N, N-Dimethyltryptamine, a natural hallucinogen, ameliorates Alzheimer's disease by restoring neuronal Sigma-1 receptor-mediated endoplasmic reticulum-mitochondria crosstalk.

BACKGROUND: Aberrant neuronal Sigma-1 receptor (Sig-1r)-mediated endoplasmic reticulum (ER)- mitochondria signaling plays a key role in the neuronal cytopathology of Alzheimer's disease (AD). The natural psychedelic N, N-dimethyltryptamine (DMT) is a Sig-1r agonist that may have the anti-AD potential through protecting neuronal ER-mitochondrial interplay. METHODS: 3×TG-AD transgenic mice were administered with chronic DMT (2 mg/kg) for 3 weeks and then performed water maze test. The Aß accumulation in the mice brain were determined. The Sig-1r level upon DMT treatment was tested. The effect of DMT on the ER-mitochondrial contacts site and multiple mitochondria-associated membrane (MAM)-associated proteins were examined. The effect of DMT on calcium transport between ER and mitochondria and the mitochondrial function were also evaluated. RESULTS: chronic DMT (2 mg/kg) markedly alleviated cognitive impairment of 3×TG-AD mice. In parallel, it largely diminished Aß accumulation in the hippocampus and prefrontal cortex. DMT restored the decreased Sig-1r levels of 3×TG-AD transgenic mice. The hallucinogen reinstated the expression of multiple MAM-associated proteins in the brain of 3×TG-AD mice. DMT also prevented physical contact and calcium dynamic between the two organelles in in vitro and in vivo pathological circumstances. DMT modulated oxidative phosphorylation (OXPHOS) and ATP synthase in the in vitro model of AD. CONCLUSION: The anti-AD effects of DMT are associated with its protection of neuronal ER-mitochondria crosstalk via the activation of Sig-1r. DMT has the potential to serve as a novel preventive and therapeutic agent against AD.

5-MeO-DMT induces sleep-like LFP spectral signatures in the hippocampus and prefrontal cortex of awake rats.

5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is a potent classical psychedelic known to induce changes in locomotion, behaviour, and sleep in rodents. However, there is limited knowledge regarding its acute neurophysiological effects. Local field potentials (LFPs) are commonly used as a proxy for neural activity, but previous studies investigating psychedelics have been hindered by confounding effects of behavioural changes and anaesthesia, which alter these signals. To address this gap, we investigated acute LFP changes in the hippocampus (HP) and medial prefrontal cortex (mPFC) of freely behaving rats, following 5-MeO-DMT administration. 5-MeO-DMT led to an increase of delta power and a decrease of theta power in the HP LFPs, which could not be accounted for by changes in locomotion. Furthermore, we observed a dose-dependent reduction in slow (20-50 Hz) and mid (50-100 Hz) gamma power, as well as in theta phase modulation, even after controlling for the effects of speed and theta power. State map analysis of the spectral profile of waking behaviour induced by 5-MeO-DMT revealed similarities to electrophysiological states observed during slow-wave sleep (SWS) and rapid-eye-movement (REM) sleep. Our findings suggest that the psychoactive effects of classical psychedelics are associated with the integration of waking behaviours with sleep-like spectral patterns in LFPs.

Chronic Behavioral and Neurochemical Effects of Four Novel N-Benzyl-2-phenylethylamine Derivatives Recently Identified as "Psychoactive" in Adult Zebrafish Screens.

Potently affecting human and animal brain and behavior, hallucinogenic drugs have recently emerged as potentially promising agents in psychopharmacotherapy. Complementing laboratory rodents, the zebrafish (Danio rerio) is a powerful model organism for screening neuroactive drugs, including hallucinogens. Here, we tested four novel N-benzyl-2-phenylethylamine (NBPEA) derivatives with 2,4- and 3,4-dimethoxy substitutions in the phenethylamine moiety and the -F, -Cl, and -OCF3 substitutions in the ortho position of the phenyl ring of the N-benzyl moiety (34H-NBF, 34H-NBCl, 24H-NBOMe(F), and 34H-NBOMe(F)), assessing their behavioral and neurochemical effects following chronic 14 day treatment in adult zebrafish. While the novel tank test behavioral data indicate anxiolytic-like effects of 24H-NBOMe(F) and 34H-NBOMe(F), neurochemical analyses reveal reduced brain norepinephrine by all four drugs, and (except 34H-NBCl) - reduced dopamine and serotonin levels. We also found reduced turnover rates for all three brain monoamines but unaltered levels of their respective metabolites. Collectively, these findings further our understanding of complex central behavioral and neurochemical effects of chronically administered novel NBPEAs and highlight the potential of zebrafish as a model for preclinical screening of small psychoactive molecules.

The 'PSILAUT' protocol: an experimental medicine study of autistic differences in the function of brain serotonin targets of psilocybin.

BACKGROUND: The underlying neurobiology of the complex autism phenotype remains obscure, although accumulating evidence implicates the serotonin system and especially the 5HT2A receptor. However, previous research has largely relied upon association or correlation studies to link differences in serotonin targets to autism. To directly establish that serotonergic signalling is involved in a candidate brain function our approach is to change it and observe a shift in that function. We will use psilocybin as a pharmacological probe of the serotonin system in vivo. We will directly test the hypothesis that serotonergic targets of psilocybin - principally, but not exclusively, 5HT2A receptor pathways-function differently in autistic and non-autistic adults. METHODS: The 'PSILAUT' "shiftability" study is a case-control study autistic and non-autistic adults. How neural responses 'shift' in response to low doses (2 mg and 5 mg) of psilocybin compared to placebo will be examined using multimodal techniques including functional MRI and EEG. Each participant will attend on up to three separate visits with drug or placebo administration in a double-blind and randomized order. RESULTS: This study will provide the first direct evidence that the serotonin targets of psilocybin function differently in the autistic and non-autistic brain. We will also examine individual differences in serotonin system function. CONCLUSIONS: This work will inform our understanding of the neurobiology of autism as well as decisions about future clinical trials of psilocybin and/or related compounds including stratification approaches. TRIAL REGISTRATION: NCT05651126.

Psilocybin for dementia prevention? The potential role of psilocybin to alter mechanisms associated with major depression and neurodegenerative diseases.

Major depression is an established risk factor for subsequent dementia, and depression in late life may also represent a prodromal state of dementia. Considering current challenges in the clinical development of disease modifying therapies for dementia, the focus of research is shifting towards prevention and modification of risk factors to alter the neurodegenerative disease trajectory. Understanding mechanistic commonalities underlying affective symptoms and cognitive decline may reveal biomarkers to aid early identification of those at risk of progressing to dementia during the preclinical phase of disease, thus allowing for timely intervention. Adult hippocampal neurogenesis (AHN) is a phenomenon that describes the birth of new neurons in the dentate gyrus throughout life and it is associated with spatial learning, memory and mood regulation. Microglia are innate immune system macrophages in the central nervous system that carefully regulate AHN via multiple mechanisms. Disruption in AHN is associated with both dementia and major depression and microgliosis is a hallmark of several neurodegenerative diseases. Emerging evidence suggests that psychedelics promote neuroplasticity, including neurogenesis, and may also be immunomodulatory. In this context, psilocybin, a serotonergic agonist with rapid-acting antidepressant properties has the potential to ameliorate intersecting pathophysiological processes relevant for both major depression and neurodegenerative diseases. In this narrative review, we focus on the evidence base for the effects of psilocybin on adult hippocampal neurogenesis and microglial form and function; which may suggest that psilocybin has the potential to modulate multiple mechanisms of action, and may have implications in altering the progression from major depression to dementia in those at risk.

Psychedelics and the 'inner healer': Myth or mechanism?

BACKGROUND: Reference to an intrinsic healing mechanism or an 'inner healer' is commonplace amongst psychedelic drug-using cultures. The 'inner healer' refers to the belief that psychedelic compounds, plants or concoctions have an intrinsically regenerative action on the mind and brain, analogous to intrinsic healing mechanisms within the physical body, for example, after sickness or injury. AIMS: Here, we sought to test and critique this idea by devising a single subjective rating item pertaining to perceived 'inner healing' effects. METHODS: The item was issued to 59 patients after a single high (25 mg, n = 30) or 'placebo' (1 mg, n = 29) dose of psilocybin in a double-blind randomised controlled trial of psilocybin for depression. RESULTS: Inner healer scores were higher after the high versus placebo dose of psilocybin (t = 3.88, p < 0.001). Within the high-dose sub-sample only, inner healer scores predicted improved depressive symptomatology at 2 weeks post-dosing. CONCLUSIONS: The principle of activating inner healing mechanisms via psychedelics is scientifically nascent; however, this study takes a positivist and pragmatic step forward, asking whether it warrants further examination.

Exploring psychedelic-assisted therapy in the treatment of functional seizures: A review of underlying mechanisms and associated brain networks.

Functional seizures (FS), the most common subtype of functional neurological disorder (FND), cause serious neurological disability and significantly impact quality of life. Characterized by episodic disturbances of functioning that resemble epileptic seizures, FS coincide with multiple comorbidities and are treated poorly by existing approaches. Novel treatment approaches are sorely needed. Notably, mounting evidence supports the safety and efficacy of psychedelic-assisted therapy (PAT) for several psychiatric conditions, motivating investigations into whether this efficacy also extends to neurological disorders. Here, we synthesize past empirical findings and frameworks to construct a biopsychosocial mechanistic argument for the potential of PAT as a treatment for FS. In doing so, we highlight FS as a well-defined cohort to further understand the large-scale neural mechanisms underpinning PAT. Our synthesis is guided by a complexity science perspective which we contend can afford unique mechanistic insight into both FS and PAT, as well as help bridge these two domains. We also leverage this perspective to propose a novel analytic roadmap to identify markers of FS diagnostic specificity and treatment success. This endeavor continues the effort to bridge clinical neurology with psychedelic medicine and helps pave the way for a new field of psychedelic neurology.

Mechanisms of therapeutic change after psychedelic treatment in OCD.

Novel treatments are required for the 30-50% of individuals with obsessive-compulsive disorder (OCD) who remain resistant to first-line pharmacological and psychotherapeutic treatments. Recent pilot data suggest benefit from psilocybin-assisted psychotherapy (PAP) and from imagery rescripting (ImRs). We explore psychological mechanisms of change underpinning both interventions that appear to allow for reprocessing of negative emotions and core beliefs associated with past aversive events. A next critical step in PAP is the development of psychotherapeutic frameworks grounded in theory. We propose that basing PAP on an ImRs framework may provide synergistic benefits in symptom reduction, modification of core beliefs, and value-based living.

In the new era of psychedelic assisted therapy: A systematic review of study methodology in randomized controlled trials.

Recent years have seen a resurgence in randomized, placebo controlled trials (RCTs) utilizing non-classical psychedelics (e.g. 3,4-methyl enedioxy methamphetamine [MDMA]), and classical psychedelics (e.g. psilocybin, lysergic acid diethylamide [LSD], and N,N-dimethyltryptamine [DMT/ayahuasca]) in conjunction with assisted therapy (AT) for psychiatric disorders. A notable methodological challenge in psychedelic AT, however, is the complexity of blinding procedures. The lack of efficacious blinding can introduce considerable response bias, reduce internal validity, and compromise participant retention. This systematic review examines design and blinding techniques in RCTs utilizing psychedelics and placebo for the treatment of psychiatric disorders. The aim of this work is to identify factors that may inform future RTC design for conducting psychedelics research. We conducted a systematic review of PubMed, MEDLINE, CINAHL, Cochrane Central Register of Controlled Trials (CENTRAL), Psycinfo, Embase, and Web of Science Core Collection to examine: (1) placebo selection, (2) study design, and (3) integrity of blinding measures. Sixteen publications were identified as meeting the criteria for a systematic review. Our findings suggest that traditional placebo administration is insufficient to control for expectancy confounds. Consequently, experimental methodology that limits personnel unblinding and the use of an active placebo are important considerations when designing prospective clinical studies involving psychedelics.

The potential of 5-methoxy-N,N-dimethyltryptamine in the treatment of alcohol use disorder: A first look at therapeutic mechanisms of action.

Alcohol use disorder (AUD) remains one of the most prevalent psychiatric disorders worldwide with high economic costs. Current treatment options show modest efficacy and relapse rates are high. Furthermore, there are increases in the treatment gap and few new medications have been approved in the past 20 years. Recently, psychedelic-assisted therapy with psilocybin and lysergic acid diethylamide has garnered significant attention in the treatment of AUD. Yet, they require significant amounts of therapist input due to prolonged subjective effects (~4-12 h) leading to high costs and impeding implementation. Accordingly, there is an increasing interest in the rapid and short-acting psychedelic 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT). This paper offers a first look at potential therapeutic mechanisms for AUD by reviewing the current literature on 5-MeO-DMT. Primarily, 5-MeO-DMT is able to induce mystical experiences and ego-dissolution together with increases in psychological flexibility and mindfulness. This could decrease AUD symptoms through the alleviation of psychiatric mood-related comorbidities consistent with the negative reinforcement and self-medication paradigms. In addition, preliminary evidence indicates that 5-MeO-DMT modulates neural oscillations that might subserve ego-dissolution (increases in gamma), psychological flexibility and mindfulness (increases in theta), and the reorganization of executive control networks (increases in coherence across frequencies) that could improve emotion regulation and inhibition. Finally, animal studies show that 5-MeO-DMT is characterized by neuroplasticity, anti-inflammation, 5-HT2A receptor agonism, and downregulation of metabotropic glutamate receptor 5 with clinical implications for AUD and psychiatric mood-related comorbidities. The paper concludes with several recommendations for future research to establish the purported therapeutic mechanisms of action.

LSD increases sleep duration the night after microdosing.

Microdosing psychedelic drugs at a level below the threshold to induce hallucinations is an increasingly common lifestyle practice. However, the effects of microdosing on sleep have not been previously reported. Here, we report results from a Phase 1 randomized controlled trial in which 80 healthy adult male volunteers received a 6-week course of either LSD (10 µg) or placebo with doses self-administered every third day. Participants used a commercially available sleep/activity tracker for the duration of the trial. Data from 3231 nights of sleep showed that on the night after microdosing, participants in the LSD group slept an extra 24.3 min per night (95% Confidence Interval 10.3-38.3 min) compared to placebo-with no reductions of sleep observed on the dosing day itself. There were no changes in the proportion of time spent in various sleep stages or in participant physical activity. These results show a clear modification of the physiological sleep requirements in healthy male volunteers who microdose LSD. The clear, clinically significant changes in objective measurements of sleep observed are difficult to explain as a placebo effect. Trial registration: Australian New Zealand Clinical Trials Registry: A randomized, double-blind, placebo-controlled trial of repeated microdoses of lysergic acid diethylamide (LSD) in healthy volunteers; https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=381476 ; ACTRN12621000436875.

Single administration of a psychedelic [(R)-DOI] influences coping strategies to an escapable social stress.

Psychedelic compounds have potentially rapid, long-lasting anxiolytic, antidepressive and anti-inflammatory effects. We investigated whether the psychedelic compound (R)-2,5-dimethoxy-4-iodoamphetamine [(R)-DOI], a selective 5-HT2A receptor partial agonist, decreases stress-related behavior in male mice exposed to repeated social aggression. Additionally, we explored the likelihood that these behavioral changes are related to anti-inflammatory properties of [(R)-DOI]. Animals were subjected to the Stress Alternatives Model (SAM), an escapable social stress paradigm in which animals develop reactive coping strategies - remaining in the SAM arena (Stay) with a social aggressor, or dynamically initiated stress coping strategies that involve utilizing the escape holes (Escape) to avoid aggression. Mice expressing these behavioral phenotypes display behaviors like those in other social aggression models that separate animals into stress-vulnerable (as for Stay) or stress-resilient (as for Escape) groups, which have been shown to have distinct inflammatory responses to social stress. These results show that Stay animals have heightened cytokine gene expression, and both Stay and Escape mice exhibit plasma and neural concentrations of the inflammatory cytokine tumor necrosis factor-α (TNFα) compared to unstressed control mice. Additionally, these results suggest that a single administration of (R)-DOI to Stay animals in low doses, can increase stress coping strategies such as increasing attention to the escape route, promoting escape behavior, and reducing freezing during socially aggressive interaction in the SAM. Lower single doses of (R)-DOI, in addition to shifting behavior to suggest anxiolytic effects, also concomitantly reduce plasma and limbic brain levels of the inflammatory cytokine TNFα.

Effects of discontinuation of serotonergic antidepressants prior to psilocybin therapy versus escitalopram for major depression.

BACKGROUND: There is growing evidence for the therapeutic effects of the psychedelic drug psilocybin for major depression. However, due to the lack of safety data on combining psilocybin with selective serotonin reuptake inhibitors (SSRIs) and serotonin-norepinephrine reuptake inhibitors (SNRIs) and concerns that there may be a negative interaction on efficacy, participants enrolling in psychedelic trials are usually required to discontinue SNRI/SNRIs prior to enrolling. AIMS: Using data from a recent clinical trial examining the comparative efficacy the psychedelic drug psilocybin (P) combined with approximately 20 h of psychological support to a 6-week (daily) course of the SSRI escitalopram plus matched psychological support for major depressive disorder, we explored the effects of discontinuing SSRI/SNRIs prior to study enrolment on study outcomes. METHODS: Exploratory post hoc analyses using linear mixed effects model were performed to investigate the discontinuation effect on various validated depression symptom severity scales and well-being. The impact of SSRI/SNRIs discontinuation on the acute psychedelic experience was also explored. RESULTS/OUTCOMES: In the psilocybin group, there was a reduced treatment effect on all outcome measures for SSRI/SNRIs discontinuers compared with unmedicated patients at trial entry. However, no effects of discontinuation on measures of the acute psychedelic experience were found. CONCLUSION: Discontinuation of SSRI/SNRIs before psilocybin might diminish response to treatment; however, as we did not test SSRI/SNRI continuation in our trial, we cannot infer such causation. Moreover, the exploratory nature of the analyses makes them hypothesis generating, and not confirmatory. A controlled trial of SSRI/SNRI discontinuation versus continuation prior to psilocybin is urgently required.

Chemistry and Pharmacology of Delta-8-Tetrahydrocannabinol.

Cannabis sativa is one of the oldest plants utilized by humans for both economic and medical purposes. Although the use of cannabis started millennia ago in the Eastern hemisphere, its use has moved and flourished in the Western nations in more recent centuries. C. sativa is the source of psychoactive cannabinoids that are consumed as recreational drugs worldwide. The C21 aromatic hydrocarbons are restricted in their natural occurrence to cannabis (with a few exceptions). Delta-9-tetrahydrocannabinol (Δ9-THC) is the main psychoactive component in cannabis, with many pharmacological effects and various approved medical applications. However, a wide range of side effects are associated with the use of Δ9-THC, limiting its medical use. In 1966, another psychoactive cannabinoid, Delta-8-tetrahydrocannabinol (Δ8-THC) was isolated from marijuana grown in Maryland but in very low yield. Δ8-THC is gaining increased popularity due to its better stability and easier synthetic manufacturing procedures compared to Δ9-THC. The passing of the U.S. Farm Bill in 2018 led to an increase in the sale of Δ8-THC in the United States. The marketed products contain Δ8-THC from synthetic sources. In this review, methods of extraction, purification, and structure elucidation of Δ8-THC will be presented. The issue of whether Δ8-THC is a natural compound or an artifact will be discussed, and the different strategies for its chemical synthesis will be presented. Δ8-THC of synthetic origin is expected to contain some impurities due to residual amounts of starting materials and reagents, as well as side products of the reactions. The various methods of analysis and detection of impurities present in the marketed products will be discussed. The pharmacological effects of Δ8-THC, including its interaction with CB1 and CB2 cannabinoid receptors in comparison with Δ9-THC, will be reviewed.

Psychedelic Therapy: A Primer for Primary Care Clinicians-Historical Perspective and Overview.

BACKGROUND: Psychedelic drugs have recently emerged as plausibly effective pharmacological agents for the management of depression, anxiety, and other neuropsychiatric conditions, including those that are treatment-resistent. The latter half of the 20th century marked a revolution in the treatment of mental illnesses, exemplified by the introduction of selective serotonin reuptake inhibitors and other pharmacological agents. Nevertheless, mental illness remains a major public health crisis, affecting nearly one billion individuals worldwide. AREAS OF UNCERTAINTY: Because of the decades-long status of several psychedelics as Schedule I drugs, there have not been very many large, double-blind, randomized controlled trials of psychedelics. Owing to small sample sizes, there may be rare yet serious adverse events that have not been reported in the clinical trials thus far. THERAPEUTIC ADVANCES: Esketamine, a dissociative hallucinogen drug, was approved for the management of major depressive disorder by the Food and Drug Administration in 2019. As of January 2024, two Phase III trials of 3,4-methylenedioxymethamphetamine (MDMA), a synthetic drug that inhibits the serotonin transporter, have been completed; the results indicate that MDMA is superior to existing pharmacological treatments for post-traumatic stress disorder. A phase III trial of psilocybin, a naturally occurring serotonin receptor partial agonist, is currently underway. The following series details the current state of research in psychedelic therapeutics, including lysergic acid diethylamide (LSD), N-N-dimethyltryptamine (DMT) and ayahuasca, psilocybin, ibogaine, MDMA, and ketamine. LIMITATIONS: While initial clinical trials of psychedelics for depression were very promising, trials of psilocybin with larger sample sizes (100+ participants) suggest that its remission rate is 25%-29%. This is about the same as the remission rate of antidepressants, which is roughly 30% according to the landmark STAR*D trial. CONCLUSIONS: Psychedelic drugs and structural derivatives offer a great deal of promise for the management of a wide range of psychiatric morbidities. It is imperative that clinicians become familiar with these novel agents and learn how to integrate psychedelic therapy with the rest of their care through open communication and referral.

Psychedelic Therapy: A Primer for Primary Care Clinicians-N,N-Dimethyltryptamine and Ayahuasca.

BACKGROUND: N,N-dimethyltryptamine (DMT) is a naturally occurring serotonergic psychedelic found in natural plants around the globe. As the main psychoactive component in ayahuasca, which also contains monoamine oxidase inhibitors, DMT has been consumed as plant-based brew by indigenous peoples for centuries. Further research is required to delineate the therapeutic utility of DMT. AREAS OF UNCERTAINTY: Although previous research has shown that DMT is synthesized endogenously, it may not be produced at physiologically relevant concentrations. Additionally, the phenomenological similarities between the DMT-induced state and near-death experiences led to the popular hypothesis that endogenous DMT is released during the dying process. However, this hypothesis continues to be debated. Generally, DMT and ayahuasca seem to be physiologically and psychiatrically safe, although ayahuasca is known to cause transient vomiting. THERAPEUTIC ADVANCES: A double-blind, randomized controlled trial showed that, within 1 week, ayahuasca causes remission in 36% of patients with treatment-resistant depression. According to top-line results from a recent phase IIa trial, 57% of patients with major depressive disorder experienced remission 12 weeks after receiving a single intravenous dose of DMT. LIMITATIONS: There has only been a single published double-blind randomized controlled trial on ayahuasca and 2 on DMT. All clinical trials have had small sample sizes (≤34 participants). DMT requires further research to understand its therapeutic and clinical potential as a psychedelic. CONCLUSIONS: Preliminary evidence indicates that ayahuasca and DMT may be more effective than existing antidepressants for treating major depressive disorder and treatment-resistant depression.

Psychedelic Therapy: A Primer for Primary Care Clinicians-The Strengths, Weaknesses, Opportunities, and Threats of Psychedelic Therapeutics.

The reviews in this special edition have presented a primer on the state of the literature for 7 different psychedelic compounds and their plausible roles in medicine. In a common format underscoring strengths, weakness, opportunities, and threats (SWOT), this article addresses how psychedelic compounds fit into the broader health care landscape for indicated conditions. Historically, psychiatric pathologies have been treated with small-molecule compounds that have limited effect sizes and carry a variety of adverse effect profiles. Psychedelic medicines offer the opportunity to provide more potent and rapidly acting treatments. It is crucial to note that this is an emerging field of medicine, and only one of these compounds (esketamine) is currently Food and Drug Administration-approved for depression. The other compounds discussed are investigational, and this discussion is both imaginative and prospective in nature.

Characterization of the Neurochemical and Behavioral Effects of the Phenethylamine 2-Cl-4,5-MDMA in Adolescent and Adult Male Rats.

BACKGROUND: The proliferation of novel psychoactive substances (NPS) in the drug market raises concerns about uncertainty on their pharmacological profile and the health hazard linked to their use. Within the category of synthetic stimulant NPS, the phenethylamine 2-Cl-4,5-methylenedioxymethamphetamine (2-Cl-4,5-MDMA) has been linked to severe intoxication requiring hospitalization. Thereby, the characterization of its pharmacological profile is urgently warranted. METHODS: By in vivo brain microdialysis in adolescent and adult male rats we investigated the effects of 2-Cl-4,5-MDMA on dopamine (DA) and serotonin (5-HT) neurotransmission in two brain areas critical for the motivational and rewarding properties of drugs, the nucleus accumbens (NAc) shell and the medial prefrontal cortex (mPFC). Moreover, we evaluated the locomotor and stereotyped activity induced by 2-Cl-4,5-MDMA and the emission of 50-kHz ultrasonic vocalizations (USVs) to characterize its affective properties. RESULTS: 2-Cl-4,5-MDMA increased dialysate DA and 5-HT in a dose-, brain area-, and age-dependent manner. Notably, 2-Cl-4,5-MDMA more markedly increased dialysate DA in the NAc shell and mPFC of adult than adolescent rats, while the opposite was observed on dialysate 5-HT in the NAc shell, with adolescent rats being more responsive. Furthermore, 2-Cl-4,5-MDMA stimulated locomotion and stereotyped activity in both adolescent and adult rats, although to a greater extent in adolescents. Finally, 2-Cl-4,5-MDMA did not stimulate the emission of 50-kHz USVs. CONCLUSIONS: This is the first pharmacological characterization of 2-Cl-4,5-MDMA demonstrating that its neurochemical and behavioral effects may differ between adolescence and adulthood. These preclinical data could help understanding the central effects of 2-Cl-4,5-MDMA by increasing awareness on possible health damage in users.