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Home , MDMA, Tryptamine

Ps>cnopnarmaco,og: I_v?.,, . _.,:._,,* _'_, lPsyclloptlarmacology 4 _' Sprmger-Verlag 1993

Behavioral characterization of -ethyltryptamine, a derivative with MDMA-Iike properties in rats

Kirsten M. Krebs _, Mark A. Geyer 2 _Department of Psychology, University of California at San Diego, La JoiN, CA 92093. USA 2Department of Psychiatr3. University of California at San Diego, La Jolla, CA 92093. USA

Received 19 July 1993 Final version 3 August 1993 _,

Abstract. Several reports have speculated that the {MDMA: Ecstasy) in street sates(Federal Register 1993); a tryptamme-derived alpha-ethyltryptamine IAET_ :\ET may produce neurotoxicit) {Huang et al. iz max imve effects similar to those of the - 199l): and deaths have been reported after ingestion of b derived drug 3 4-methylenedioxymethamphetan_ine AET (Daldrup et al. 1986: Federal Register 1993). Both a (MDIVlA). Indeed, the US Drug Enforcement Adminis- in vivo and in vitro studies of AET have revealed that it b il tration has recently placed AET on the Schedule I list inhibits activity and increases sero- ti because of its putative similarity to MDMA. The Behav- tonin and release (Greig et al. 1959; Baker et e: ioral Pattern Monitor, which quantifies locomotor and at. 1980}. Systematic behavioral studies of AET are few, ti investigatory responses of rats, was used to characterize however. The behaviors that have been examined, such ti the effects of AET in a paradigm that distinguishes be- as ejaculator>, response (Renyi 1986), serotonin syn- si tween the effects of traditional , am- drome (Renyi and Ross 1985), and cage-leaving r.esponse h phetamine-like , and MDMA-like . First, (Renyi et al. 1986), have not been designed to evaluate d, a dose-response study revealed that all doses of AET AET in relation to other drugs with abuse liability, with tested (5, 10, 20 rog/kg) significantly increased locomotor the exception of amphetamine (Renyi and Ross 1985; activity. Locomotor hyperactivity is produced by MD- Renyi 1986; Renyi et al. 1986), In order to test this drug MA or amphetamine-like stimulants, but not by classical in a paradigm that has been used extensively to profile u hallucinogens, such as LSD or . Additionally, hallucinogens and MDMA-like drugs, as well as stimu- c AET significantly decreased measures of investigatory lants, we performed a dose-response study of AET on behavior. Similar decreases occur with MDMA or hallu- rats tested in the Behavioral Pattern Monitor (BPM), a cv cinogen administration, but not with amphetamine-like system for quantifying the locomotor and investigatory a administration. Second, as with MDMA, the responses of rodents. Previous studies have shown that it c_ locomotor hyperactivity induced by AET was attenuated is the serotonin-releasing properties of MDMA and re- by pretreatment (10 rog/kg) with the serotonin reuptake lated drugs that are responsible for their locomotor acti- inhibitor fiuoxetine. Thus, AET, a tryptamine-derived rating effects (Callaway et al. 1990, 1991}. First, a deriva- drug, appears to produce an MDMA-like profile of be- tive of MDMA, MBDB, that releases serotonin but not havioral changes by virtue of releasing presynaptic sero- dopamine, has behavioral effects very similar to those of tonin. MDMA(Callawayet al. 1991).Second,the hyperactivity produced by MDMA is prevented by depletion of brain Re)' words: Alpha-ethyltryptamine - Monase - Fluox- serotonin or pretreatment with serotonin antagonists etine MDMA Locomotion Rats-Serotonin (Callaway et al. 1990). Third, the locomotor activating effects of MDMA could be attenuated by pretreatment with or other inhibitors of serotonin uptake v (Callaway et al. 1990). Hence, the locomotor hyperactivi- u Recent interest in the compound alpha-ethyltryptamine ty produced by MDMA-like drugs is attributable to their -_, (AET), a tryptamine derivative previously marketed as actions as indirect serotonin . --- Monase, has been stimulated by its inclusion on the After observing that AET produced an MDMA-like _, Schedule I list by the US Drug Enforcement Administra- profile of behavioral changes in the BPM, pretreatment tion (DEA) (Federal Register 1993). The scheduling of with fluoxetine was used to further investigate the simi- AET was justified by the DEA because: AET has been larities between AET and MDMA. This experiment test- substituted for 3,4-methylenedioxymethamphetamine ed the hypothesis that, like MDMA, AET produces loco-

Correspondem'toe .' K. M. Krebs pendent release of presynaptic serotonin.

motor hyperactivity by virtue of an uptake carrier,de_ i,,I_i, ,_ 285 f

Materials and methods Procedure. Animals were tested in the dark and during the dark phase of their light cycle. Approximately 24 h before testing, ant- Animals. Male Sprague-Dawley rats (Harlan Industries, San Diego, reals were taken to the testing room, weighed, handled briefly, Calif.} weighing 275-350 g were used. Animals were housed two or placed in a clear Plexiglas 'box (24 x 46 em) for 30 s, and then three per cage under a 12-h reverse light cycle (lights off: 0700 returned to their cages in the animal room. On the day of testing, hours). Food and water were available ad libitum. Animals were animals were brought to the testing room and allowed to sit for 60 allowed to acclimatize for approximately 1 week after arrival, min. Injections were administered under red lights in the testing room. Animals were tested for 120 min. In experiment 1, four Behat,ioral testing. All behavior was measured in the BPM, a groups (n = 7-10) of 35 rats received saline, 5, 10, or 20 mg/kg AET 30.5 x 61,0ce black Plexiglas chamber housed in a wooden 10 mtn before testing. sound-attenuating box. Ten 2.5-cm holes are placed in the chamber Based on the results from experiment 1, 5 mg/kg AET was se- (three in each long wall, one in one short 'wall, and three in the leered for further study because it had submaximal effects, but reit- floor). Photocells in each hole detect investigatory holepokes. A ably increased activity. Because previous research with MDMA has touchplate. 1'5.2cm above the floor, allows detection of rearings demonstrated that only the locomotor activating effects of MDMA when contact is made by the animal between the metal floor and the were attenuated by fluoxetine pretreatment (Callaway et al. 1990), metal touchplate. A 4 x 8 grid of infrared photobeams detects the we examined only locomotor activity in experiment 2. The most animal's position in an X-Y plane. A computer continuously mon- sensitive time resolution for activity was determined to be the sec- itors all photobeams and the touchplate and stores the data for later ond hour of the 2-h test. Four groups of ten rats each received analysis, pretreatments of either saline or 10 mg/kg fiuoxetine 60 mtn before testing. Ten minutes before testing, rats were treated with either Ana/rsix. Tile raw data were reduced to: the X and '5' coordinates saline or 5 mg/kg AET. One rat, for unknown reasons, failed to of the rat in the chamber: the occurrence of hotepokes or rearings: respond throughout the test session and was omitted because it was and tile amounl of time spent at a particular coordmatc or perform- more than two standard deviations below the group mean for sever- lng a particular bch;.lxi,or. LOCOIllOtOFactivitx x_a_ quantified both al measures. Nevertheless. specific comparisons belween groups by thc total number of photohcam breaks in the BPM mlovementsJ were unaffected by Ibis exclusion. as a measure of total motor activity, and bv thc number of crossmgs between any of eight equal square sectors within the BPM {cross- Drt_gs. Drugs used were: AET (Sigma Chemicals, St. Louis. Mo3 ings) as a measure of horizontal locomotion. The number and dura- and fluoxetine (Lilly, Indianapolis, Ind.). Drugs were prepared fresh tion of both holepokes and rearings were calculated. Data were first daily. AET (salt weight) was dissolved in nitrogen-purged 0.9% examined in lO-mtn time resolutions by two-way ANOVA with saline and injected subcutaneously in a volume of I ml/kg. Fluox- time as a repeated measure. When there were significant interac- etine was dissolved in 0.9% saline and injected intraperitoneally in tions with time (including Greenhouse-Geisser corrections for non- a volume of 2 mi/kg. sphericity), further time resolutions were examined. Specific post hoc comparisons were done using Newman-Keuls, Significance was demonstrated by surpassing an alpha level of 0.05.

200 60

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0 ' ' ' 0 ' ' ' [ ' ' i t i _ i 1 2 a 4 6 e ? . g lOllta _, a 4 _ e 7 · * 1o 1112 10 Minute Blocke 10 Minute Blocks (A)

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800 80

400 m Fig. 1. All doses of AET (5, 10, 20 mg/kg) signifi- , * Z 4o cantly increased horizontal locomotor activity (A) u, a00 _: and global motor activity (C) over the 2-h test ses- _ sion.AlldosesofAETsignificantlydecreasedin- O a00 _ zo vestigatory behaviors of holepokes (B) and rearings

for successive 10-mtn intervals. · Saline; 57 5 mg/ o ...... o kg AET; · 10 rog/kg AET; [] 20 rog/kg AET ,00 2 3 _¥4 /0'xl)..OxO8 7 8/O-IDii 10 11 12 _o il .._xX.xl______4 _' 7 8 Ii 10 11 1_2 (D) over 2 h. Data are presented as group means 10 Mlnute Block,, 10 Minute Blocks (c) (O) FLUOXETINE ANTAGONISM Discussion OF AET .** The behavioralprofileproduced by AET is strikingly

7** from theeffectsof eithertraditional hallucinogensor am- co '** phetamine-like stimulants. The present studies of the ef- t9 Il00 feetsofacutelyadministeredAETdemonstratedthat all z doses of AET significantly increased locomotor activity co 400 I ' similarto that of MDMA and is readilydistinguishable ne similarstructure,may providea tool that couldshed O 2.0 light on the structure-activityrelationshipmediatingthe _o.[- common behavioral effects of these disparate com- O 5.0o-l_ _ _ pounds.mimic theSimilareffectseffectsof MDMA-Iikeare alsoseendrugs,in ratsdespitetreateditswithdis- coPRETRt/ EATS: AL SAL FLU FL_1 U 'MDMAover the 2(Gold-h testetsession.al. 1988;TheCallawayability ofet AETal. 1990,to closely1991) '_ TREATMENT:SAL AE:T $AL AET and amphetamine (Callaway et al. 1990), but not in rats , Fig. 2. Acute treatment with AET (SAL/AET)15mg/kg}significant- treated with classical hallucinogens, such as LSD or mes- ly increased locomotor activity in the second hour of a 2-h test. This caline (Adams and Geyer 1985: Geyer and Krebs 1993). effect was attenuated by pretreatment with fluoxetine (FLU.4E23. Additionally, AET significantly decreased measures of Data are presented as group means _+SEM. *Significantlydifferent investigatory behavior. Again. similar effects are ob- from all other groups. P 0.05 served with MDMA administration IGold et al. 1988; Callaway et al. 1990. 1991) and with admin- Results istration (Adams and Geyer 1985:Geyer and Krebs 1993; Krebs and Geyer 19931, but not with stimulant Experiment I administration (Callaway et al. 1990). Therefore, the pat- tern of results observed with AET administration, name- AET significantly increased locomotor activity over the ly increased locomotor activity and decreased investiga- 2-h test session, as measured both by crossings [F(3, tory behavior, is rerfiarkably similar to the pattern pro- 31) = 6.54] (Fig. lA) and movements [F(3, 31) = 7.27] duced by MDMA, but not by traditional hallucinogen or (Fig. lC). AET also significantly decreased investigatory stimulant administration. activity, as measured by holepokes [F(3, 31)= 8.02] Experiment 2 demonstrated that tire serotonin reup- (Fig. IB) and rearings [F(3, 31) = 25.42] (Fig. ID) over take inhibitor fluoxetine attenuated the locomotor hy- the 2-h test session. Because activity measures had signif- peractivity produced by AET. A similar effect was also leant time-by-AET interactions [F(33, 341) = 8.97- seen with MDMA (Callaway et al. 1990), indicating that 10.43], data were examined for the first and second hour. serotonin release is necessary for MDMA-Iike drugs to The second hour was the more sensitive measure of exert their locomotor activating effects. Accordingly, the AET's locomotor activating effects. AET significantly in- fluoxetine-induced attenuation of AET's effects on loco- creased activity, as measured by both crossings [F(3, motor hyperactivity indicates that AET acts via seroton- 31) = 14.81] and movements [F(3, 31) = 16.89]. Post hoc in release. analyses revealed that all doses were significantly differ- Thus, in the present studies in rats, AET has been ent from saline. For movements, post hoc analyses also found to be similar to MDMA-Iike drugs in two respects: demonstrated dose-dependent effects of AET because be- (1) AET increases locomotor activity and decreases in- tween doses, 5 mg/kg AET was significantly different vestigatory activity; and (2) AET's effects on locomotor from saline and both 10 mg/kg and 20 mg/kg. Lower activity can be attenuated by pretreatment with fluox- doses of AET (1.25-2.5 rog/kg) produce qualitatively sim- etine. These results lead to the conclusion that AET ex- ilar, but diminished, effects (K.M. Krebs and M.A. Geyer, erts these effects through serotonin release. This apparent unpublished observations), similarity in the effects of AET and MDMA in rats is consistent with reports that AET has been sold on the street as an MDMA-like drug (Federal Register 1993). Experiment 2 These observations of AET's similarity in effects to MDMA are especially interesting when we consider the There was a significant interaction between fluoxetine differences in structure between these compounds. MD- pretreatment and AET treatment for crossings IF(I, MA and its related drugs are amphetamine derivatives, 35) = 4.51] and for movements IF(l, 35) = 7.04] for the whereas AET is a tryptamine derivative that is much second hour of testing (Fig. 2), indicating that the in- more closely related to serotonin. This observation could crease in both crossings and movements produced by be important in the understanding of the nature of the AET was reduced by fluoxetine pretreatment. Post hoc structure-activity relationships of these drugs. Further analyses revealed that, for both crossings and move- studies should now be undertaken to provide converging ments, the fluoxetine-pretreated/AET-treated group was evidence for the relevance of presynaptic serotonin re- different only from the saline-pretreated/AET-treated lease to the effects of AET and to determine whether group, related derivativesof tryptaminehave similareffects.If ' ih_r_ is a common mechanism for producing the locomo- Daldrup T Heller C, Matthiesen U, Honus S, Bresges A, Haarhof K tor activating effects seen with both AET and the (19861 Etryptamine, a new with a fatal effect. Z -substituted phenalkylamines, then the Rechtsmed97:61-68 combined study of both classes of compounds could FederalRegister (1993)58(47): 13533,March 12 Geyer MA, Krebs KM (19931 Serotonin receptor involvement in an provide critical insights into the pharmacological actions animal model of the acute effects of hallucinogens. In: Lin G (ed) subserving the subjective effects of indirect serotonin ag- Hallucinogens: an update. NIDA Research Monograph Series, onists. National Institute on Drug Abuse, Rocky)lie, Maryland (in press) Acknowledgements. The authors thank V. Lehmann-Masten and Gold LH, Koob GF, Geyer MA (19881 Stimulant and hallucino- D. Giracello for technical assistance and Dr. L. Erinofffor suggest- genic behavioral profiles of 3,4-methylenedioxymetham- lng investigations of AET. This work was supported by a grant from phetamine and N-ethyl-3,4-methylenedioxymethamphetamine the National Institute on Drug Abuse (DA02925). M. Geyer was m rats. J Pharmacol Exp Ther 247:547-555 supported by a Research Scientist Development Award from the Grieg ME, Walk RA, Gibbons AJ (19591 The effect of three National Institute of Mental Health (MH001881. tryptamine derivatives on serotonin in vitro and in vivo. J Pharmacol Exp Ther 127:110-115 Huang X, Johnson MP. Nichols DE (19911 Reduction in brain serotonin markers by alpha-ethyltryptamine (Monase). Eur J References Pharmacol200:187-190 Krebs KM, Geyer MA (19931 Cross-tolerance studies of serotonin :tdams I,M. (;ever MA (1985} A proposed animal model for hallu- receptors involved ill behavioral effects of LSD in rats. Psy- cmogem, based on LSD's effects on patterns of exploration ill chopharmacology (in press) rats. Bchav Neurosci 99:881 900 Renvi L {1986}The effects of monoamine oxidase inhibitors on the Baker (;B. Hiob kIi. I)exvhursl W(, (198(})Effccts of monoaminc cjaculalory response induced bx 5-methoxy.-N,N-dimethyl- oxtdasc mhib,ors on rclcasc oF dopammc and 5-hxdrox- tryptamine in the rat. Br J Pharmacol 88:827-835 .xtrvp. lammc from rat strmtum in xitro. ('cll Mol Bio] _6.'_ · I82 RenyiL, RossSB (19851Effectsof am)flare)neand relatedcom- 186 poundson the accumulationof biogenicmonoaminesin rat Callaway ('W. Wing LL (ieyer MA (19901 Serotonin release con- brain slices in vitro and ex vivo in relation to their behavioural tributes to thc locomotor stimulant effects of 3.4-methylene- effects. Acta Pharmacol Toxicol 56:416-426 dioxymethanlphetamme in rals. .1 Pharmacol Exp Ther Renyi L, Archer T, Minor BG, Tandberg B, Fredriksson A, Ross SB 254:456 464 (1986) The inhibition of the cage-leaving response - a model for ('alJawa.x ('W, .Iohnson MP, (iold LH, Nichols DE, Geyer MA studies of thc neurotransmission in the rat. J Neu- ( 19911Amphetanfinc derivatives )nd ucc locomotor hyperactivi- ral Transm 65:193-210 ly by acting as indirect scrolonin agonists. 1(}4:293 301