Synthetic cannabinoid CP-55,940 induces apoptosis in a human skeletal muscle model via regulation of CB1 receptors and L-type Ca2+ channels.

Rhabdomyolysis has been reported in patients who abuse synthetic cannabinoids. However, no studies have yet assessed whether these cases reflect the direct cytotoxicity of synthetic cannabinoids on skeletal muscle, a possibility that the present study sought to address. Specifically, this study investigated the cytotoxicity of the synthetic cannabinoid CP-55,940, a compound that acts equally on both types of cannabinoid receptors (CB1 and CB2), in a human embryonic rhabdomyosarcoma (RD) cell line. Exposure of these cells to CP-55,940 resulted in concentration-dependent decreases in cell viability. These effects were attenuated by pre-incubation with AM251 (30 µM), a selective CB1 receptor antagonist, but not by pre-incubation with AM630 (30 µM), a selective CB2 receptor antagonist. Following treatment with CP-55,940, RD cells exhibited apoptosis, as indicated by the accumulation of annexin-V, activation of caspase-3, and a loss of the mitochondrial membrane potential. Additionally, CP-55,940 treatment of RD cells led to increases in intracellular Ca2+ levels. CP-55,940-induced cell death was significantly attenuated in the absence of extracellular Ca2+, and was partially decreased by pre-incubation with verapamil (5 µM) or diltiazem (5 µM), compounds that block the L-type Ca2+ channel. Our results indicate that the cytotoxicity of CP-55,940 towards RD cells (skeletal muscle cells) is mediated by the CB1 receptor, but not by the CB2 receptor. Our results further suggest that calcium influx through the L-type channel may play an important role in the apoptosis induced by these compounds.

Synthetic cannabinoids enhanced ethanol-induced motor impairments through reduction of central glutamate neurotransmission.

Marijuana or synthetic cannabinoids and alcohol are often used together, with these combinations causing motor impairments that can subsequently lead to motor vehicle accidents. This study investigated the combined use of both synthetic cannabinoids and ethanol and their effect on motor coordination in mice in addition to examining the neurochemical changes in the cerebellum. Ethanol (2 g/kg, i.p.) significantly induced motor impairment in the accelerating rotarod test in mice. Furthermore, ethanol-induced motor impairments were further accentuated when combined with the synthetic cannabinoid, JWH-018 or AB-CHMINACA. The enhancement effects of the synthetic cannabinoids were completely antagonized by pretreatment with the selective CB1 receptor antagonist AM251, but not by the selective CB2 receptor antagonist AM630. Neurochemical study results showed that ethanol caused a reduction in the extracellular glutamate levels in the cerebellum during periods of ethanol-induced motor impairment. In addition to the enhanced motor impairment seen when ethanol was combined with JWH-018, these combinations also enhanced the reduction of the extracellular glutamate levels in the cerebellum. We additionally used microelectrode array recordings to examine the effects of ethanol and/or JWH-018 on the spontaneous network activity in primary cultures from mouse cerebellum. Results showed that ethanol combined with JWH-018 significantly reduced spontaneous neuronal network activity in the primary cerebellar culture. Our findings demonstrate that ethanol-induced motor impairments are enhanced by synthetic cannabinoids, with these effects potentially mediated by CB1 receptors. An accentuated reduction of neurotransmissions in the cerebellum may play an important role in motor impairments caused by ethanol combined with synthetic cannabinoids.

Synthetic cannabinoid AM2201 induces seizures: Involvement of cannabinoid CB1 receptors and glutamatergic transmission.

Abuse of synthetic cannabinoids is a serious social problem worldwide. Intentional ingestion of synthetic cannabinoids can cause severe toxicity, including seizures. Here we investigated the effects of acute administration of synthetic cannabinoids on the induction of epileptic seizures by monitoring electroencephalographic activity in freely moving mice. The synthetic cannabinoid, AM2201, induced abnormal, high-amplitude (>2-fold baseline amplitude), sharp-wave activity. The abnormal spike-wave discharges were accompanied by epileptiform behavior: rigid posture, tail extension, rearing with forepaws extended, jumping, and intermittent tonic-clonic jerking movements. The abnormal spike-wave discharges and behavioral changes were suppressed by pretreatment with the selective CB1 receptor antagonist AM251, but not with the selective CB2 receptor antagonist AM630 or the vanilloid receptor antagonist, capsazepine. Furthermore, the group 1 metabotropic glutamate receptor antagonist SIB1757 eliminated AM2201-induced spike-wave discharges and episodes of epileptiform behavior. AM2201 markedly increased the extracellular glutamate concentration in the hippocampus during periods of AM2201-induced abnormal spike-wave discharges and behavioral changes. These findings are the first evidence that AM2201 induces epileptic seizures by enhancing glutamatergic transmission in the hippocampus. Our findings demonstrate that induction of epileptic seizures by synthetic cannabinoids is mediated by CB1 receptors, but not by CB2 receptors, and further suggest that rapid elevation of glutamatergic transmission may play an important role in the induction of seizures following intentional ingestion of synthetic cannabinoids.

[Pharmacological properties of law-evading chemical substances].

In recent years, frequent cases of people suffering disturbed consciousness, dyspnea, etc. due to abuse of synthetic cannabis and being transported by ambulance or causing traffic accidents are occurring and are becoming a serious social problem in Japan. Most law-evading herbal products have colorful illustrations and logos and are sold as incense or herbs. Law-evading herbal products consist of finely chopped dry vegetative matter mixed with chemical substances (drugs), and the drugs are injurious to health. Analysis of chemical substances in herbal products clarified that they contain synthetic cannabinoid, a chemical component that exhibits action similar to that of hemp. There are many affinity compounds of cannabinoid, so presently, even if a particular drug is regulated, similar compounds that partially differ in structure will propagate. There is thus a cat-and-mouse game between regulations on chemical substances and their propagation. This paper summarizes the pharmacological actions and danger of chemical substances contained in law-evading herbal products by focusing on synthetic cannabinoid or synthetic cathinone, a chemical substance it contains.

Cytotoxicity of synthetic cannabinoids on primary neuronal cells of the forebrain: the involvement of cannabinoid CB1 receptors and apoptotic cell death.

The abuse of herbal products containing synthetic cannabinoids has become an issue of public concern. The purpose of this paper was to evaluate the acute cytotoxicity of synthetic cannabinoids on mouse brain neuronal cells. Cytotoxicity induced by synthetic cannabinoid (CP-55,940, CP-47,497, CP-47,497-C8, HU-210, JWH-018, JWH-210, AM-2201, and MAM-2201) was examined using forebrain neuronal cultures. These synthetic cannabinoids induced cytotoxicity in the forebrain cultures in a concentration-dependent manner. The cytotoxicity was suppressed by preincubation with the selective CB1 receptor antagonist AM251, but not with the selective CB2 receptor antagonist AM630. Furthermore, annexin-V-positive cells were found among the treated forebrain cells. Synthetic cannabinoid treatment induced the activation of caspase-3, and preincubation with a caspase-3 inhibitor significantly suppressed the cytotoxicity. These synthetic cannabinoids induced apoptosis through a caspase-3-dependent mechanism in the forebrain cultures. Our results indicate that the cytotoxicity of synthetic cannabinoids towards primary neuronal cells is mediated by the CB1 receptor, but not by the CB2 receptor, and further suggest that caspase cascades may play an important role in the apoptosis induced by these synthetic cannabinoids. In conclusion, excessive synthetic cannabinoid abuse may present a serious acute health concern due to neuronal damage or deficits in the brain.

Dopaminergic hyperactivity accompanied by hyperlocomotion in C57BL/6J-bg(J)/bg(J) (beige-J) mice.

Genetic factors affect locomotor activity, which mainly depends on the activation of dopaminergic systems. C57BL/6J-bg(J)bg(J) (beige-J) mice, which exhibit deficiencies in immunological function, show behavioral hyperactivity. The present study was designed to investigate the locomotor activity of beige-J mice accompanied by a change in the dopaminergic system. Beige-J mice showed higher locomotor activity and dopamine turnover, whereas splenectomy reduced this hyperlocomotion and dopamine turnover. These results suggest that beige-J mice could be suitable as an experimental animal model for investigating hyperactivation of the dopaminergic system, and the spleen may contribute to the susceptibility of dopaminergic systems to activation.

Fentanyl-Type Antagonist of the µ-Opioid Receptor: Important Role of Axial Chirality in the Active Conformation.

In recent years, synthetic opioids have emerged as a predominant cause of drug-overdose-related fatalities, causing the "opioid crisis." To design safer therapeutic agents, we accidentally discovered µ-opioid receptor (MOR) antagonists based on fentanyl with a relatively uncomplicated chemical composition that potentiates structural modifications. Here, we showed the development of novel atropisomeric fentanyl analogues that exhibit more potent antagonistic activity against MOR than naloxone, a morphinan MOR antagonist. Derivatives displaying stable axial chirality were synthesized based on the amide structure of fentanyl. The aS- and aR-enantiomers exerted antagonistic and agonistic effects on the MOR, respectively, and each atropisomer interacted with the MOR by assuming a distinct binding mode through molecular docking. These findings suggest that introducing atropisomerism into fentanyl may serve as a key feature in the molecular design of future MOR antagonists to help mitigate the opioid crisis.

Ecstasy (3, 4-methylenedioxymethamphetamine) use among Japanese rave population.

AIMS: The aim of this study was to examine the prevalence of 3, 4-methylenedioxymethamphetamine (MDMA) use and to identify characteristics of MDMA users among rave attendees in Japan. This is the first rave-population study focusing on MDMA use in Japan. METHODS: The anonymous self-administrative questionnaire was conducted using laptop computers at four rave parties at three different venues in Tokyo, Japan. Participants were asked about lifetime use of MDMA and other club drug use, characteristics of rave attendance, and their demographics. RESULTS: Questionnaires were completed by 300 rave attendees (47.3% female), 68.3% of whom were aged 20-29 years, and 92.3% of whom had completed high school. Among the participants, 8.0% reported MDMA use. Compared with 'non-drug controls' (the participants who had never used any illicit drugs), MDMA users were significantly more likely to be 30-39-year-old men. In addition, compared with 'cannabis users' (non-MDMA users who had used cannabis), MDMA users were significantly more likely to use other drugs and reported more adverse health effects due to 'polydrug use'. Furthermore, MDMA users were significantly more likely to go to raves and preferred smaller venues. CONCLUSIONS: Our results clearly suggest that rave attendees have a higher lifetime prevalence of MDMA use than the Japanese general population (0.2% reported in 2007). MDMA users are deeply involved in rave parties, and MDMA use may have high potential to generate close relationships among rave attendees. Therefore, MDMA users may have more opportunities to access MDMA than cannabis users and non-drug controls.

[Drug discrimination properties and cytotoxicity of the cannabinoid receptor ligands].

The worldwide distribution of smokable herbal mixtures called "Spice" that contain synthetic cannabinoids with a pharmacological activity similar to delta 9-tetrahydrocannabinol (delta 9-THC) has been reported. The synthetic cannabinoids induce behavior and have biochemical properties similar to naturally occurring cannabinoids such as delta 9-THC. In drug discrimination procedures, animal behavior is differentially reinforced depending on the presence or absence of specific drug stimuli. This review seeks to establish an animal model to serve as a discriminative stimulus of the synthetic cannabinoids, to determine whether this discriminative stimulus is identical to that of delta 9-THC. Much data have been obtained in drug discrimination experiments with various synthetic cannabinoids. In the discriminative study, synthetic cannabinoids such as CP-55,940 and WIN-55,212-2 were substituted for delta 9-THC in rats trained to discriminate delta 9-THC from the vehicle. These discriminative effects of synthetic cannabinoids were antagonized by CB1 antagonist SR-141,716A. The discriminative effects of synthetic cannabinoids may overlap with the delta 9-THC cue mediated by CB1 receptors. In in vitro study using NG 108-15 cell lines, synthetic cannabinoids have produced strong cytotoxicities that were suppressed by pretreatment with the CB1 receptor antagonist. Furthermore, pretreatment with caspase inhibitors suppressed these synthetic-cannabinoid-induced cytotoxicities in NG 108-15 cells. These findings indicate that the cytotoxicity of synthetic cannabinoids towards NG 108-15 cells is mediated by the CB1 receptors and further suggest that caspase cascades may play an important role in the cytotoxicities induced by these synthetic cannabinoids. In conclusion, synthetic cannabinoid abuse could be a health hazard for humans.

[Pharmacological properties and dependence liabilities of synthetic cannabinoids].

Since 2000, types of abused drugs have increased in Japan and abused drugs are diversifying. Chemicals with structures similar to the drugs and stimulants restricted by law are sold as medicines that are not so restricted. Furthermore, drugs that have evaded attention of the law and abuse of these drugs are increasing rapidly. Serious health hazards are occurring from abuse of these chemicals as well as existing abused drugs: they are thus causing a major social problem. Synthetic cannabinoids, one of these illegal drugs, is called "spice" and its abuse is spreading all over the world. So far, Germany and Japan have analyzed the ingredients of the spice series. Synthetic cannabinoids that have actually been detected include CP-47497 and its variations with different numbers of carbon atoms in the side chain, i.e., CP-47497-(C8), JWH-018, etc. Cases of drug dependence and health hazards, such as mental diseases, caused by abuse of spice have been reported, revealing their danger. Numerous synthetic cannabinoid derivatives emerged during the development of medicines, and the methods for synthesizing them and the strength of their binding affinity with the cannabinoid receptor have also been clarified. Under current conditions, when particular synthetic cannabinoid derivatives are restricted, others soon replace them. Although the biochemical characteristics of synthetic cannabinoid derivatives, such as binding affinity with receptors, have been clarified, actions of synthetic cannabinoid derivatives that are taken are still unknown for most derivatives. Some cannabinoid derivatives exhibit tens of times the strength of delta 9-THC, an ingredient of hemp, so it is feared that taking them may cause unexpected serious health hazards.

[Animal models for evaluating of drug dependence liability].

Assessment of psychological dependence liability of drug is essential for regulatory compliance and the drug development process. The most reliable method for evaluating the psychic dependence liability of drug is the self-administration paradigm. The conditioned place preference paradigm is also useful method for evaluating the rewarding effect of drugs and is widely used. Furthermore, the drug discrimination paradigm, which analyzes the similarity with abused drugs by utilizing the effect of consciousness when taking drugs, has also been implemented. This paper summarizes the characteristics of each method and outlines the methods for evaluating drug dependence using animals. It also introduces recent findings about the studies of molecular mechanisms of drug dependence using gene-knockout and transgenic mice.

[Present Conditions of Marijuana Regulation in USA: Medical and Recreational Use].

In most countries marijuana is regulated by the Single Convention on Narcotic Drugs. In Japan marijuana use is illegal under the Marijuana Control Law. In USA, marijuana is also classified as a schedule I drug, which is the most stringent regulation category under federal law. On the other hand, California became the first state to legalize marijuana for medical uses in 1996. Since then, several other US states have approved marijuana for medical or recreational use. However, marijuana remains completely illegal in most states, while some allow only cannabidiol (CBD) extracted from marijuana for medical use. In June 2018, the US Food and Drug Administration approved Epidiolex, the first marijuana-derived drug, containing purified CBD, to treat certain rare childhood seizure syndromes. Therefore the situation surrounding control of marijuana in USA is complex. Recently, a definite trend toward reconsidering marijuana regulation has been seen around the world, which could have a major impact on marijuana policy in Japan. In this review, we investigated existing medical and recreational marijuana laws in various US states, with a focus on California, which approved recreational use in 2018. Here, we describe the current state of marijuana regulation in terms of both medical and recreational use. In addition, we discuss public safety issues associated with marijuana, including crime, traffic accidents, and emergency department visits from possible marijuana exposure, as well as generated tax revenues, from official marijuana-related reports in Colorado, which legalized marijuana use in 2012.

[Dependence and Cytotoxicity of Components of Cannabis].

Cannabis use among the younger population in Japan has been steadily increasing. The aim of the present review is to highlight recent knowledge regarding the molecular mechanisms of action and health risks associated with cannabis and synthetic cannabinoid consumption. We investigated the effects of Δ9-tetrahydrocannabinol (THC) and synthetic cannabinoids on place conditioning in ICR mice. Both Δ9-THC and synthetic cannabinoids produce a significant conditioned place preference. These rewarding effects were completely suppressed by the cannabinoid CB1 receptor type antagonist AM251. The cytotoxicological effects of Δ9-THC and synthetic cannabinoids were also characterized in the limbic forebrain of mice in primary culture in vitro. Δ9-THC and synthetic cannabinoids caused cell death in a dose-dependent manner. The rank order of cytotoxicological potency was synthetic cannabinoids>Δ9-THC and related to the agonistic activities of the CB1 receptor. A recent review on the harmful effects of cannabis use in humans reported that behavioral impairments, especially in terms of attention, memory, and complex information-processing ability, can last for many weeks after cessation of cannabis use among heavy users. In addition, cannabis use could be a risk factor for drug dependence and later psychosis among adolescents. The results of animal and human studies suggest that CB1 receptors play an important role in the expression of harmful effects of cannabis and synthetic cannabinoid use. Moreover, concern regarding increasing concentrations of Δ9-THC in cannabis in many countries has been noted, because more potent cannabis may be associated with worse adverse effects.

Δ9-Tetrahydrocannabinol elicited 22-kHz ultrasonic vocalization changes after air puff stimulus through CB1 receptor in adult rats.

Δ9-Tetrahydrocannabinol (THC) is known to have various pharmacological effects mediated through activation of cannabinoid CB1 and CB2 receptors in rodents. In adult rats, 22- and 50-kHz ultrasonic vocalizations (USVs) serve as an effective communication system and as indicators of negative and positive states, respectively. The present study was performed to determine whether THC affects USVs in adult rats, and to determine the roles of cannabinoid receptors in these effects. THC (1, 3 mg/kg) was administered intraperitoneally to adult male Wistar rats 60 min before measurement of USVs. The CB1 antagonist, SR141716 (3, 6 mg/kg), or CB2 antagonist, AM630 (1, 10 mg/kg), was administered intraperitoneally 10 min before THC. USVs were measured during a 5-minute period without air puff stimulus or with air puff stimulus. THC did not affect 22- or 50-kHz USVs without air puff stimulus. On the other hand, THC significantly increased the number of 22-kHz USVs, but not 50-kHz USVs, after air puff stimulus. Moreover, SR141716 at 6 mg/kg, but not AM630 at either dose, inhibited the increase in number of 22-kHz USVs induced by THC after air puff stimulus. These results suggest that THC induced changes in sensitivity to aversive air puff stimuli through CB1 receptors, and as a result increased emission of 22-kHz USVs in rats.

[Evaluation of the rewarding effects of drugs by conditioned place preference (CPP) paradigm: properties of volatile organic solvents and uncontrolled newly-abused drugs].

The conditioned place preference paradigm has been used to evaluate the motivational effects of drugs. The primary motivational effects of a drug are conditioned to prominent environmental stimuli of a conditioned place preference box, and after several conditionings, animals acquire secondary motivational effects of drugs. If the drug experience produces a positive effect, it is expected that animals will spend more time in the drug-paired place. This technique only requires that the animals carry out a simple operation to approach or avoid the drug-paired place. Place conditioning procedures have been used to assess rewarding and aversive properties of drugs. Abuse of volatile organic solvents among young people is a serious social issue in Japan. Organic solvents are cheap and relatively easy to obtain; hence, they carry the risk of becoming "gate-way drugs" for users. We developed an airtight conditioned place preference system for drug inhalation. Using this system, we found that toluene inhalation produced a rewarding effect in mice. The conditioned place preference paradigm has been used as a tool to delineate the neurochemical substrates mediating the rewarding effect of drugs. This method may be of great help in evaluating the abuse potential of substances (e.g., volatile organic solvents and uncontrolled newly-abused drugs) in animals.

[Neurochemical mechanisms for development of psychological dependence on volatile organic solvents].

Abuse of volatile organic solvents among youth remains a major social problem. Organic solvents are cheap and relatively easy to obtain, so they carry the risk of becoming a "gateway drug" for users. The effect of repeated inhalation of toluene on subsequent responses to other drugs of abuse is unclear. In the present study, we investigated the effect of toluene inhalation on methamphetamine-induced behavioral change using a newly developed sealed inhalation shuttlebox. The influence of the cyclic AMP response element binding (CREB) protein expression following toluene inhalation was also examined. Mice were exposed to toluene or air once daily for five days. Methamphetamine produced significant hyperlocomotion in air-exposed mice. This stimulatory effect of methamphetamine was significantly enhanced following repeated inhalation of toluene. Furthermore, repeated toluene inhalation increased the levels of CREB proteins in the limbic forebrain. The present study demonstrated that adaptation of the adenylate cyclase system following repeated toluene inhalation might be involved in the expression of behavioral sensitization to subsequent methamphetamine administration. Inhalant abuse could thus be associated with the risk of other substances of abuse.

Pyrolysis of UR-144, a synthetic cannabinoid, augments an affinity to human CB1 receptor and cannabimimetic effects in mice.

Drug abusers most often smoke 'herbal incense' as a cigarette or inhale it using a smoking tool. Smoking may cause pyrolysis of the drug and produce decomposed products of which biological effect has never been investigated. The synthetic cannabinoid UR-144 is known to undergo thermal degradation, giving a ring-opened isomer, so-called UR-144 degradant. The present study demonstrates by using UR-144 as a model drug that the smoke of burned UR-144 contains the UR-144 degradant. The UR-144 degradant showed approximately four fold higher agonist activity to human CB1 receptor and augmented hypothermic and akinetic actions in mice compared to UR-144. These results indicate that smoking behavior may increase psychological actions of the certain synthetic cannabinoids.

Oxidative metabolism of 5-methoxy-N,N-diisopropyltryptamine (Foxy) by human liver microsomes and recombinant cytochrome P450 enzymes.

In vitro quantitative studies of the oxidative metabolism of (5-methoxy-N,N-diisopropyltryptamine, 5-MeO-DIPT, Foxy) were performed using human liver microsomal fractions and recombinant CYP enzymes and synthetic 5-MeO-DIPT metabolites. 5-MeO-DIPT was mainly oxidized to O-demethylated (5-OH-DIPT) and N-deisopropylated (5-MeO-IPT) metabolites in pooled human liver microsomes. In kinetic studies, 5-MeO-DIPT O-demethylation showed monophasic kinetics, whereas its N-deisopropylation showed triphasic kinetics. Among six recombinant CYP enzymes (CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A4) expressed in yeast or insect cells, only CYP2D6 exhibited 5-MeO-DIPT O-demethylase activity, while CYP1A2, CYP2C8, CYP2C9, CYP2C19 and CYP3A4 showed 5-MeO-DIPT N-deisopropylase activities. The apparent Km value of CYP2D6 was close to that for 5-MeO-DIPT O-demethylation, and the Km values of other CYP enzymes were similar to those of the low-Km (CYP2C19), intermediate-Km (CYP1A2, CYP2C8 and CYP3A4) and high-Km phases (CYP2C9), respectively, for N-deisopropylation in human liver microsomes. In inhibition studies, quinidine (1 microM), an inhibitor of CYP2D6, almost completely inhibited human liver microsomal 5-MeO-DIPT O-demethylation at a substrate concentration of 10 microM. Furafylline, a CYP1A2 inhibitor, quercetin, a CYP2C8 inhibitor, sulfaphenazole, a CYP2C9 inhibitor and ketoconazole, a CYP3A4 inihibitor (5 microM each) suppressed about 60%, 45%, 15% and 40%, respectively, of 5-MeO-DIPT N-deisopropylation at 50 microM substrate. In contrast, omeprazole (10 microM), a CYP2C19 inhibitor, suppressed only 10% of N-deisopropylation by human liver microsomes, whereas at the same concentration the inhibitor suppressed the reaction by recombinant CYP2C19 almost completely. These results indicate that CYP2D6 is the major 5-MeO-DIPT O-demethylase, and CYP1A2, CYP2C8 and CYP3A4 are the major 5-MeO-DIPT N-deisopropylase enzymes in the human liver.

[Role of the monoamine system in the brain on the development of psychological dependence on toluene].

Abuse of volatile organic solvents among youth remains a major social problem in Japan. Organic solvents are cheap and relatively easy to obtain, so they carry the risk of becoming a so-called "gate-way drug" for users. Psychological dependence assessment systems have been established for drug inhalation using the conditioned place preference (CPP) procedure. We found toluene produced the rewarding effect in this new CPP system. The mesolimbic dopamine pathway, which includes dopaminergic neurons in the ventral tegmental area (VTA) of the midbrain and their targets in the limbic forebrain, especially the nucleus accumbens (NAC), is one of the most important substrates for the development of psychological dependence on drugs such as stimulants, cocaine, and heroin. Recently, it has indicated that the VTA-NAC pathway (monoamine system) may play an important role of the expression of psychological dependence on the volatile organic solvent toluene. Clarification of organic solvent's mechanism for the development of psychological dependence focusing on the monoamine system can be exploited for the new medicine and useful treatments for dependence on toluene.