brain sciences Article Acute DOB and PMA Administration Impairs Motor and Sensorimotor Responses in Mice and Causes Hallucinogenic Effects in Adult Zebrafish Micaela Tirri 1 , Luisa Ponzoni 2, Sabrine Bilel 1, Raffaella Arfè 1,3, Daniela Braida 2, Mariaelvina Sala 4 and Matteo Marti 1,5,* 1 Department of Morphology, Surgery and Experimental Medicine, Section of Legal Medicine and LTTA Centre, University of Ferrara, 44121 Ferrara, Italy; [email protected] (M.T.); [email protected] (S.B.); rfarfl@unife.it (R.A.) 2 Department of Medical Biotechnology and Translational Medicine, Università degli Studi di Milano, 20133 Milan, Italy; [email protected] (L.P.); [email protected] (D.B.) 3 Institute of Public Health, Section of Legal Medicine, Catholic University of Rome, 00168 Rome, Italy 4 CNR, Neuroscience Institute, 20162 Milan, Italy; [email protected] 5 Collaborative Center for the Italian National Early Warning System, Department of Anti-Drug Policies, Presidency of the Council of Ministers, 00184 Rome, Italy * Correspondence: [email protected]; Tel.: +39-0532-455781; Fax: +39-0532-455777 Received: 23 July 2020; Accepted: 20 August 2020; Published: 24 August 2020 Abstract: The drastic increase in hallucinogenic compounds in illicit drug markets of new psychoactive substances (NPS) is a worldwide threat. Among these, 2, 5-dimetoxy-4-bromo-amphetamine (DOB) and paramethoxyamphetamine (PMA; marketed as “ecstasy”) are frequently purchased on the dark web and consumed for recreational purposes during rave/dance parties. In fact, these two substances seem to induce the same effects as MDMA, which could be due to their structural similarities. According to users, DOB and PMA share the same euphoric effects: increasing of the mental state, increasing sociability and empathy. Users also experienced loss of memory, temporal distortion, and paranoia following the repetition of the same thought. The aim of this study was to investigate the effect of the acute systemic administration of DOB and PMA (0.01–30 mg/kg; i.p.) on motor, sensorimotor (visual, acoustic, and tactile), and startle/PPI responses in CD-1 male mice. Moreover, the pro-psychedelic effect of DOB (0.075–2 mg/kg) and PMA (0.0005–0.5 mg/kg) was investigated by using zebrafish as a model. DOB and PMA administration affected spontaneous locomotion and impaired behaviors and startle/PPI responses in mice. In addition, the two compounds promoted hallucinatory states in zebrafish by reducing the hallucinatory score and swimming activity in hallucinogen-like states. Keywords: DOB; hallucinogens; mice; PMA; prepulse inhibition; sensorimotor responses; zebrafish 1. Introduction In recent years, novel psychoactive substances (NPS) have emerged in response to legislative control and market trends [1]. NPS, also known as designer drugs, bath salts, or legal highs, are analogous or derivatives of controlled substances. Since 2012, the number of NPS seized in Europe has rapidly increased [2]. The large demand for stimulants and hallucinogens such as amphetamine, methamphetamine, 3, 4-metyhlendioxy methamphetamine (MDMA/Ecstasy), and LSD increased their production and trafficking in many EU Member States. The local synthesis and distribution of these substances limited their prohibition. Brain Sci. 2020, 10, 586; doi:10.3390/brainsci10090586 www.mdpi.com/journal/brainsci Brain Sci. 2020, 10, 586 2 of 20 As a central European product, MDMA/ecstasy gained popularity among adults and also teenagers. In 2018, the number of European MDMA/ecstasy users (aged 15–64) reached approximately 13.7 million. MDMA use isBrain more Sci. 2020 common, 10, x FOR PEER among REVIEW young people. In fact, it is estimated that 2%2 of 20 young adults (aged 15–34) haveAs used a central ecstasy European in 2019 product, [3]. InMDMA/ecstasy addition, numerous gained popularity NPS withamong stimulant adults and andalso entactogenic properties haveteenagers. been designed In 2018, tothe substitutenumber of MDMAEuropean andMDMA/ecstasy/or LSD in users the illicit(aged drug15–64) market. reached Among them are dimethoxylated-amphetaminesapproximately 13.7 million. MDMA such use as is 4-bromo-2,more common among 5-dimethoxyamphetamine young people. In fact, it is estimated (DOB; Figure1), that 2% of young adults (aged 15–34) have used ecstasy in 2019 [3]. In addition, numerous NPS with which is a syntheticstimulant analogous and entactogenic of mescaline properties have belonging been designed to the to substitute class of MDMA phenethylamines. and/or LSD in the Recently, DOB gained more popularityillicit drug market. as a drugAmong of them abuse are indimethox Italy [ylated-amphetamines4]. This drug is usuallysuch as 4-bromo-2, sold online 5- in the form of tablets anddimethoxyamphetamine blotters under a variety (DOB; Figure of names 1), which such is a synthetic as “Golden” analogous or of “LSD-25”. mescaline belonging Yet, DOB to belongs to the class of phenethylamines. Recently, DOB gained more popularity as a drug of abuse in Italy [4]. the D-series ofThis substituted drug is usually amphetamines sold online in the form with of hallucinogen-liketablets and blotters under e ffa ectsvariety similar of names to such that as of LSD, high in vivo potency,“Golden” and aor long “LSD-25”. duration Yet, DOB of action.belongs However,to the D-series their of substitu pharmacologicalted amphetamines eff withects are not fully hallucinogen-like effects similar to that of LSD, high in vivo potency, and a long duration of action. established [5].However, their pharmacological effects are not fully established [5]. Figure 1. FigureChemical 1. Chemical structures structures of of DOB DOB (2,(2, 5-di 5-dimetoxy-4-bromo-amphetamine)metoxy-4-bromo-amphetamine) and PMA and PMA (paramethoxyamphetamine). (paramethoxyamphetamine). It has been reported that synthetic analogous of mescaline, including DOB, are more potent to It has beeninduce reported hallucinations that than synthetic those of many analogous naturallyof occurring mescaline, hallucinogens including [6]. DOB DOB, interacts are with more potent to induce hallucinationsserotonergic than receptors, those with of the many highest naturally affinity for 5-HT2A occurring receptors hallucinogens [7,8]. Studies in vivo [6]. revealed DOB interacts with that DOB is 10 times more potent than 4-bromo-2, 5-dimethoxyphenethylamine (2C-B) in humans [9] serotonergic receptors,and double/triple with to thethe phenethylamine highest affinity analogous for 5-HT2A(2C-B) in mice receptors [10]. In addition, [7,8]. Studiesthe behavioralin vivo revealed that DOB is 10effects times of DOB more seem potent to be 5-HT2A-dependent than 4-bromo-2, [11–13]. 5-dimethoxyphenethylamine (2C-B) in humans [9] PMA (Figure 1) has been also designed to mimic the effects of MDMA. In fact, this drug is and double/tripleillegally to sold the in phenethylamine the form of powder or tablets analogous and is labeled (2C-B) as “amphetamine” in mice [10 ].or “ecstasy”. In addition, The ring the behavioral effects of DOBsubstitution seem to beenhances 5-HT2A-dependent the serotonergic potency, [11–13 hallucinogenic,]. and MDMA-like properties of PMA (Figuremethamphetamine/amphetamine1) has been also designed drugs [14]. Incidental to mimic deaths the have e beenffects reported of MDMA. after PMA and In its fact, this drug derivative PMMA having a high toxicity, but systemic data are not available [15–17]. In particular, is illegally soldPMA in appears the form to be more of powder toxic than orMDMA tablets at low and doses is [18] labeled and often, as the “amphetamine” effect is delayed by or “ecstasy”. The ring substitutioningestion [19]. enhances In vitro studies the serotonergic confirm that PMA potency, potency hallucinogenic,is 100 times more potent and than MDMA-like MDMA [20] properties of methamphetamineand this/amphetamine is due to PMA’s ability drugs to release [14]. and Incidental inhibit the reuptake deaths of have 5-HT been[21]. reported after PMA and its Effects and dosage of DOB and PMA have been reported by psychonauts [22]. Both are identified derivative PMMAas “pattern having recognition a high enhancement” toxicity, but and systemic considered dataas psychedelic are not availablesubstances with [15 –strong17]. In particular, PMA appearscognitive to be moreeffects, toxiceven at than low MDMAdoses, as anxiety, at low agitation, doses [mild18] andparanoia, often, increased the emusicalffect is delayed by perception, temporal distortion, and double vision, are some of the most reported effects by users ingestion [19].[23].In However, vitro studies there are confirm no currently that available PMA pr potencyeclinical data is on 100 thetimes major psychopharmacological more potent than MDMA [20] and this is dueeffects to PMA’s of DOB and ability PMA. to release and inhibit the reuptake of 5-HT [21]. Effects and dosageOn this basis, of DOB the purpose and PMA of this havestudy is been to evaluate reported in vivo by the psychonauts acute sensorimotor [22 alteration]. Both are identified and hallucinogenic properties, typical of phenethylamine and amphetamine-like compounds in the as “pattern recognitionclassical murine enhancement” and the zebrafish and models. considered as psychedelic substances with strong cognitive effects, even at low doses, as anxiety, agitation, mild paranoia, increased musical perception, temporal distortion, and double vision, are some of the most reported effects by users [23]. However, there are no currently available preclinical data on the major psychopharmacological