Chaboz S,Hoener MC, and Liechti ME (2013) Pharmacological characterization of designercathinones in vitro. Br J Pharmacol 168:458–470.
Simmler LD, Rickli A, Hoener MC, and Liechti ME (2014a) Monoamine transporterand receptor interaction profiles of a new series of designer cathinones. Neuro-pharmacology 79:152–160.
Simmler LD, Rickli A, Schramm Y, Hoener MC, and Liechti ME (2014b) Pharma-cological profiles of aminoindanes, piperazines, and pipradrol derivatives. BiochemPharmacol 88:237–244.
Wainscott DB, Little SP, Yin T, Tu Y, Rocco VP, He JX, and Nelson DL (2007)Pharmacologic characterization of the cloned human trace amine-associated re-ceptor1 (TAAR1) and evidence for species differences with the rat TAAR1.J Pharmacol Exp Ther 320:475–485.
Wolinsky TD, Swanson CJ, Smith KE, Zhong H, Borowsky B, Seeman P, Branchek T,and Gerald CP (2007) The trace amine 1 receptor knockout mouse: an animalmodel with relevance to schizophrenia. Genes Brain Behav 6:628–639.
Xie Z and Miller GM (2007) Trace amine-associated receptor 1 is a modulator of thedopamine transporter. J Pharmacol Exp Ther 321:128–136.
Zaczek R, Culp S, and De Souza EB (1991) Interactions of [3H]amphetamine with ratbrain synaptosomes. II. Active transport. J Pharmacol Exp Ther 257:830–835.
Zucchi R, Chiellini G, Scanlan TS, and Grandy DK (2006) Trace amine-associatedreceptors and their ligands. Br J Pharmacol 149:967–978.
Address correspondence to: Dr. Matthias E. Liechti, Division of ClinicalPharmacology and Toxicology, Department of Biomedicine, University Hospi-tal Basel, Hebelstrasse 2, CH-4031 Basel, Switzerland. E-mail: [email protected]
144 Simmler et al.
at ASPE
T Journals on January 11, 2020
jpet.aspetjournals.orgD
ownloaded from