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Cannabis-based treatment induces polarity-reversing plasticity assessed by theta burst stimulation in humans.

In animal models, the cannabinoid system has been convincingly implicated in the regulation of long-lasting synaptic plasticity. Both long-term potentiation (LTP) and depression (LTD) phenomena can be induced in the human motor cortex by transcranial magnetic theta burst stimulation (TBS). Here, we explored the potential involvement of the cannabinoid system in TBS-induced synaptic plasticity in humans. We tested the effects of a cannabis-based preparation (Sativex) on continuous TBS (cTBS) and intermittent TBS (iTBS) protocols in subjects with multiple sclerosis. We observed a shift in the polarity of synaptic plasticity induced by cTBS. In these subjects, in fact, cTBS induced the expected inhibition of motor-evoked potentials (MEPs) before Sativex exposure, whereas it caused a persisting enhancement of MEP amplitude 4 weeks after. The LTP-like phenomenon induced by iTBS was conversely unaffected by Sativex. Our results indicate that cannabis ingredients have metaplastic effects on the motor cortex, and strongly suggest that the cannabinoid system is involved in the modulation of synaptic plasticity not only in rodents but also in humans.

Neuroprotective strategies in drug abuse-evoked encephalopathy.

Encephalopathy is evidenced as an altered mental state with various neurological symptoms, such as memory and cognitive problems. The type of a substance-evoked encephalopathy will depend on the drug, substance, or combination being abused. The categories into which we could place the various abused substances could be tentatively divided into stimulants, amphetamines, hallucinogens, narcotics, inhalants, anesthetics, anabolic steroids, and antipsychoticsantidepressants. Other factors that may underlie encephalopathy, such as infectious agents, environmental, and other factors have also to be taken into account. Drugs of abuse can be highly toxic to the CNS following acute, but more so in chronic exposure, and can produce significant damage to other organs, such as the heart, lungs, liver, and kidneys. The damage to these organs may be at least partially reversible when drug abuse is stopped but CNS damage from repeated or prolonged abuse is often irreversible. The major pathways for the organ and CNS toxicity could be related to ischemic events as well as increased cell damage due to metabolic or mitochondrial dysfunction resulting in increased excitotoxicity, reduced energy production, and lowered antioxidant potential. These susceptibilities could be strengthened by the use of antioxidants to combat free radicals (e.g., vitamin E, lipoic acid) trying to improve energy generation by using mitochondriotropicmetabolic compounds (e.g., thiamine, coenzyme Q10, carnitine, riboflavin) by reducing excitotoxicity (e.g., glutamate antagonists) and other possible strategies, such as robust gene response, need to be investigated further. The drug-abuse-evoked encephalopathy still needs to be studied further to enable better preventative and protective strategies.

Chronic adolescent exposure to Δ-9-tetrahydrocannabinol in COMT mutant mice impact on psychosis-related and other phenotypes.

Cannabis use confers a two-fold increase in the risk for psychosis, with adolescent use conferring even greater risk. A high-low activity catechol-O-methyltransferase (COMT) polymorphism may modulate the effects of adolescent Δ-9-tetrahydrocannabinol (THC) exposure on the risk for adult psychosis. Mice with knockout of the COMT gene were treated chronically with THC (4.0 and 8.0 mgkg over 20 days) during either adolescence (postnatal days (PDs) 32-52) or adulthood (PDs 70-90). The effects of THC exposure were then assessed in adulthood across behavioral phenotypes relevant for psychosis exploratory activity, spatial working memory (spontaneous and delayed alternation), object recognition memory, social interaction (sociability and social novelty preference), and anxiety (elevated plus maze). Adolescent THC administration induced a larger increase in exploratory activity, greater impairment in spatial working memory, and a stronger anti-anxiety effect in COMT knockouts than in wild types, primarily among males. No such effects of selective adolescent THC administration were evident for other behaviors. Both object recognition memory and social novelty preference were disrupted by either adolescent or adult THC administration, independent of genotype. The COMT genotype exerts specific modulation of responsivity to chronic THC administration during adolescence in terms of exploratory activity, spatial working memory, and anxiety. These findings illuminate the interaction between genes and adverse environmental exposures over a particular stage of development in the expression of the psychosis phenotype.

Activation of α7 nicotinic receptors improves phencyclidine-induced deficits in cognitive tasks in rats implications for therapy of cognitive dysfunction in schizophrenia.

Nicotinic α7 acetylcholine receptors (nAChRs) have been highlighted as a target for cognitive enhancement in schizophrenia. To investigate whether the deficits induced by sub-chronic phencyclidine (PCP) in reversal learning and novel object recognition could be attenuated by the selective α7 nAChR full agonist, PNU-282987. Adult female hooded-Lister rats received sub-chronic PCP (2mgkg) or vehicle i.p. twice daily for 7days, followed by 7 days washout. In cohort 1, PCP-treated rats then received PNU-282987 (5, 10, 20mgkg s.c.) or vehicle and were tested in the reversal-learning task. In cohort 2, PCP-treated rats received PNU-282987 (10mgkg s.c.) or saline for 15days and were tested in the novel object recognition test on day 1 and on day 15, to test for tolerance. Sub-chronic PCP produced significant deficits in both cognitive tasks (P<0.01-0.001). PNU-282987 attenuated the PCP-induced deficits in reversal learning at 10mgkg (P<0.01) and 20mgkg (P<0.001), and in novel object recognition at 10mgkg on day 1 (P<0.01) and on day 15 (P<0.001). These data show that PNU-282987 has efficacy to reverse PCP-induced deficits in two paradigms of relevance to schizophrenia. Results further suggest that 15-day once daily dosing of PNU-282987 (10mgkg s.c.) does not cause tolerance in the rat. This study suggests that activation of α7 nAChRs, may represent a suitable strategy for improving cognitive deficits of relevance to schizophrenia.

A structural insight into the molecular recognition of a (-)-Delta9-tetrahydrocannabinol and the development of a sensitive, one-step, homogeneous immunocomplex-based assay for its detection.

(-)-Delta9-tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC-bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 A and 2.0 A resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C(10) monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C(10) monoterpene moiety, 11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol and 11-hydroxy-(9)-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab-Delta(9)-THC immunocomplex binding antibody from a naïve antibody phage display library, the specificity of the Delta(9)-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ngml from saliva samples.

Transfer of the discriminative stimulus effects of Δ9-THC and nicotine from one operant response to another in rats.

Transfer of the discriminative stimulus effects of two drugs from one operant (original-response) to a topographically different response (transfer-response) that was spared drug discrimination training was investigated. Eight rats were trained in a counterbalanced one manipulandum (lever press and nose poke) drug discrimination procedure. Counterbalanced IP administered nicotine (0.3 mgkg) or Δ(9)-tetrahydrocannabinol (3.0 mgkg) functioned as discriminative stimuli. S(D) drugs occasioned sessions of food-reinforcement (variable-interval 30-s schedule) S(Δ) drugs occasioned non-reinforcement. The original-response (lever-pressing or nose-poking) was initially reinforced during 30-min S(D) drug sessions, and non-reinforced on the other alternating S(Δ)-drug sessions. Two separate 5-min non-reinforcement tests, counterbalanced by drug order, revealed stimulus control over the original-response by both drugs, which transferred to the transfer-response. Subsequent extinction training of the transfer-response attenuated the original-response response rates with the S(D) drug conditions but had little impact on discriminative control. Discriminative control was reversed for the transfer-response but had little impact on the original-response but, again, reduced response rate. These data demonstrate that (a) discriminative control by two distinct drug states can transfer and modulate a topographically different free-operant response and, (b) as is true for exteroceptive stimuli, drug states that function as antecedents embedded within the operant three-term contingency have differing relationships with the response and the primary reinforcer.

LC-MS-MS method for simultaneous determination of THCCOOH and THCCOOH-glucuronide in urine Application to workplace confirmation tests.

Cannabis is the one of the most frequently detected drug in workplace drug testing and in cases of driving under influence of drugs, and there is a greater demand for sensitive, rapid and reliable methods for confirming the presence of this drug in biological samples. This paper describes an LC-MS-MS procedure for direct analysis of cTHC and cTHC-glucuronide in urine, without hydrolysis of the samples or derivatisation. The sample preparation is very simple and consist only in a dilution of urine with methanol 11 (vv) after adding the deuterated internal standard (cTHC-d3) then 10μl of the final solution is injected in LC-MS-MS. Chromatographic separation was performed using a reversed-phase column and gradient elution with 0.01% formic acidacetonitrilemethanol and two MS-MS transitions for each substance were monitored. The method was fully validated and proved to be accurate (RSD <15%), precise (intra-day CV <10%) and sensitive with a limit of detection (LOD) of 5ngml for both the compounds studied. Linearity was tested in the range 5-1000ngml and the values of R(2) resulted >0.99. The developed method was applied to several authentic samples of urine which tested positive in the immunoassay screening and 98% of them was confirmed.

Use patterns and self-reported effects of Salvia divinorum an internet-based survey.

There is growing use of Salvia divinorum (SD), a psychoactive plant that produces hallucinogen-like effects through a kappa opioid receptor (KOR) mechanism. Little is known about KOR agonist effects in humans and about users of SD. To characterize the reasons, methods, and reported consequences of SD use. Individuals reading SD-related pages of a drug-information website were invited to anonymously complete an online questionnaire if they had used SD. Participants (N500) were 92.6% male and 23.4 ± 8.7 (mean ± s.d.) years old. They had used a median of six times (range 1-250). 80.6% probably or definitely would use SD again. Most participants (92.6%) typically smoked or vaporized SD product. When smoked, the drugs main effects were estimated to last 14.1 ± 12.8 (range 0.5-120) minutes. When asked to compare SD effects to other methods of altering consciousness, the most common answer was that SD was unique (38.4%). 25.8% reported persisting (≥ 24 h) positive effects (often described as increased sense of well-being) on at least one occasion. 4.4% reported persisting negative effects (most often anxiety). SD is typically smoked, acute effects are brief, and persistent adverse effects are uncommon. In addition to acute hallucinogenic effects, SD may produce subacute increases in subjective well-being. Such a subacute effect would be unusual for a drug that is used non-medically, as withdrawal from other drugs typically either does not affect mood or causes dysphoria. Findings from this convenience sample should be confirmed and extended using surveys of random samples and controlled clinical studies.

Adolescent brain maturation, the endogenous cannabinoid system and the neurobiology of cannabis-induced schizophrenia.

Cannabis use during adolescence increases the risk of developing psychotic disorders later in life. However, the neurobiological processes underlying this relationship are unknown. This review reports the results of a literature search comprising various neurobiological disciplines, ultimately converging into a model that might explain the neurobiology of cannabis-induced schizophrenia. The article briefly reviews current insights into brain development during adolescence. In particular, the role of the excitatory neurotransmitter glutamate in experience-dependent maturation of specific cortical circuitries is examined. The review also covers recent hypotheses regarding disturbances in strengthening and pruning of synaptic connections in the prefrontal cortex, and the link with latent psychotic disorders. In the present model, cannabis-induced schizophrenia is considered to be a distortion of normal late postnatal brain maturation. Distortion of glutamatergic transmission during critical periods may disturb prefrontal neurocircuitry in specific brain areas. Our model postulates that adolescent exposure to Δ9-tetrahydrocannabinol (THC), the primary psychoactive substance in cannabis, transiently disturbs physiological control of the endogenous cannabinoid system over glutamate and GABA release. As a result, THC may adversely affect adolescent experience-dependent maturation of neural circuitries within prefrontal cortical areas. Depending on dose, exact time window and duration of exposure, this may ultimately lead to the development of psychosis or schizophrenia. The proposed model provides testable hypotheses which can be addressed in future studies, including animal experiments, reanalysis of existing epidemiological data, and prospective epidemiological studies in which the role of the dose-time-effect relationship should be central.

Longitudinal study of cognition among adolescent marijuana users over three weeks of abstinence.

Cognitive deficits that persist up to a month have been detected among adult marijuana users, but decrements and their pattern of recovery are less known in adolescent users. Previously, we reported cognitive deficits among adolescent marijuana users after one month of abstinence (Medina, Hanson, Schweinsburg, Cohen-Zion, Nagel, Tapert, 2007). In this longitudinal study, we characterized neurocognitive changes among marijuana-using adolescents across the first three weeks of abstinence. Participants were adolescent marijuana users with limited alcohol and other drug use (n19) and demographically similar non-using controls (n21) ages 15-19. Participants completed a brief neuropsychological battery on three occasions, after 3days, 2weeks, and 3weeks of stopping substance use. Abstinence was ascertained by decreasing tetrahydrocannabinol metabolite values on serial urine drug screens. Verbal learning, verbal working memory, attention and vigilance, and time estimation were evaluated. Marijuana users demonstrated poorer verbal learning (p<.01), verbal working memory (p<.05), and attention accuracy (p<.01) compared to controls. Improvements in users were seen on word list learning after 2weeks of abstinence and on verbal working memory after 3weeks. While attention processing speed was similar between groups, attention accuracy remained deficient in users throughout the 3-week abstinence period. This preliminary study detected poorer verbal learning and verbal working memory among adolescent marijuana users that improved during three weeks of abstinence, while attention deficits persisted. These results implicate possible hippocampal, subcortical, and prefrontal cortex abnormalities.

Pharmacological evaluation of the natural constituent of Cannabis sativa, cannabichromene and its modulation by Δ(9)-tetrahydrocannabinol.

In contrast to the numerous reports on the pharmacological effects of Δ(9)-tetrahydrocannabinol (THC), the pharmacological activity of another substituent of Cannabis sativa, cannabichromene (CBC) remains comparatively unknown. In the present study, we investigated whether CBC elicits cannabinoid activity in the tetrad assay, which consists of the following four endpoints hypomotility, antinociception, catalepsy, and hypothermia. Because cannabinoids are well documented to possess anti-inflammatory properties, we examined CBC, THC, and combination of both phytocannabinoids in the lipopolysaccharide (LPS) paw edema assay. CBC elicited activity in the tetrad that was not blocked by the CB(1) receptor antagonist, rimonabant. Moreover, a behaviorally inactive dose of THC augmented the effects of CBC in the tetrad that was associated with an increase in THC brain concentrations. Both CBC and THC elicited dose-dependent anti-inflammatory effects in the LPS-induced paw edema model. The CB(2) receptor, SR144528 blocked the anti-edematous actions of THC, but not those produced by CBC. Isobolographic analysis revealed that the anti-edematous effects of these cannabinoids in combination were additive. Although CBC produced pharmacological effects, unlike THC, its underlying mechanism of action did not involve CB(1) or CB(2) receptors. In addition, there was evidence of a possible pharmacokinetic component in which CBC dose-dependently increased THC brain levels following an i.v. injection of 0.3mgkg THC. In conclusion, CBC produced a subset of behavioral activity in the tetrad assay and reduced LPS-induced paw edema through a noncannabinoid receptor mechanism of action. These effects were augmented when CBC and THC were co-administered.

Serotoninergic modulation of cortical and respiratory responses to episodic hypoxia.

Biphasic respiratory response to hypoxia in anesthetized animals is accompanied by changes in the EEG mostly in the low EEG frequency bands. Serotonin is a potent modulator of cortical and respiratory activity through 5-HT(2) receptors. Present study investigated whether 5-HT(2) receptors might be involved in the EEG and respiratory relationship during normoxic and hypoxic respiration assessed from integrated phrenic (Phr) and hypoglossal (HG) nerve activities. EEG signal recorded from the frontal cortex was subjected to power spectral analysis in delta, theta, alpha, and beta frequency bands. Systemic administration of 5-HT(2) agonist DOI (1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane) enhanced tonic and lowered peak phasic respiratory activity, and increased frequency of bursts of Phr and HG activity. At the same time, EEG activity became desynchronized and arterial blood pressure (ABP) increased. Following DOI pretreatment, 11% hypoxia induced an augmented respiratory response in comparison with the response in the baseline condition. ABP fell less then in the control hypoxia. EEG pattern changed less than in the baseline state. Subsequent administration of ketanserin, a 5-HT(2) antagonist increased respiratory activity, elicited a synchronization of EEG activity and hypotension. The respiratory response to hypoxia was attenuated and cortical response was more potent in comparison with that after DOI injection. Arterial blood pressure decreased more then during baseline hypoxic response. The results suggest that modulation of cortical synchronization and desynchronization through 5-HT(2) receptor active agents may impact to hypoxic respiratory response.

Neurotoxic effects of MDMA (ecstasy) on the developing rodent brain.

The incidence of methamphetamine abuse is particularly high in adolescents and is a common problem among women of childbearing age, leading to an increasing number of children with prenatal exposure. MDMA (3,4-methylenedioxymethamphetamine, ecstasy) is an amphetamine-like stimulant and is known to induce apoptotic damage to fine serotonergic fibers in the adult rat brain. Little is known about toxic effects of MDMA and potential underlying molecular mechanisms in the developing brain. Here, we investigated whether MDMA exposure during the period of rapid brain growth causes neurodegeneration in the developing rat brain. MDMA significantly enhanced neuronal death in the brains of 6-day-old rat pups at a dose of 60 mgkg, but no significant toxicity was detected at the ages of 14 and 21 days. Brain regions mainly affected were the cortex, septum, thalamus, hypothalamus and the cornu ammonis 1 region. To explore possible molecular mechanisms involved in this neurodegenerative process, we investigated the impact of MDMA on the expression of the neurotrophins brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and nerve growth factor. Neonatal exposure of 6-day-old rats to MDMA triggered a considerable increase in cortical BDNF and NT-3 levels. Moreover, P7 CD1BDNF knockout mice were noticeably more sensitive to MDMA exposure as compared to their wild-type age-matched littermates. These data suggest that a single injection of MDMA causes neurodegeneration in the neonatal rat brain. The upregulation of BDNF and NT-3 expression may indicate an important compensatory mechanism leading to the survival of neuronal cells in the developing brain.

Procedural and declarative memory task performance, and the memory consolidation function of sleep, in recent and abstinent ecstasyMDMA users.

EcstasyMDMA use has been associated with various memory deficits. This study assessed declarative and procedural memory in ecstasyMDMA users. Participants were tested in two sessions, 24 h apart, so that the memory consolidation function of sleep on both types of memory could also be assessed. Groups were drug-naive controls (n 24) recent ecstasyMDMA users, who had taken ecstasyMDMA 2-3 days before the first testing session (n 25), and abstinent users, who had not taken ecstasyMDMA for at least 8 days before testing (n 17). Procedural memory did not differ between groups, but greater lifetime consumption of ecstasy was associated with poorer procedural memory. Recent ecstasyMDMA users who had taken other drugs (mainly cannabis) 48-24 h before testing exhibited poorer declarative memory than controls, but recent users who had not taken other drugs in this 48-24-h period did not differ from controls. Greater lifetime consumption of ecstasy, and of cocaine, were associated with greater deficits in declarative memory. These results suggest that procedural, as well as declarative, memory deficits are associated with the extent of past ecstasy use. However, ecstasyMDMA did not affect the memory consolidation function of sleep for either the declarative or the procedural memory task.

Behavioral consequences of NMDA antagonist-induced neuroapoptosis in the infant mouse brain.

Exposure to NMDA glutamate antagonists during the brain growth spurt period causes widespread neuroapoptosis in the rodent brain. This period in rodents occurs during the first two weeks after birth, and corresponds to the third trimester of pregnancy and several years after birth in humans. The developing human brain may be exposed to NMDA antagonists through drug-abusing mothers or through anesthesia. We evaluated the long-term neurobehavioral effects of mice exposed to a single dose of the NMDA antagonist, phencyclidine (PCP), or saline, on postnatal day 2 (P2) or P7, or on both P2 and P7. PCP treatment on P2 P7 caused more severe cognitive impairments than either single treatment. Histological examination of acute neuroapoptosis resulting from exposure to PCP indicated that the regional pattern of degeneration induced by PCP in P2 pups was different from that in P7 pups. The extent of damage when evaluated quantitatively on P7 was greater for pups previously treated on P2 compared to pups treated only on P7. These findings signify that PCP induces different patterns of neuroapoptosis depending on the developmental age at the time of exposure, and that exposure at two separate developmental ages causes more severe neuropathological and neurobehavioral consequences than a single treatment.

Dopamine uptake inhibitors but not dopamine releasers induce greater increases in motor behavior and extracellular dopamine in adolescent rats than in adult male rats.

Most life-long drug addiction begins during adolescence. Important structural and functional changes in brain occur during adolescence and developmental differences in forebrain dopamine systems could mediate a biologic vulnerability to drug addiction during adolescence. Studies investigating age differences in psychostimulant responses have yielded mixed results, possibly because of different mechanisms for increasing extracellular dopamine. Recent research from our laboratory suggests that adolescent dopamine systems may be most affected by selective dopamine uptake inhibitors. We investigated age-related behavioral responses to acute administration of several dopamine uptake inhibitors methylphenidate, 1-2-bis-(4-fluorophenyl)methoxyethyl-4-(3-phenylpropyl)piperazine (GBR12909), and nomifensine and releasing agents amphetamine and methylenedioxymethamphetamine (MDMA) in adolescent and adult male rats. Methylphenidate and amphetamine effects on stimulated dopamine efflux were determined using fast-scan cyclic voltammetry in vivo. Dopamine uptake inhibitors but not dopamine releasing agents induced more locomotion andor stereotypy in adolescent relative to adult rats. MDMA effects were greater in adults at early time points after dosing. Methylphenidate but not amphetamine induced much greater dopamine efflux in periadolescent relative to adult rats. Periadolescent male rats are particularly sensitive to psychostimulants that are DAT inhibitors but are not internalized and do not release dopamine. Immaturity of DAT andor DAT associated signaling systems in adolescence specifically enhances behavioral and dopaminergic responses in adolescence.

Behavioural and molecular consequences of chronic cannabinoid treatment in Huntingtons disease transgenic mice.

Early loss of CB1 receptors is a hallmark of human Huntingtons disease. Data from rodent studies suggest that preservation and activation of CB1 receptors may be protective against disease progression. R61 transgenic mice are considered to be a model of early pathogenic changes in Huntingtons disease. We have shown previously that levels of CB1 in R61 mice prior to the onset of motor symptoms (12 weeks of age) remain high enough to justify commencement of cannabinoid drug treatment. Eight weeks of daily treatment with the cannabinoid agonists HU210 (0.01 mgkg) and Delta(9)-tetrahydrocannabinol (THC, 10.00 mgkg), or the inhibitor of endocannabinoid metabolism URB597 (0.30 mgkg), did not alter the progressive deterioration of performance observed in motor behavioural testing. HU210-treated R61 mice experienced a significant increase in seizure events suggesting that this therapy may lower the seizure threshold and cautioning against highly efficacious agonists as potential therapy in this disease. Molecular characterisation of brains at the end of the study showed that there were no significant effects of HU210 or THC treatment on the ligand binding of cannabinoid CB1, dopamine D1, D2, serotonin 5HT2A or GABA(A) receptors, nor CB1 or fatty acid amide hydrolase (FAAH) mRNA expression in R61 mice. Intriguingly, a significant increase in the number of ubiquitinated aggregates was observed in the striatum with HU210 treatment, indicating an influence of CB1 on the disease process. Chronic URB597 treatment preserved CB1 receptors in the R61 striatum, suggesting that the manipulation of endocannabinoid levels warrants further exploration.

Antipsychotic drugs prevent the motor hyperactivity induced by psychotomimetic MK-801 in zebrafish (Danio rerio).

Glutamate N-methyl-d-aspartate (NMDA) receptor antagonists, such as dizocilpine (MK-801), elicit schizophrenia-like symptoms in humans and a behavioral syndrome in rodents, characterized by hyperlocomotion and stereotyped actions, which is antagonized by antipsychotic drugs. Animal models of schizophrenia have been established and used for the development of new antipsychotic drugs. In this work we characterized the behavioral effects of MK-801 and investigated the effect of typical and atypical antipsychotic treatments on locomotor activity as well on the hyperlocomotion induced by MK-801 in zebrafish. MK-801 (20 microM) increased the locomotor behavior as measured by the number of line crossings, distance traveled, and the mean speed in the tank test after 15, 30, and 60 min of exposure. All tested antipsychotics counteracted MK-801-induced hyperactivity on all parameters analyzed and at doses that, given alone, had no effect on spontaneous locomotor activity. The results suggest a similar profile between typical and atypical antipsychotics in the reversal of locomotor disorders induced by MK-801. Moreover, an anxiolytic effect was verified at 30 and 60 min of MK-801 exposure, which was not reversed by antipsychotics tested in this work. In addition, olanzapine, which alone caused an anxiolytic response, when given with MK-801 potentiated the latters effect on anxiety. In this work we demonstrated the value of the zebrafish, a simple to use animal model, in developing some behavioral features observed in schizophrenia, which may indicate a new approach for drug screening.

Neurophysiological and cognitive effects of smoked marijuana in frequent users.

Previously, we reported that acute marijuana intoxication minimally affected complex cognitive performance of daily marijuana smokers. It is possible that the cognitive tests used were insensitive to marijuana-related cognitive effects. In the current study, electroencephalographic (EEG) signals were recorded as daily marijuana users performed additional tests of immediate working memory and delayed episodic memory, before and after smoking marijuana. Research volunteers (N24), who reported smoking approximately 24 marijuana cigarettesweek, completed this study. Participants completed baseline computerized cognitive tasks, smoked a single marijuana cigarette (0%, 1.8%, or 3.9% (9)-THC ww), and completed additional cognitive tasks sessions were separated by at least 72-hours. Cardiovascular and subjective effects were also assessed throughout sessions. Overall performance accuracy was not significantly altered by marijuana, although the drug increased response times during task performance and induced a response bias towards labeling new words as having been previously seen in the verbal episodic memory task. Marijuana reduced slow wave evoked potential amplitude in the episodic memory task and decreased P300 amplitude and EEG power in the alpha band in the spatial working memory task. Heart rate and positive subjective-effect ratings were increased in a (9)-THC concentration-dependent manner. Relative to previous findings with infrequent marijuana users, the frequent users in the current study exhibited similar neurophysiological effects but more subtle performance effects. These data emphasize the importance of taking into account the drug-use histories of research participants and examining multiple measures when investigating marijuana-related effects on cognitive functioning.

Involvement of mitochondriallysosomal toxic cross-talk in ecstasy induced liver toxicity under hyperthermic condition.

The initial objectives of this study were to evaluate the extent of 3, 4-methylenedioxymethamphetamine (MDMA) induced loss of cell viability (cytotoxicity), induction of reactive oxygen species formation and damage to sub-cellular organelles (e.g. mitochondrialysosomes) in freshly isolated rat hepatocytes under normothermic conditions (37 degrees C) and to compare the results with the effects obtained under hyperthermic conditions (41 degrees C). MDMA induced cytotoxicity, reactive oxygen species formation, mitochondrial membrane potential decline and lysosomal membrane leakiness in isolated rat hepatocytes at 37 degrees C. A rise in incubation temperature from 37 degrees C to 41 degrees C had an additivesynergic effect on the oxidative stress markers. We observed variations in mitochondrial membrane potential and lysosomal membrane stability that are significantly (P<0.05) higher than those under normothermic conditions. Antioxidants, reactive oxygen species scavengers, lysosomal inactivators, mitochondrial permeability transition (MPT) pore sealing agents, NADPH P450 reductase inhibitor, and inhibitors of reduced CYP2E1 and CYP2D6 prevented all MDMA induced hepatocyte oxidative stress cytotoxicity markers. It is therefore suggested that metabolic reductive activation of MDMA by reduced cytochrome P450s and glutathione could lead to generation of some biological reactive intermediates which could activate reactive oxygen species generation and cause mitochondrial and lysosomal oxidative stress membrane damages. We finally concluded that hyperthermia could potentiate MDMA induced liver toxicity probably through a mitochondriallysosomal toxic cross-talk in freshly isolated rat hepatocytes.

GABA(A) receptor density is altered by cannabinoid treatment in the hippocampus of adult but not adolescent rats.

Cannabinoids are known to induce transient psychotic symptoms and cognitive dysfunction in healthy individuals and contribute to trigger schizophrenia in vulnerable individuals, particularly during adolescence. Converging preclinical evidence suggests important interactions between cannabinoid and GABAergic systems. In the present study, we compared the effects of cannabinoid treatment on GABA(A) receptor binding in the brain of adolescent and adult rats. Adolescent (5 weeks old) and adult (10 weeks old) rats were treated with the synthetic cannabinoid HU210 (25, 50 or 100 microgkgday) or vehicle for 1, 4 or 14 days. Rats were sacrificed 24 h after the last injection and GABA(A) receptor density was measured in several brain regions using (35)STBPS and in vitro autoradiography. Adolescent rats had higher numbers of GABA(A) receptors compared to adults. A 24% increase of binding in adult rats treated with 100 microgkg HU210 for 14 days compared to controls was observed in the CA1 region of the hippocampus (16.1 versus 12.9 fmolmg tissue equivalent, t2.720, p<0.05). HU210 did not affect GABA(A) receptors in adolescent rats in any treatment regimen and in adult rats treated with HU210 for 1 or 4 days. These data suggest that long-term, high-dose treatment with HU210 increases GABA(A) receptors in the hippocampus of adult rats, changes that may interfere with associated hippocampal cognitive functions such as learning and memory. In addition, our results suggest that the adolescent brain does not display the same compensatory mechanisms that are activated in the adult brain following cannabinoid treatment.

Effects of cannabinoids on capsaicin receptor activity following exposure of primary sensory neurons to inflammatory mediators.

Activation of the cannabinoid 1 (CB1) receptor in cultured primary sensory neurons reduces responses mediated through the transient receptor potential vanilloid type 1 receptor (TRPV1), which plays a pivotal role in the development of heat hyperalgesia and visceral hyper-reflexia in inflammatory conditions. Here, we studied the effect of cannabinoid-evoked inhibitory effect on TRPV1 in inflammatory conditions. The effect of anandamide (1 nM-30 nM) and 1,1-dimethylheptyl-11-hydroxytetrahydrocannabinol (HU210 1 microM-10 microM) was assessed on capsaicin (10 nM or 100 nM)-evoked cobalt uptake in rat cultured primary sensory neurons following the incubation of the cells in an inflammatory environment created by the major inflammatory mediators, bradykinin (5 microM), prostaglandin E(2) (5 microM) and nerve growth factor (100 ngml) for 10 min. 1 nM and 10 nM anandamide significantly reduced the 10 nM but not the 100 nM capsaicin-evoked responses. HU210 did not produce a significant change in responses evoked by capsaicin at either of its concentrations. The anandamide-induced inhibitory effect could not be reversed by the CB1 receptor antagonist, rimonabant (200 nM) or the membrane-permeable cAMP analogue, 8Br-cAMP (100 microM). These findings suggest that anandamide may inhibit TRPV1-mediated responses in a non-CB1non-cannabinoid 2 receptor-dependent manner in primary sensory neurons in inflammatory conditions.

Pharmacological effects of cannabinoids on the Caco-2 cell culture model of intestinal permeability.

Activation of cannabinoid receptors decreases emesis, inflammation, gastric acid secretion, and intestinal motility. However, the effects of cannabinoids on intestinal permeability have not yet been established. The aim of the present study is to examine the effects of cannabinoids on intestinal permeability in an in vitro model. Caco-2 cells were grown until fully confluent on inserts in 12-well plates. Transepithelial electrical resistance (TEER) measurements were made as a measure of permeability. EDTA (50 μM) was applied to reversibly increase permeability (reduce TEER). The effects of cannabinoids on permeability in combination with EDTA, or alone, were assessed. Potential target sites of action were investigated using antagonists of the cannabinoid (CB)(1) receptor, CB(2) receptor, transient receptor potential vanilloid subtype 1 (TRPV1), peroxisome proliferator-activated receptor (PPAR)γ, PPARα, and a proposed cannabinoid receptor. When applied to the apical or basolateral membrane of Caco-2 cells, Δ(9)-tetrahydrocannabinol (THC) and cannabidiol (CBD) enhanced the speed of recovery of EDTA-induced increased permeability. This effect was sensitive to cannabinoid CB(1) receptor antagonism only. Apical application of endocannabinoids caused increased permeability, sensitive to cannabinoid CB(1) receptor antagonism. By contrast, when endocannabinoids were applied basolaterally, they enhanced the recovery of EDTA-induced increased permeability, and this involved additional activation of TRPV1. All cannabinoids tested increased the mRNA of the tight junction protein zona occludens-1, but only endocannabinoids also decreased the mRNA of claudin-1. These findings suggest that endocannabinoids may play a role in modulating intestinal permeability and that plant-derived cannabinoids, such as THC and CBD, may have therapeutic potential in conditions associated with abnormally permeable intestinal epithelium.

The role of monoamines in the changes in body temperature induced by 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) and its derivatives.

Hyperthermia is probably the most widely known acute adverse event that can follow ingestion of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) by recreational users. The effect of MDMA on body temperature is complex because the drug has actions on all three major monoamine neurotransmitters 5-hydroxytryptamine (5-HT), dopamine and noradrenaline, both by amine release and by direct receptor activation. Hyperthermia and hypothermia can be induced in laboratory animals by MDMA, depending on the ambient temperature, and involve both central thermoregulation and peripheral changes in blood flow and thermogenesis. Acute 5-HT release is not directly responsible for hyperthermia, but 5-HT receptors are involved in modulating the hyperthermic response. Impairing 5-HT function with a neurotoxic dose of MDMA or p-chlorophenylalanine alters the subsequent MDMA-induced hyperthermic response. MDMA also releases dopamine, and evidence suggests that this transmitter is involved in both the hyperthermic and hypothermic effects of MDMA in rats. The noradrenergic system is also involved in the hyperthermic response to MDMA. MDMA activates central alpha(2A)-adrenoceptors and peripheral alpha(1)-adrenoceptors to produce cutaneous vasoconstriction to restrict heat loss, and beta(3)-adrenoceptors in brown adipose tissue to increase heat generation. The hyperthermia occurring in recreational users of MDMA can be fatal, but data reviewed here indicate that it is unlikely that any single pharmaceutical agent will be effective in reversing the hyperthermia, so careful body cooling remains the principal clinical approach. Crucially, educating recreational users about the potential dangers of hyperthermia and the control of ambient temperature should remain key approaches to prevent this potentially fatal problem.

Mechanisms mediating the ability of caffeine to influence MDMA (Ecstasy)-induced hyperthermia in rats.

Caffeine exacerbates the hyperthermia associated with an acute exposure to 3,4 methylenedioxymethamphetamine (MDMA, Ecstasy) in rats. The present study investigated the mechanisms mediating this interaction. Adult male Sprague-Dawley rats were treated with caffeine (10 mg x kg(-1) i.p.) and MDMA (15 mg x kg(-1) i.p.) alone and in combination. Core body temperatures were monitored before and after drug administration. Central catecholamine depletion blocked MDMA-induced hyperthermia and its exacerbation by caffeine. Caffeine provoked a hyperthermic response when the catecholamine releaser d-amphetamine (1 mg x kg(-1)) was combined with the 5-HT releaser D-fenfluramine (5 mg x kg(-1)) or the non-selective dopamine receptor agonist apomorphine (1 mg x kg(-1)) was combined with the 5-HT(2) receptor agonist DOI (2 mg x kg(-1)) but not following either agents alone. Pretreatment with the dopamine D(1) receptor antagonist Schering (SCH) 23390 (1 mg x kg(-1)), the 5-HT(2) receptor antagonist ketanserin (5 mg x kg(-1)) or alpha(1)-adreno- receptor antagonist prazosin (0.2 mg x kg(-1)) blocked MDMA-induced hyperthermia and its exacerbation by caffeine. Co-administration of a combination of MDMA with the PDE-4 inhibitor rolipram (0.025 mg x kg(-1)) and the adenosine A(12) receptor antagonist 9-chloro-2-(2-furanyl)-1,2,4triazolo1,5-Cquinazolin-5-amine 15943 (10 mg x kg(-1)) or the A(2A) receptor antagonist SCH 58261 (2 mg x kg(-1)) but not the A(1) receptor antagonist DPCPX (10 mg x kg(-1)) exacerbated MDMA-induced hyperthermia. A mechanism comprising 5-HT and catecholamines is proposed to mediate MDMA-induced hyperthermia. A combination of adenosine A(2A) receptor antagonism and PDE inhibition can account for the exacerbation of MDMA-induced hyperthermia by caffeine.

Neuroprotective potential of CB1 receptor agonists in an in vitro model of Huntingtons disease.

The therapeutic potential of cannabinoids in Huntingtons disease (HD) has been investigated by several groups with complex and sometimes contrasting results. We sought to examine key points of intersection between cannabinoid receptor 1 (CB(1)) signalling, survival and the formation of mutant huntingtin aggregates in HD. Using a simplified pheochromocytoma (PC12) cell model of HD expressing exon 1 of wild-type or mutant huntingtin, we assayed cell death and aggregate formation using high-throughput cytotoxicity and image-based assays respectively. CB(1) activation by HU210 conferred a small but significant level of protection against mutant huntingtin-induced cell death. Pertussis toxin uncoupled HU210 from the inhibition of cAMP, preventing rescue of cell death. Phosphorylation of extracellular signal-regulated kinase (ERK) was also critical to CB(1)-mediated rescue. Conversely, treatments that elevated cAMP exacerbated mutant huntingtin-induced cell death. Despite opposing effects on HD cell survival, both HU210 and compounds that elevated cAMP increased the formation of mutant huntingtin aggregates. The increase in aggregation by HU210 was insensitive to Pertussis toxin and UO126, suggesting a G-protein alpha subtype s (G(s))-linked mechanism. We suggest that the CB(1) receptor, through G-protein alpha subtype io (G(io))-linked, ERK-dependent signal transduction, is a therapeutic target in HD. However the protective potential of CB(1) may be limited by promiscuous coupling to G(s), the stimulation of cAMP formation and increased aggregate formation. This may underpin the poor therapeutic efficacy of cannabinoids in more complex model systems and suggest that therapies that are selective for the G(io), ERK pathway may be of most benefit in HD.

The effects of Delta-tetrahydrocannabinol and cannabidiol alone and in combination on damage, inflammation and in vitro motility disturbances in rat colitis.

Cannabis is taken as self-medication by patients with inflammatory bowel disease for symptomatic relief. Cannabinoid receptor agonists decrease inflammation in animal models of colitis, but their effects on the disturbed motility is not known. (-)-Cannabidiol (CBD) has been shown to interact with Delta(9)-tetrahydrocannabinol (THC) in behavioural studies, but it remains to be established if these cannabinoids interact in vivo in inflammatory disorders. Therefore the effects of CBD and THC alone and in combination were investigated in a model of colitis. The 2,4,6-trinitrobenzene sulphonic acid (TNBS) model of acute colitis in rats was used to assess damage, inflammation (myeloperoxidase activity) and in vitro colonic motility. Sulphasalazine was used as an active control drug. Sulphasalazine, THC and CBD proved beneficial in this model of colitis with the dose-response relationship for the phytocannabinoids showing a bell-shaped pattern on the majority of parameters (optimal THC and CBD dose, 10 mg.kg(-1)). THC was the most effective drug. The effects of these phytocannabinoids were additive, and CBD increased some effects of an ineffective THC dose to the level of an effective one. THC alone and in combination with CBD protected cholinergic nerves whereas sulphasalazine did not. In this model of colitis, THC and CBD not only reduced inflammation but also lowered the occurrence of functional disturbances. Moreover the combination of CBD and THC could be beneficial therapeutically, via additive or potentiating effects.

Cannabinoid-1 receptor activation induces reactive oxygen species-dependent and -independent mitogen-activated protein kinase activation and cell death in human coronary artery endothelial cells.

Impaired endothelial activity andor cell death play a critical role in the development of vascular dysfunction associated with congestive heart failure, diabetic complications, hypertension, coronary artery disease and atherosclerosis. Increasing evidence suggests that cannabinoid 1 (CB(1)) receptor inhibition is beneficial in atherosclerosis and cardiovascular inflammation both in experimental models, as well as in humans. Here, we investigated the effects of CB(1) receptor activation with the endocannabinoid anandamide (AEA) or synthetic agonist HU210 on cell death and interrelated signal transduction pathways in human primary coronary artery endothelial cells (HCAECs). Cell death, CB(1) receptor expression, reactive oxygen species (ROS) generation and activation of signal transduction pathways in HCAECs were determined by flow cytometry and molecular biology tools. In HCAECs expressing CB(1) receptors (demonstrated by Western immunoblot and flow cytometry) AEA (5-15 microM) or HU210 (30-1000 nM) triggered concentration- and time-dependent activation of p38 and c-Jun NH(2)-terminal protein kinase (JNK)-mitogen-activated protein kinases (MAPKs), cell death and ROS generation. The AEA- or HU210-induced cell death and MAPK activation were attenuated by CB(1) antagonists SR141716 (rimonabant) and AM281, inhibitors of p38 and JNK-MAPKs or the antioxidant N-acetylcysteine. N-acetylcysteine alone prevented AEA- or HU210-induced ROS generation, but only partially attenuated MAPK activation and cell death. In contrast, in combination with CB(1) antagonists, N-acetylcysteine completely prevented these effects. CB(1) receptor activation in endothelial cells may amplify the ROS-MAPK activation-cell death pathway in pathological conditions when the endocannabinoid synthetic or metabolic pathways are dysregulated by excessive inflammation andor oxidativenitrosative stress, thereby contributing to the development of endothelial dysfunction and pathophysiology of multiple cardiovascular diseases.

The plant cannabinoid Delta9-tetrahydrocannabivarin can decrease signs of inflammation and inflammatory pain in mice.

The phytocannabinoid, Delta(9)-tetrahydrocannabivarin (THCV), can block cannabinoid CB(1) receptors. This investigation explored its ability to activate CB(2) receptors, there being evidence that combined CB(2) activationCB(1) blockade would ameliorate certain disorders. We tested the ability of THCV to activate CB(2) receptors by determining whether (i) it inhibited forskolin-stimulated cyclic AMP production by Chinese hamster ovary (CHO) cells transfected with human CB(2) (hCB(2)) receptors (ii) it stimulated (35)SGTPgammaS binding to hCB(2) CHO cell and mouse spleen membranes (iii) it attenuated signs of inflammationhyperalgesia induced in mouse hind paws by intraplantar injection of carrageenan or formalin and (iv) any such anti-inflammatory or anti-hyperalgesic effects were blocked by a CB(1) or CB(2) receptor antagonist. THCV inhibited cyclic AMP production by hCB(2) CHO cells (EC(50) 38 nM), but not by hCB(1) or untransfected CHO cells or by hCB(2) CHO cells pre-incubated with pertussis toxin (100 ng.mL(-1)) and stimulated (35)SGTPgammaS binding to hCB(2) CHO and mouse spleen membranes. THCV (0.3 or 1 mg.kg(-1) i.p.) decreased carrageenan-induced oedema in a manner that seemed to be CB(2) receptor-mediated and suppressed carrageenan-induced hyperalgesia. THCV (i.p.) also decreased pain behaviour in phase 2 of the formalin test at 1 mg.kg(-1), and in both phases of this test at 5 mg.kg(-1) these effects of THCV appeared to be CB(1) and CB(2) receptor mediated. THCV can activate CB(2) receptors in vitro and decrease signs of inflammation and inflammatory pain in mice partly via CB(1) andor CB(2) receptor activation.

Physical and functional interaction between CB1 cannabinoid receptors and beta2-adrenoceptors.

The CB(1) cannabinoid receptor and the beta(2)-adrenoceptor are G protein-coupled receptors (GPCRs) co-expressed in many tissues. The present study examined physical and functional interactions between these receptors in a heterologous expression system and in primary human ocular cells. Physical interactions between CB(1) receptors and beta(2)-adrenoceptors were assessed using bioluminescence resonance energy transfer (BRET). Functional interactions between these receptors were evaluated by examining receptor trafficking, as well as extracellular signal-regulated kinase (ERK) and cyclic AMP response element binding protein (CREB) signalling. Physical interactions between CB(1) receptors and beta(2)-adrenoceptors were demonstrated using BRET. In human embryonic kidney (HEK) 293H cells, co-expression of beta(2)-adrenoceptors tempered the constitutive activity and increased cell surface expression of CB(1) receptors. Co-expression altered the signalling properties of CB(1 )receptors, resulting in increased Galpha(i)-dependent ERK phosphorylation, but decreased non-Galpha(i)-mediated CREB phosphorylation. The CB(1) receptor inverse agonist AM251 (N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide) attenuated beta(2)-adrenoceptor-pERK signalling in cells expressing both receptors, while the CB(1) receptor neutral antagonist O-2050 ((6aR,10aR)-3-(1-methanesulfonylamino-4-hexyn-6-yl)-6a,7,10,10a-tetrahydro-6,6,9-trimethyl-6H-dibenzob,dpyran) did not. The actions of AM251 and O-2050 were further examined in primary human trabecular meshwork (HTM) cells, which are ocular cells endogenously co-expressing CB(1) receptors and beta(2)-adrenoceptors. In HTM cells, as in HEK 293H cells, AM251 but not O-2050, altered the beta(2)-adrenoceptor-pERK response. A complex interaction was demonstrated between CB(1) receptors and beta(2)-adrenoceptors in HEK 293H cells. As similar functional interactions were also observed in HTM cells, such interactions may affect the pharmacology of these receptors in tissues where they are endogenously co-expressed.

Evidence for both inverse agonism at the cannabinoid CB1 receptor and the lack of an endogenous cannabinoid tone in the rat and guinea-pig isolated ileum myenteric plexus-longitudinal muscle preparation.

Cannabinoid receptor agonists reduce intestinal propulsion in rodents through the CB(1) receptor. In addition to its antagonistic activity at this receptor, rimonabant (N-(piperidino)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-3-pyrazole-carboxyamide) alone augments intestinal transit. Using rat and guinea-pig ileum MPLM (myenteric plexus-longitudinal muscle) preparations, we investigated whether the latter effect was through inverse agonism or antagonism of endocannabinoid agonist(s). Inverse agonism was investigated by comparing the maximal enhancement of electrically evoked contractions of the MPLM by two CB(1) receptor antagonists, AM 251 (N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide) and O-2050 (6aR,10aR)-3-(1-methanesulphonylamino-4-hexyn-6-yl)-6a,7,10,10a-tetrahydro-6,6,9-trimethyl-6-H-dibenzob,dpyran, with that produced by rimonabant. To reveal ongoing endocannabinoid activity, effects of inhibiting endocannabinoid hydrolysis by fatty acid amide hydrolase (FAAH) using AA-5HT (arachidonyl-5-hydroxytryptamine), PMSF (phenylmethylsulphonyl fluoride) or URB-597 (3-carbamoyl-biphenyl-3-yl-cyclohexylcarbamate), or putative uptake using VDM-11 (5Z,8Z,11Z,14Z)-N-(4-hydroxy-2-methylphenyl)-5,8,11,14-eicosatetraenamide was evaluated. The presence of CB(1) receptors was revealed by antagonism of exogenous anandamide, arachidonylethanolamide (AEA) and WIN 55,212-2 (R)-()-2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)-pyrrolo1,2,3-de-1,4-benzoxazin-6-yl-1-naphthalenylmethanone mesylate by rimonabant. The rank order of potentiation of contractions was AM 251 > rimonabant > O-2050. Neither the FAAH inhibitors nor VDM-11 affected electrically evoked contractions. Each FAAH inhibitor increased the potency of AEA but not WIN 55,212-2. VDM-11 did not alter the inhibitory effect of AEA. The different levels of maximal potentiation of contractions by the CB(1) receptor antagonists suggest inverse agonism. The potentiation of the action of AEA by the FAAH inhibitors showed that FAAH was present. The lack of effect of FAAH inhibitors and VDM-11 alone on electrically evoked contractions, and on the potency of exogenous AEA suggests that pharmacologically active endocannabinoids were not released and the endocannabinoid transporter was absent. Thus, the CB(1) receptor antagonists behave as inverse agonists.

Structural MRI findings in long-term cannabis users what do we know

In animal studies, tetrahydrocannabinol (THC) has been found to affect brain morphology, particularly within areas rich in cannabinoid receptors (e.g., hippocampus, cerebral cortex). While cannabis remains the most widely used illicit drug worldwide, there has been limited work investigating its effects on human brain tissue. In this paper, we conducted a systematic review of existing structural magnetic resonance imaging studies to examine whether cannabis use is associated with significant changes in brain anatomy. We identified only 13 structural neuroimaging studies, which were diverse in terms of sample characteristics (e.g., age of participants, duration and frequency of use) and methodology (e.g., image analysis). No study found global structural changes in cannabis users, although six studies reported regional alterations. While changes in the hippocampus and parahippocampus were frequently identified, the findings were inconsistent across studies. The available literature also provides some evidence that regional structural changes are associated with cannabis use patterns (particularly cumulative dosage and frequency of use), as well as measures of psychopathology (e.g., measures of depressive and psychotic symptoms). Together, these structural imaging findings suggest that THC exposure does affect brain morphology, especially in medial-temporal regions. Given the small literature available and the limitations of studies to date, further research is clearly required, particularly given the prevalence of cannabis use worldwide.

Rhabdomyolysis After LSD Ingestion.

Rhabdomyolysis involves the release of intracellular contents secondary to muscle cell injury it generally presents with muscle pain and weakness. Illicit drugs, including phencyclidine, MDMA (ecstasy), and cocaine, are frequently documented as a cause of rhabdomyolysis. The authors review the literature on LSD-associated rhabdomyolysis. The authors provide a new case report of a previously health patient who suffered rhabdomyolysis after LSD ingestion. Although frequently listed as a cause of rhabdomyolysis, there are only limited reports of rhabdomyolysis in patients who have ingested LSD. The discussion outlines potential mechanisms and management of LSD-associated rhabdomyolysis. Consultation psychiatrists may be called to assist in management of acute mental-status changes or agitation associated with LSD intoxication in addition to facilitating subsequent chemical-dependency treatment.

Transition to and from injecting drug use among regular ecstasy users.

There is a scant amount of research investigating injecting drug use among people not selected on the basis of their injecting behaviour, and less attention has been given to stimulant users who may have a different experience with injecting drug use than opioid users who are more commonly studied. The current study aimed to investigate initiation to, and transition from, injecting drug use among a sentinel sample of regular ecstasy users in Australia. Participants were regular ecstasy users recruited across Australia in 2007 who were administered a structured interview that contained questions regarding initiation to injecting, reasons for injecting cessation, and likelihood of future injecting. Among those with a history of injecting drug use, injecting first occurred at a similar age to that of first ecstasy use. The majority did not inject themselves at the first occasion, and two-fifths were under the influence of other drugs at the time. Two-fifths of injectors had not injected in the past 6 months, with many relating this to concerns surrounding stigma. Route of drug administration is clearly not static, and the findings from this study suggest that some who have ceased injecting may still be at risk for future injecting.

A fast method for screening andor quantitation of tetrahydrocannabinol and metabolites in urine by automated SPELCMSMS.

Marijuana is one of the most commonly used illicit substances. The high usage of this substance results in it being commonly encountered in clinical samples throughout the USA and Europe. Due to its wide availability and use, marijuana is also commonly encountered in forensic toxicology laboratories. The proposed method utilized an automated solid phase extraction (SPE) coupled to liquid chromatographymass spectrometry (LCMS). The automated SPE procedure was developed using Hysphere C8-EC sorbent, and the high performance liquid chromatography (HPLC) separation was performed using an Xterra MS C(18) column with a total runtime of 10 min. The standard curves linearity generally fell between 6 and 500 ngmL. The limits of detection ranged from 2 to 4 ngmL, and the limits of quantitation ranged from 8 to 12 ngmL. The bias and imprecision were determined using a simple analysis of variance (single factor). The results demonstrate bias as <11% and percent imprecision as <12% for all components at four quality control levels. This method has been in use for over 2 years and has been applied to numerous forensic samples. When compared to other published methods, it exceeds others in its simplicity and speed of analysis. This method takes advantage of robotics and automation for a total analysis time of 10 min, including sample preparation, separation, and detection.

Does dopamine mediate the psychosis-inducing effects of cannabis A review and integration of findings across disciplines.

General population epidemiological studies have consistently found that cannabis use increases the risk of developing psychotic disorders in a dose-dependent manner. While the epidemiological signal between cannabis and psychosis has gained considerable attention, the biological mechanism whereby cannabis increases risk for psychosis remains poorly understood. Animal research suggests that delta-9-tetrahydrocannabinol (THC, the main psychoactive component of cannabis) increases dopamine levels in several regions of the brain, including striatal and prefrontal areas. Since dopamine is hypothesized to represent a crucial common final pathway between brain biology and actual experience of psychosis, a focus on dopamine may initially be productive in the examination of the psychotomimetic effects of cannabis. Therefore, this review examines the evidence concerning the interactions between THC, endocannabinoids and dopamine in the cortical as well as subcortical regions implicated in psychosis, and considers possible mechanisms whereby cannabis-induced dopamine dysregulation may give rise to delusions and hallucinations. It is concluded that further study of the mechanisms underlying the link between cannabis and psychosis may be conducted productively from the perspective of progressive developmental sensitization, resulting from gene-environment interactions.

Changes in 5-HT2A-mediated behavior and 5-HT2A- and 5-HT1A receptor binding and expression in conditional brain-derived neurotrophic factor knock-out mice.

Changes in brain-derived neurotrophic factor (BDNF) expression have been implicated in the etiology of psychiatric disorders. To investigate pathological mechanisms elicited by perturbed BDNF signaling, we examined mutant mice with central depletion of BDNF (BDNF(2L2LCk-cre)). A severe impairment specific for the serotonin 2A receptor (5-HT(2A)R) in prefrontal cortex was described previously in these mice. This is of much interest, as 5-HT(2A)Rs have been linked to neuropsychiatric disorders and anxiety-related behavior. Here we further characterized the serotonin receptor alterations triggered by BDNF depletion. 5-HT(2A) ((3)H-MDL100907) and 5-HT(1A) ((3)H-WAY100635) receptor autoradiography revealed site-specific alterations in BDNF mutant mice. They exhibited lower 5-HT(2A) receptor binding in frontal cortex but increased binding in hippocampus. Additionally, 5-HT(1A) receptor binding was decreased in hippocampus of BDNF mutants, but unchanged in frontal cortex. Molecular analysis indicated corresponding changes in 5-HT(2A) and 5-HT(1A) mRNA expression but normal 5-HT(2C) content in these brain regions in BDNF(2L2LCk-cre) mice. We investigated whether the reduction in frontal 5-HT(2A)R binding was reflected in reduced functional output in two 5-HT(2A)-receptor mediated behavioral tests, the head-twitch response (HTR) and the ear-scratch response (ESR). BDNF(2L2LCk-cre) mutants treated with the 5-HT(2A) receptor agonist (-)-2,5-dimethoxy-4-iodoamphetamine (DOI) showed a clearly diminished ESR but no differences in HTR compared to wildtypes. These findings illustrate the context-dependent effects of deficient BDNF signaling on the 5-HT receptor system and 5-HT(2A)-receptor functional output.

HPC-1syntaxin 1A gene knockout mice show abnormal behavior possibly related to a disruption in 5-HTergic systems.

HPC-1syntaxin 1A (STX1A) is thought to regulate the exocytosis of synaptic vesicles in neurons. In recent human genetic studies, STX1A has been implicated in neuropsychological disorders. To examine whether STX1A gene ablation is responsible for abnormal neuropsychological profiles observed in human psychiatric patients, we analysed the behavioral phenotype of STX1A knockout mice. Abnormal behavior was observed in both homozygotes (STX1A(--)) and heterozygotes (STX1A(-)) in a social interaction test, a novel object exploring test and a latent inhibition (LI) test, but not in a pre-pulse inhibition test. Interestingly, attenuation of LI, which is closely related to human schizotypic symptoms, was restored by administration of the selective serotonin reuptake inhibitor, fluoxetine, but not by the dopamine reuptake inhibitor, GBR12935, or the noradrenalin reuptake inhibitor, desipramine. We also observed that LI attenuation was restored by DOI (a 5-HT(2A) receptor agonist), but not by 8-OH-DPAT (a 5-HT(1A) receptor agonist), mCPP (a 5-HT(2C) receptor agonist), SKF 38393 (a D(1) receptor agonist), quinpirole (a D(2)D(3) receptor agonist) or haloperidol (a D(2)D(3) receptor antagonist). Thus, attenuation of LI is mainly caused by disruption of 5-HT-ergic systems via 5-HT(2A) receptors. In addition, 5-HT release from hippocampal and hypothalamic slices was significantly reduced. Therefore, ablation of STX1A may cause disruption of 5-HT-ergic transmission and induce abnormal behavior.

A follow-up study acute behavioural effects of Delta(9)-THC in female heterozygous neuregulin 1 transmembrane domain mutant mice.

Heavy cannabis use is linked with an increased risk for schizophrenia. We showed previously that male heterozygous neuregulin 1 transmembrane domain (Nrg1 HET) mice are more sensitive to some effects of the psychotropic cannabis constituent Delta(9)-tetrahydrocannabinol (THC). We report data from a follow-up study in female Nrg1 HET mice, investigating THC effects on behaviours with some relevance to schizophrenia. Mice were injected with THC (0, 5 or 10 mgkg i.p.) 30 min before a test battery open field, elevated plus maze, novel object recognition (set 1) or light-dark, social interaction (SI) and prepulse inhibition (PPI 1 variable interstimulus interval (ISI) set 2). Another set (set 3) was injected with the same doses of THC before a fixed interstimulus interval PPI test (PPI 2). Female Nrg1 HETs displayed the hallmark increased locomotor activity at 5 months and anxiolytic-like behaviour in the open field at 3 and 5 months. THC decreased locomotor activity in both genotypes. THC selectively reduced some SI behaviours in WT mice. Baseline PPI was enhanced in mutants under a variable ISI, while THC had no effect on PPI using either protocol. This study reports novel findings on the baseline PPI profile and resistance to THC-induced social withdrawal in female Nrg1 HET mice. This is the first description of THC effects in females of this mouse model and suggests that the transmembrane domain Nrg1 mutation does not appear to have a severe impact on the behavioural sensitivity to THC in female mice.

Ecstasy use among US adolescents from 1999 to 2008.

To investigate trends in rates of ecstasy use among US adolescents from 1999 to 2008, and to examine the associations between the major sociodemographic factors, especially gender, and ecstasy use, during this period. The adolescent subsamples (age 12-17) from 1999 to 2008 NHSDANSDUH surveys were used for the current study. Data from adolescents self-reports on use of ecstasy and of other drugs, as well as sociodemographic characteristics, were used in the analyses. There was an increasing trend in adolescent ecstasy use from 1999 to 2002, which was followed by a decreasing trend from 2002 to 2005, and a slight rise from 2005 to 2008. In contrast to some other drugs, ecstasy was more likely to be used by girls than by boys. This gender difference persisted over the 10-year period and could not be explained by other demographic factors. Given the known health consequences of ecstasy use, especially for females, the observed gender difference in adolescent ecstasy use should be taken into account by drug prevention and intervention programs.

Dimethyltryptamine (DMT) subjective effects and patterns of use among Australian recreational users.

Dimethyltryptamine (DMT) is an endogenous hallucinogen with traditional use as a sacrament in the orally active preparation of ayahuasca. Although the religious use of ayahuasca has been examined extensively, very little is known about the recreational use of DMT. In this study, Australian participants (n121) reporting at least one lifetime use of DMT completed an online questionnaire recording patterns of use, subjective effects and attitudes towards their DMT use. Smoking DMT was by far the most common route of administration (98.3%) with a comparatively smaller proportion reporting use of ayahuasca (30.6%). The reasons for first trying DMT were out of a general interest in hallucinogenic drugs (46.6%) or curiosity about DMTs effects (41.7%), while almost one-third (31.1%) cited possible psychotherapeutic benefits of the drug. An increase in psychospiritual insight was the most commonly reported positive effect of both smoked DMT (75.5%) and ayahuasca (46.7%), a finding that is consistent with other studies examining the ritualised use of ayahuasca in a religious context. Although previous studies of DMT use have examined ayahuasca use exclusively, the present study demonstrates the ubiquity of smoking as the most prevalent route of administration among recreational DMT users.

Lu AE58054, a 5-HT6 antagonist, reverses cognitive impairment induced by subchronic phencyclidine in a novel object recognition test in rats.

The in-vitro potency and selectivity, in-vivo binding affinity and effect of the 5-HT(6)R antagonist Lu AE58054 (2-(6-fluoro-1H-indol-3-yl)-ethyl-3-(2,2,3,3-tetrafluoropropoxy)-benzyl-amine) on impaired cognition were evaluated. Lu AE58054 displayed high affinity to the human 5-HT(6) receptor (5-HT(6)R) with a Ki of 0.83 nm. In a 5-HT(6) GTPgammaS efficacy assay Lu AE58054 showed no agonist activity, but demonstrated potent inhibition of 5-HT-mediated activation. Besides medium affinity to adrenergic alpha(1A)- and alpha(1B)-adrenoreceptors, Lu AE58054 demonstrated >50-fold selectivity for more than 70 targets examined. Orally administered Lu AE58054 potently inhibited striatal in-vivo binding of the 5-HT(6) antagonist radioligand (3)HLu AE60157 ((3)H8-(4-methylpiperazin-1-yl)-3-phenylsulfonylquinoline), with an ED(50) of 2.7 mgkg. Steady-state modelling of an acute pharmacokinetic5-HT(6)R occupancy time-course experiment indicated a plasma EC(50) value of 20 ngml. Administration of Lu AE58054 in a dose range (5-20 mgkg p.o.) leading to above 65% striatal 5-HT(6)R binding occupancy in vivo, reversed cognitive impairment in a rat novel object recognition task induced after subchronic treatment for 7 d with phencyclidine (PCP 2 mgkg b.i.d., i.p. for 7 d, followed by 7 d drug free). The results indicate that Lu AE58054 is a selective antagonist of 5-HT(6)Rs with good oral bioavailability and robust efficacy in a rat model of cognitive impairment in schizophrenia. Lu AE58054 may be useful for the pharmacotherapy of cognitive dysfunction in disease states such as schizophrenia and Alzheimers disease.

Contextualising psychological distress among regular ecstasy users the importance of sociodemographic factors and patterns of drug use.

Considerable concern has been raised about associations between ecstasy use and mental health. Studies of ecstasy users typically investigate varying levels of lifetime use of ecstasy, and often fail to account for other drug use and sociodemographic characteristics of participants, which may explain mixed findings. The current study aimed to examine the relationship between patterns of recent (last six months) ecstasy use and psychological distress among current, regular ecstasy users, controlling for sociodemographic risk factors and patterns of other drug use. Data were collected from regular ecstasy users (n 752) recruited from each capital city in Australia as part of the Ecstasy and related Drugs Reporting System (EDRS). Psychological distress was assessed using the Kessler Psychological Distress Scale (K10). Data were analysed using multinomial logistic regression. Seven per cent of the sample scored in the high distress category and 55% in the medium distress category. Patterns of ecstasy use were not independently associated with psychological distress. The strongest predictors of distress were female sex, lower education, unemployment, binge drug use including ecstasy (use for >48 h without sleep), frequent cannabis use and daily tobacco use. Regular ecstasy users have elevated levels of psychological distress compared with the general population however, ecstasy use per se was not independently related to such distress. Other factors, including sociodemographic characteristics and other drug use patterns, appear to be more important. These findings highlight the importance of targeting patterns of polydrug use in order to reduce drug-related harm among regular ecstasy users.

Cannabis and psychiatric disorders.

There are connection between use of cannabis and many psychiatric disturbances in adolescents, especially cannabis psychosis, depression, panic attacks and suicide. Negative effects could occur either as a result of a specific pharmacological effect of cannabis, or as the result of stressful experiences during the intoxication of cannabis in young people. Potentially is very dangerous high frequency suicidal ideation among cannabis users.

Effects of asenapine, olanzapine, and risperidone on psychotomimetic-induced reversal-learning deficits in the rat.

Asenapine is a new pharmacological agent for the acute treatment of schizophrenia and bipolar disorder. It has relatively higher affinity for serotonergic and alpha(2)-adrenergic than dopaminergic D(2) receptors. We evaluated the effects of asenapine, risperidone, and olanzapine on acute and subchronic psychotomimetic-induced disruption of cued reversal learning in rats. After operant training, rats were treated acutely with d-amphetamine (0.75 mgkg intraperitoneally i.p.) or phencyclidine (PCP 1.5mgkg i.p.) or subchronically with PCP (2mgkg i.p. for 7 days). We assessed the effects of acute coadministration of asenapine, risperidone, or olanzapine on acute d-amphetamine- and PCP-induced deficits and the effects of long-term coadministration of these agents (for 28 additional days) on the deficits induced by subchronic PCP. Deficits in reversal learning induced by acute d-amphetamine were attenuated by risperidone (0.2mgkg i.p.). Acute PCP-induced impairment of reversal learning was attenuated by acute asenapine (0.025 mgkg subcutaneously s.c.), risperidone (0.2mgkg i.p.), and olanzapine (1.0mgkg i.p.). Subchronic PCP administration induced an enduring deficit that was attenuated by acute asenapine (0.075 mgkg s.c.) and by olanzapine (1.5mgkg i.p.). Asenapine (0.075 mgkg s.c.), risperidone (0.2mgkg i.p.), and olanzapine (1.0mgkg i.p.) all showed sustained efficacy with chronic (29 days) treatment to improve subchronic PCP-induced impairments. These data suggest that asenapine may have beneficial effects in the treatment of cognitive symptoms in schizophrenia. However, this remains to be validated by further clinical evaluation.

Characterization of behavioral and endocrine effects of LSD on zebrafish.

Lysergic acid diethylamide (LSD) is a potent hallucinogenic drug that strongly affects animal and human behavior. Although adult zebrafish (Danio rerio) are emerging as a promising neurobehavioral model, the effects of LSD on zebrafish have not been investigated previously. Several behavioral paradigms (the novel tank, observation cylinder, light-dark box, open field, T-maze, social preference and shoaling tests), as well as modern video-tracking tools and whole-body cortisol assay were used to characterize the effects of acute LSD in zebrafish. While lower doses (5-100 microgL) did not affect zebrafish behavior, 250 microgL LSD increased top dwelling and reduced freezing in the novel tank and observation cylinder tests, also affecting spatiotemporal patterns of activity (as assessed by 3D reconstruction of zebrafish traces and ethograms). LSD evoked mild thigmotaxis in the open field test, increased light behavior in the light-dark test, reduced the number of arm entries and freezing in the T-maze and social preference test, without affecting social preference. In contrast, LSD affected zebrafish shoaling (increasing the inter-fish distance in a group), and elevated whole-body cortisol levels. Overall, our findings show sensitivity of zebrafish to LSD action, and support the use of zebrafish models to study hallucinogenic drugs of abuse.

Disruption of thermal nociceptive behaviour in mice mutant for the schizophrenia-associated genes NRG1, COMT and DISC1.

Abnormalities in pain perception, especially altered warmth and heat pain sensitivity, have been reported in schizophrenia. Therefore, genes associated with schizophrenia, including neuregulin-1 (NRG1), catechol-O-methyltranferase (COMT) and disrupted-in-schizophrenia-1 (DISC1), may play a role in modulating the physiological and psychological effects of pain stimuli in such patients. Thermal pain sensitivity was assessed in NRG1, COMT and DISC1 mutant mice, and the anti-nociceptive effects of acute Delta(9)-tetrahydrocannabinol (THC) were compared in NRG1 and COMT mutants. At baseline, deletion of NRG1 and DISC1 each reduced thermal pain sensitivity, while deletion of COMT increased pain sensitivity. Neither NRG1 nor COMT deletion altered the anti-nociceptive effects of acute systemic THC (8.0mgkg). These results indicate a differential contribution of NRG1 and DISC1 vis-à-vis COMT to the processing of thermal nociceptive stimuli and extend their phenotypic relationship to psychotic illness.

Meta-analysis of the efficacy and safety of Sativex (nabiximols), on spasticity in people with multiple sclerosis.

To determine the efficacy of Sativex (USAN nabiximols) in the alleviation of spasticity in people with multiple sclerosis. The results from three randomized, placebo-controlled, double-blind parallel group studies were combined for analysis. 666 patients with multiple sclerosis and spasticity. A 0-100 mm Visual Analogue Scale (VAS, transformed to a 0-10 scale) or a 0-10 Numerical Rating Scale (0-10 NRS) was used to measure spasticity. Patients achieving a > or 30% improvement from baseline in their spasticity score were defined as responders. Global impression of change (GIC) at the end of treatment was also recorded. The patient populations were similar. The adjusted mean change of the numerical rating scale from baseline in the treated group was -1.30 compared with -0.97 for placebo. Using a linear model, the treatment difference was -0.32 (95% CI -0.61, -0.04, p 0.026). A statistically significant greater proportion of treated patients were responders (odds ratio (OR) 1.62, 95% CI 1.15, 2.28 p 0.0073) and treated patients also reported greater improvement odds ratio 1.67 (95% CI 1.05, 2.65 p 0.030). High numbers of subjects experienced at least one adverse event, but most were mild to moderate in severity and all drug-related serious adverse events resolved. The meta-analysis demonstrates that nabiximols is well tolerated and reduces spasticity.

Immunohistochemical study of vesicle monoamine transporter 2 in the hippocampal formation of PCP-treated mice.

The exact pathophysiology of schizophrenia is unknown despite intensive scientific studies using molecular biology, psychopharmacology, neuropathology, etc. It is thought that neurodevelopmental failures such as neuronal network incompetence and the inappropriate formation of neurons affect the neurotransmitters. Several animal models have been created to investigate the etiology of this disease. In this study, we investigated the expression of vesicle monoamine transporter 2 (VMAT2), which has a significant role in neurotransmission, in the hippocampal formation in 1-phenylcyclohexylpiperazine (PCP)-treated mice using immunohistochemical staining technique to clarify neuronal abnormalities. PCP-treated mice are thought to be one of novel animal models for schizophrenia. The expression of VMAT2 in the hippocampal formation was significantly reduced overall in the PCP-treated mice compared to that in control (saline-treated) mice, also these reductions were observed throughout the brain. These facts implied that the pathophysiology of this disease involves abnormal monoaminergic transmission through VMAT2, despite PCP was the N-methyl-d-aspartate (NMDA) receptor antagonist that might induce glutamatergic abnormality. Since insufficient or excess release of neurotransmitter might alter neurochemical function and neurotransmission, VMAT2 might be an important target for biological research in psychiatric disease including schizophrenia.

Brain-derived neurotrophic factor controls cannabinoid CB1 receptor function in the striatum.

The role of brain-derived neurotrophic factor (BDNF) in emotional processes suggests an interaction with the endocannabinoid system. Here, we addressed the functional interplay between BDNF and cannabinoid CB(1) receptors (CB(1)Rs) in the striatum, a brain area in which both BDNF and CB(1)s play a role in the emotional consequences of stress and of rewarding experiences. BDNF potently inhibited CB(1)R function in the striatum, through a mechanism mediated by altered cholesterol metabolism and membrane lipid raft function. The effect of BDNF was restricted to CB(1)Rs controlling GABA-mediated IPSCs (CB(1)R(GABA)), whereas CB(1)Rs modulating glutamate transmission and GABA(B) receptors were not affected. The action of BDNF on CB(1)R(GABA) function was tyrosine kinase dependent and was complete even after receptor sensitization with cocaine or environmental manipulations activating the dopamine (DA)-dependent reward system. In mice lacking one copy of the BDNF gene (BDNF(-)), CB(1)R(GABA) responses were potentiated and were preserved from the action of haloperidol, a DA D(2) receptor (D(2)R) antagonist able to fully abolish CB(1)R(GABA) function in rewarded animals. Haloperidol also enhanced BDNF levels in the striatum, suggesting that this neurotrophin may act as a downstream effector of D(2)Rs in the modulation of cannabinoid signaling. Accordingly, 5 d cocaine exposure both reduced striatal BDNF levels and increased CB(1)R(GABA) activity, through a mechanism dependent on D(2)Rs. The present study identifies a novel mechanism of CB(1)R regulation mediated by BDNF and cholesterol metabolism and provides some evidence that DA D(2)R-dependent modulation of striatal CB(1)R activity is mediated by this neurotrophin.

Determining the subjective effects of TFMPP in human males.

Trifluoromethylphenyl piperazine (TFMPP) is an active constituent of a relatively new group of recreational drugs known as party pills. TFMPP has been anecdotally reported to induce mild psychedelic effects similar to lysergic acid diethylamide and psilocybin. There has been no research about the subjective effects of TFMPP in humans. This study aimed to investigate the subjective effects of TFMPP in human males. A randomised, double-blind, placebo-controlled trial design was used to investigate the subjective effects of TFMPP in 30 healthy, non-smoking male volunteers (mean age 24 - 4 years). Participants were randomised into two groups and given either TFMPP 60 mg (n 15) or placebo (n 15). Each participant completed three rating scales, the Addiction Research Centre Inventory (ARCI), the Profile of Mood States (POMS) and the Visual Analogue Scales (VAS), both before and 120 min after drug administration. Results from the ARCI indicated that TFMPP produced increases in dysphoria and dexamphetamine-like effects. TFMPP also increased ratings of tensionanxiety and confusionbewilderment as rated on the POMS. Results from the VAS indicated increases in drug liking, high and stimulated ratings relevant to placebo. Increased ratings of dexamphetamine-like effects, tensionanxiety, stimulated and high following TFMPP administration resemble the subjective effects of common amphetamine-type stimulants. However, increases in dysphoria and confusionbewilderment ratings following TFMPP are more commonly associated with drugs that have greater effects on serotonin release, binding and reuptake such as 1-3-chlorophenyl-piperazine, fenfluramine and lysergic acid diethylamide.

MDMA (ecstasy) delays pubertal development and alters sperm quality after developmental exposure in the rat.

3,4-Methylenedioxymethamphetamine, MDMA or ecstasy is consumed mainly by young population at childbearing age. Therefore, there may be a risk of exposure of some pregnant women. The effects of the developmental exposure to MDMA on the sexual development and long-term sexual behaviourfertility were assessed in Sprague-Dawley rats. MDMA was administered subcutaneously at 0 (control), 0.5, 5 and 10 mgkg to female rats once a day, three consecutive days a week during 10 weeks, including gestation and lactation. The male offspring was evaluated for sexual maturation and mated with untreated sexually receptive females to evaluate the mating and pregnancy rates. Hormonal, haematological, biochemical, histological, genotoxicological and testicular and sperm parameters were also evaluated. A significant higher incidence of DNA damage in sperm and interstitial oedema in testes was found. There was also a significant and dose-related decrease in sperm count and a significant decrease in sperm motility at all doses. A significant delay in preputial separation onset in all treated groups was observed. This study reports by the first time an alteration of spermatogenesis after in utero and lactation MDMA exposure in the rat.

Beta-carboline alkaloids bind DNA.

Beta-carboline alkaloids present in Peganum harmala (harmal) have recently drawn attention due to their antitumor activities. The mechanistic studies indicate that beta-carboline derivatives inhibit DNA topoisomerases and interfere with DNA synthesis. They interact with DNA via both groove binding and intercalative modes and cause major DNA structural changes. The aim of this study was to examine the interactions of five beta-carboline alkaloids (harmine, harmane, harmaline, harmalol and tryptoline) with calf-thymus DNA in aqueous solution at physiological conditions, using constant DNA concentration (6.25 mM) and various alkaloidspolynucleotide (phosphate) ratios of 1240, 1160, 180, 140, 120, 110, 15, 12 and 11. Fourier transform infrared (FTIR) and UV-visible spectroscopic methods were used to determine the ligand binding modes, the binding constants, and the stability of alkaloids-DNA complexes in aqueous solution. Spectroscopic evidence showed major binding of alkaloids to DNA with overall binding constants of K(harmine)-DNA3.44x10(7) M(-1), K(harmane)-DNA1.63x10(5) M(-1), K(harmaline)-DNA3.82x10(5) M(-1), K(harmalol)-DNA6.43x10(5) M(-1) and K(tryptoline)-DNA1.11x10(5) M(-1). The affinity of alkaloids-DNA binding is in the order of harmine>harmalol>harmaline>harmane>tryptoline. No biopolymer secondary structural changes were observed upon alkaloid interaction and DNA remains in the B-family structure in these complexes.

Metabolism of designer drugs of abuse an updated review.

This paper reviews the metabolism of new designer drugs of abuse that have emerged on the black market during the last years and is an update of a review published in 2005. The presented review contains data concerning the so-called 2C compounds (phenethylamine type) such as 4-bromo-2,5-dimethoxy-beta-phenethylamine (2C-B), 4-iodo-2,5-dimethoxy-beta-phenethylamine (2C-I), 2,5-dimethoxy-4-methyl-beta-phenethylamine (2C-D), 4-ethyl-2,5-dimethoxy-beta-phenethylamine (2C-E), 4-ethylthio-2,5-dimethoxy-beta-phenethylamine (2C-T-2), and 2,5-dimethoxy-4-propylthio-beta-phenethylamine (2C-T-7), beta-keto designer drugs such as 2-methylamino-1-(3,4-methylenedioxyphenyl)butan-1-one (butylone, bk-MBDB), 2-ethylamino-1-(3,4-methylenedioxyphenyl)propan-1-one (ethylone, bk-MDEA), 2-methylamino-1-(3,4-methylene notdioxy notphenyl)propan-1-one (methylone, bk-MDMA), and 2-methylamino-1-p-tolylpropane-1-one (mephedrone, 4-methyl-methcathinone), pyrrolidino notphenones such as 4-methyl-pyrrolidinobutyrophenone (MPBP) and alpha-pyrrolidinovalerophenone (PVP), phencyclidine-derived drugs such as N (1 phenylcyclohexyl) propanamine (PCPr), N-(1-phenylcyclohexyl)-2-ethoxyethanamine (PCEEA), N-(1-phenylcyclohexyl)-3-methoxypropanamine (PCMPA), and N-(1-phenylcyclohexyl)-2-methoxyethanamine (PCMEA), tryptamines such as 5-methoxy-N,N-diisopropyl nottryptamine (5-MeO-DIPT), and finally alpha-methylfentanyl (alpha-MF) and 3-methylfentanyl (3-MF). Papers have been considered and reviewed on the identification of in vivo or in vitro human or animal metabolites and the cytochrome P450 or monoamineoxidase isoenzyme-dependent metabolism.

Ecstasy overdoses at a New Years Eve rave--Los Angeles, California, 2010.

Ecstasy (3,4-methylenedioxymethamphetamine MDMA) is an illegal synthetic amphetamine used as a stimulant and hallucinogen. On January 4, 2010, the Los Angeles County (LAC) Department of Public Health (DPH) learned of six MDMA-related emergency department (ED) visits and one death, all linked to a New Years Eve event attended by approximately 45,000 persons. LAC DPH conducted an investigation to search for additional MDMA-related ED visits, characterize the cases, and determine whether drug contamination was involved. This report summarizes the results of the investigation, which determined that 18 patients visited EDs in LAC for MDMA-related illness within 12 hours of the rave. All were aged 16-34 years, and nine were female. In addition to using MDMA, 10 of the 18 had used alcohol, and five had used other drugs. Three patients were admitted to the hospital, including one to intensive care. A tablet obtained from one of the patients contained MDMA and caffeine, without known toxic contaminants. The cluster of apparent ecstasy overdoses occurred in the context of likely increasing MDMA use in the county during 2005-2009, as indicated by increased identification of MDMA-containing forensic specimens and a large increase in LAC residents entering drug treatment programs for MDMA. Collaboration between public health, police, fire, and emergency medical service (EMS) officials on a comprehensive prevention strategy might reduce the number of overdoses at similar events.

Afferent-specific AMPA receptor subunit composition and regulation of synaptic plasticity in midbrain dopamine neurons by abused drugs.

Ventral tegmental area (VTA) dopamine (DA) neurons play a pivotal role in processing reward-related information and are involved in drug addiction and mental illness in humans. Information is conveyed to the VTA in large part by glutamatergic afferents that arise in various brain nuclei, including the pedunculopontine nucleus (PPN). Using a unique rat brain slice preparation, we found that PPN stimulation activates afferents targeting GluR2-containing AMPA receptors (AMPAR) on VTA DA neurons, and these afferents did not exhibit long-term depression (LTD). In contrast, activation of glutamate afferents onto the same DA neurons via stimulation within the VTA evoked EPSCs mediated by GluR2-lacking AMPARs that demonstrated LTD or EPSCs mediated by GluR2-containing AMPA receptors that did not express LTD. Twenty-four hours after single cocaine injections to rats, GluR2-lacking AMPARs were increased at both PPN and local VTA projections, and this permitted LTD expression in both pathways. Single injections with the main psychoactive ingredient of marijuana, Delta(9)-tetrahydrocannabinol (Delta(9)-THC), increased GluR2-lacking AMPA receptors and permitted LTD in only the PPN pathway, and these effects were prevented by in vivo pretreatment with the cannabinoid CB1 receptor antagonist AM251. These results demonstrate that cocaine more globally increases GluR2-lacking AMPA receptors at all glutamate synapses on VTA dopamine neurons, whereas Delta(9)-THC selectively increased GluR2-lacking AMPA receptors at subcortical PPN synapses. This suggests that different abused drugs may exert influence over distinct sets of glutamatergic afferents to VTA DA neurons which may be associated with different reinforcing or addictive properties of these drugs.

Hallucinogens as hard science the adrenochrome hypothesis for the biogenesis of schizophrenia.

Working in a psychiatrically innovative environment created by the Government of Saskatchewan, Canada, Abram Hoffer and Humphry F. Osmond enunciated the adrenochrome hypothesis for the biogenesis of schizophrenia in 1952, slightly later proposing and, apparently, demonstrating, in a double-blind study, that the symptoms of the illness could be reversed by administering large doses of niacin. After placing the hypothesis within its ideological framework, the author describes its emergence and elaboration and discusses the empirical evidence brought against it. Hoffers idiosyncratic diagnostic procedures, especially his creation and use of a supposed biochemical marker for schizophrenia, are examined. The author argues that Hoffers conceptualization of schizophrenia, as well as his treatment approach, depended on a tautology. Following David Healy, the author treats the adrenochrome hypothesis as a version of a transmethylation theory, thus incorporating it into mainstream psychopharmacology.

Disposition of cannabinoids in oral fluid after controlled around-the-clock oral THC administration.

Oral fluid, a promising alternative matrix for drug monitoring in clinical and forensic investigations, offers noninvasive sample collection under direct observation. Cannabinoid distribution into oral fluid is complex and incompletely characterized due to the lack of controlled drug administration studies. To characterize cannabinoid disposition in oral fluid, we administered around-the-clock oral Delta(9)-tetrahydrocannabinol (THC) (Marinol) doses to 10 participants with current daily cannabis use. We obtained oral fluid samples (n440) by use of Quantisal collection devices before, during, and after 37 20-mg THC doses over 9 days. Samples were extracted with multiple elution solvents from a single SPE column and analyzed by 2-dimensional GC-MS with electron-impact ionization for THC, 11-hydroxy-THC (11-OH-THC), cannabidiol, and cannabinol and negative chemical ionization for 11-nor-9-carboxy-THC (THCCOOH). Linear ranges were 0.5-50 microgL, with the exception of cannabinol (1-50 microgL) and THCCOOH (7.5-500 ngL). THCCOOH was the most prevalent analyte in 432 samples (98.2%), with concentrations up to 1117.9 ngL. In contrast, 11-OH-THC was not identified in any sample cannabidiol and cannabinol were quantified in 3 and 8 samples, respectively, with maximum concentrations of 2.1 and 13 microgL. THC was present in only 20.7% of samples, with highest concentrations near admission (median 4.2 microgL, range 0.6-481.9) from previously self-administered smoked cannabis. Measurement of THCCOOH in OF not only identifies cannabis exposure, but also minimizes the possibility of passive inhalation. THCCOOH may be a better analyte for detection of cannabis use.

Assessing the functional significance of ecstasy-related memory deficits using a virtual paradigm.

Previous research shows that the use of ecstasy results in working memory and executive impairments in some users. The present study sought to assess the functional significance of such deficits using a virtual reality task. Twenty-three ecstasy-polydrug users and 26 nonusers were recruited. Individuals completed a drug use questionnaire measures of sleep quality and fluid intelligence. Participants also completed a virtual reality executive function task in which they play the role of an office worker for the day completing predefined tasks such as prioritising different activities according to their importance, organising the physical office environment and managing the outgoing mail in accordance with a delivery schedule. MANOVA revealed that ecstasy users performed worse on the virtual reality task overall, and this was due to poorer performance on the planning and selection subscales. Contrary to expectations, ecstasy-polydrug users performed better on the time-based prospective memory subscale. Indices of ecstasy use were correlated with the planning subscale of the virtual task. The present study provides further support for ecstasypolydrug related deficits in executive functioning. As it is possible that this task is more ecologically valid and relevant to day-to-day activities of many users, previous research finding null results on executive function tasks may have underestimated the impact of ecstasy-polydrug use on executive functioning.

Oral delta 9-tetrahydrocannabinol improved refractory Gilles de la Tourette syndrome in an adolescent by increasing intracortical inhibition a case report.

To describe the clinical course of the Delta 9-tetrahydrocannabinol (Delta 9-THC) treatment of a boy with Gilles de la Tourette Syndrome (TS) and comorbid attention-deficithyperactivity disorder (ADHD) in relation to Delta 9-THC plasma levels and intracortical inhibition measured by transcranial magnetic stimulation. The clinical course and pharmacological and neurophysiological measures are reported in a 15-year-old boy with treatment refractory TS plus ADHD leading to severe physical and psychosocial impairment. Administration of Delta 9-THC improved tics considerably without adverse effects, allowing parallel stimulant treatment of comorbid ADHD. Along with the Delta 9-THC treatment, intracortical inhibition was increased, reflected in the enhanced short-interval intracortical inhibition and the prolongation of the cortical silent period. Our observation suggests that Delta 9-THC might be a successful alternative in patients with severe TS refractory to classic treatment. Particularly in the case of stimulant-induced exacerbation of tics, Delta 9-THC might enable successful treatment of comorbid ADHD. The enhancement of intracortical inhibition might be mediated by modulating release of several neurotransmitters including dopamine and gamma-aminobutyric acid. Further studies are needed to substantiate our findings.

Recreational drugs, 3,4-Methylenedioxymethamphetamine(MDMA), 3,4-methylenedioxyamphetamine (MDA) and diphenylprolinol, inhibit neurite outgrowth in PC12 cells.

3,4-Methylenedioxymethamphetamine (MDMA) is widely abused as a psychoactive recreational drug. It is well known that MDMA induces neurotoxic damage of serotonergic nerve endings. Although drug abuse is increasing among youths, it is unclear whether recreational drugs affect the development of nerve growth. Thus, the present study examined the effect of recreational drugs, such as MDMA, 3,4-methylenedioxyamphetamine (MDA) and diphenylprolinol, a novel recreational drug with a similar chemical structure as that of psychoactive agent pipradrol, on nerve growth factor (NGF)-induced neurite outgrowth. These recreational drugs induced a dose-dependent cell death in PC12 cells. The IC(50) values of MDMA, MDA, R-diphenylprolinol and S-diphenylprolinol were 4.11 mM, 2.75 mM, 1.00 mM and 0.77 mM, respectively, at 24 hr. To examine the effects of these recreational drugs on NGF-induced neurite outgrowth, PC12 cells were treated with NGF together with MDMA, MDA, S-diphenylprolinol or R-diphenylprolinol at low toxic concentrations. The recreational drugs significantly suppressed neurite outgrowth of PC12 cells induced by NGF. The results suggest that these psychoactive recreational drugs may inhibit neurite growth and thus be implicated in their elicited neurotoxicity.

The small molecule phenamil is a modulator of adipocyte differentiation and PPARgamma expression.

We previously described the use of a cell-based screening approach to identify small molecules that regulate adipocyte differentiation. Here we identify the amiloride derivative phenamil as an adipogenic compound. Phenamil acutely induces expression of the key transcription factor of adipogenesis, peroxisome proliferator-activated receptor gamma (PPARgamma) and, consequently, promotes the differentiation of multiple preadipocyte cell lines, including 3T3-L1 and F442A. Interestingly, the adipogenic action of phenamil is distinct from and additive with both PPARgamma ligands and the previously identified adipogenic small molecule harmine. To identify signaling pathways mediating phenamils effects, we performed transcriptional profiling of 3T3-F442A preadipocytes. ETS variant 4 (ETV4) was identified as a gene rapidly induced by phenamil but not by other adipogenic small molecules or PPARgamma agonists. Transient expression of ETV4 in preadipocytes enhances the expression of PPARgamma. Stable overexpression of ETV4 promotes expression of PPARgamma and its downstream target genes and enhances morphological differentiation. Finally, knockdown of PPARgamma expression by shRNA blocks the effects of phenamil on adipocyte differentiation and gene expression, but it does not block phenamil induction of ETV4, which suggests that ETV4 acts upstream of PPARgamma in differentiation processes. These results identify a phenamil as new small molecule tool for the probing of adipocyte differentiation that acts, at least in part, through induction of ETV4 expression.

The persistence of the subjective in neuropsychopharmacology observations of contemporary hallucinogen research.

The elimination of subjectivity through brain research and the replacement of so-called folk psychology by a neuroscientifically enlightened worldview and self-conception has been both hoped for and feared. But this cultural revolution is still pending. Based on nine months of fieldwork on the revival of hallucinogen research since the Decade of the Brain, this paper examines how subjective experience appears as epistemic object and practical problem in a psychopharmacological laboratory. In the quest for neural correlates of (drug-induced altered states of) consciousness, introspective accounts of test subjects play a crucial role in neuroimaging studies. Firsthand knowledge of the drugs flamboyant effects provides researchers with a personal knowledge not communicated in scientific publications, but key to the conduct of their experiments. In many cases, the psychedelic experience draws scientists into the field and continues to inspire their self-image and way of life. By exploring these domains the paper points to a persistence of the subjective in contemporary neuropsychopharmacology.

Factors associated with condom use among young adult ecstasy users.

This paper examines the prevalence of and the factors associated with condom use in a sample of 283 young adult ecstasy users. The study, which relied upon targeted sampling and ethnographic mapping, took place between 2002 and 2004. It entailed conducting two-hour-long, face-to-face interviews in the Atlanta, Georgia metropolitan area. Condom use was inconsistent only 35.2% of all sex acts were protected. Using multiple regression, five factors were related to condom use race (Caucasians used condoms less than other groups), income (lower income greater condom use), relationship status (persons involved in relationships reported less condom use than those who were not involved), multiple sex partners (multiple sex partners more condom use), and condom use self-efficacy (higher efficacy level more condom use). Condom use rates were not optimal in this population. In particular, targeted interventions are needed for Caucasian ecstasy users. Intervention efforts ought to address relationship (in)fidelity as it pertains to engaging in safer sex practices, especially among persons involved in relationships. Intervention efforts also need to work to increase condom use self-efficacy.

Cannabinoids inhibit cellular respiration of human oral cancer cells.

The primary cannabinoids, Delta(9)-tetrahydrocannabinol (Delta(9)-THC) and Delta(8)-tetrahydrocannabinol (Delta(8)-THC) are known to disturb the mitochondrial function and possess antitumor activities. These observations prompted us to investigate their effects on the mitochondrial O(2) consumption in human oral cancer cells (Tu183). This epithelial cell line overexpresses bcl-2 and is highly resistant to anticancer drugs. A phosphorescence analyzer that measures the time-dependence of O(2) concentration in cellular or mitochondrial suspensions was used for this purpose. A rapid decline in the rate of respiration was observed when Delta(9)-THC or Delta(8)-THC was added to the cells. The inhibition was concentration-dependent, and Delta(9)-THC was the more potent of the two compounds. Anandamide (an endocannabinoid) was ineffective suggesting the effects of Delta(9)-THC and Delta(8)-THC were not mediated by the cannabinoidreceptors. Inhibition of O(2) consumption by cyanide confirmed the oxidations occurred in the mitochondrial respiratory chain. Delta(9)-THC inhibited the respiration of isolated mitochondria from beef heart. These results show the cannabinoids are potent inhibitors of Tu183 cellular respiration and are toxic to this highly malignant tumor.

Bruxism after 3,4-methylenedioxymethamphetamine (ecstasy) abuse.

Bruxism is a recognized side effect of several licit and illicit drugs. In this report, we illustrate this phenomenon in three patients suffering from 3,4-methylenedioxymethamphetamine (ecstasy) abuse.

Role of five synthetic reaction conditions on the stable isotopic composition of 3,4-methylenedioxymethamphetamine.

The identification of links between seizures of illicit 3,4-methylenedioxymethamphetamine (MDMA or ecstasy) has been a global target of law enforcement agencies in recent years. Previous work has shown that, when the reaction conditions are carefully repeated from batch to batch, stable isotope ratios allow the discrimination of MDMA.HCl batches according to synthetic route used for manufacture. In this study, the effects of altering five reaction conditions relating to the PtH(2) reductive amination synthesis were, for the first time, systematically investigated using a two level, five factor factorial design. Results indicate that the delta(2)H values of MDMA.HCl are affected by the length of imine stir time, and the delta(15)N values are affected by the degree of excess methylamine employed. Furthermore, the delta(13)C and delta(18)O values have been shown to be affected by the efficiency of the reaction, despite the similarity in carbon and oxygen composition of the starting material and product molecules. In addition to being of theoretical importance in this field of analytical science overall, this work is essential in order to more fully contextualize the interpretation of IRMS data which may be used as potential forensic evidence.

Effects of drugs and drug combinations in pigeons trained to discriminate among pentobarbital, dizocilpine, a combination of these drugs, and saline.

Drugs with multiple actions can have complex discriminative-stimulus properties. An approach to studying such drugs is to train subjects to discriminate among drug combinations and individual drugs in the combination so that all of the complex discriminative stimuli are present during training. In the current experiments, a four-choice procedure was used to train pigeons to discriminate among dizocilpine (noncompetitive NMDA receptor blocker), pentobarbital (GABA(A) receptor agonist), a fixed-dose combination of these two drugs, and saline. Following extended training, low doses of pentobarbital or dizocilpine administered alone produced saline-appropriate responding. Higher doses of pentobarbital produced responding on the pentobarbital-appropriate key and higher doses of dizocilpine produced responding on the dizocilpine key. Administering the lowest doses of pentobarbital and dizocilpine together resulted in responding on the saline-appropriate key. Increasing the dose of pentobarbital in the presence of low doses of dizocilpine produced responding primarily on the pentobarbital-appropriate key increasing the dose of dizocilpine in the presence of the lowest dose of pentobarbital produced responding primarily on the dizocilpine-appropriate key. Combining the higher doses of pentobarbital and dizocilpine resulted in responding primarily on the drug-combination key. Low doses of phencyclidine or ethanol produced responding on the saline-appropriate key, but intermediate doses resulted in individual subjects responding predominately on either the pentobarbital key, the dizocilpine key, or the drug-combination key depending on the subject. After the highest dose of phencyclidine or ethanol, most subjects responded predominantly on the drug-combination key. Low doses of other drugs tested produced responding on the saline-appropriate key. With the highest diazepam doses responding was largely confined to the pentobarbital-appropriate key. The highest doses of dextromethorphan or dextrorphan resulted in responding on the dizocilpine key more frequently than on other keys. Across a range of doses, morphine produced responding predominantly on the saline key. The results using the four-key procedure emphasized the role of both GABA(A) and NMDA receptors in the complex discriminative stimulus properties of phencyclidine and of ethanol.

Epigallocatechin gallate-induced modulation of FoxO signaling in mammalian cells and C. elegans FoxO stimulation is masked via PI3KAkt activation by hydrogen peroxide formed in cell culture.

The green tea flavonoid epigallocatechin gallate (EGCG) is demonstrated in this study to modulate FoxO transcription factors in human skin fibroblasts in culture. EGCG at 1 microM stimulated FoxO transcription factor nuclear accumulation and DNA binding activity. This effect was masked at higher EGCG concentrations (100 microM) by EGCG-derived hydrogen peroxide generated in cell culture media that stimulates phosphoinositide-3-kinase (PI3K)Akt signaling to attenuate FoxO activity, involving FoxO phosphorylation, nuclear exclusion and attenuation of DNA binding activity. Like low concentrations of EGCG, harmine, an inhibitor of the FoxO kinase DYRK1a, stimulated FoxO nuclear accumulation and DNA binding activity. Exposure of Caenorhabditis elegans worms to EGCG caused nuclear accumulation of the FoxO ortholog, DAF-16, and enhanced expression of the DAF-16 target gene, sod-3. In line with the role of FoxODAF-16 in the control of life span, C. elegans mean and maximum life span were enhanced by 20% and 13%, respectively, by EGCG.

Therapeutical use of the cannabinoids in psychiatry.

To review the main advances related to the potential therapeutic use of cannabinoid compounds in psychiatry. A search was performed in the online databases PubMed, ScieELO, and Lilacs for studies and literature reviews concerning therapeutic applications of cannabinoids in psychiatry, especially cannabidiol, rimonabant, Delta(9)-tetrahydrocannabinol, and their analogues. Cannabidiol was found to have therapeutic potential with antipsychotic, anxiolytic, and antidepressant properties, in addition to being effective in other conditions. Delta(9)-tetrahydrocannabinol and its analogues were shown to have anxiolytic effects in the treatment of cannabis dependence and to function as an adjuvant in the treatment of schizophrenia, although additional studies are necessary to support this finding. Rimonabant was effective in the treatment of the subjective and physiological symptoms of cannabis intoxication and functioned as an adjuvant in the treatment of tobacco addiction. The potential to induce adverse reactions such as depression and anxiety restrained the clinical use of this CB(1) antagonist. Cannabinoids may be of great therapeutic interest to psychiatry however, further controlled trials are necessary to confirm the existing findings and to establish the safety of such compounds.

Interpretation of blood analysis data found after passive exposure to cannabis.

When defendants are confronted with evidence of cannabinoids in their blood suggesting consumption of cannabis they sometimes argue that this could only be due to a passive exposure. The small number of controlled studies available showed that tetrahydrocannabinol (THC), the active ingredient of cannabis, was actually found in the blood after passive exposure to cannabis smoke. The resulting blood concentrations were dependent on the applied THC doses and the size of the room in which the passive exposure occurred. However, the quantitative data indicated in the publications of the 1980s cannot be fully compared with the results of modern analytical methods. Due to the rapid distribution of THC in the body, which occurs also after passive exposure to low doses, the THC concentration in serum to be expected in a blood sample taken 1 hour after exposure is less than 1 ngmL. For assessment of an alleged passive exposure, the metabolic THC-carboxylic acid, which is excreted more slowly, must also be taken into account. After passive exposure, similar and very low serum concentrations of THC and THC-carboxylic acid are to be expected (< 2 ngmL), while higher blood levels suggest the deliberate consumption of a psychoactive dose.

The peptide hemopressin acts through CB1 cannabinoid receptors to reduce food intake in rats and mice.

Hemopressin is a short, nine amino acid peptide (H-Pro-Val-Asn-Phe-Lys-Leu-Leu-Ser-His-OH) isolated from rat brain that behaves as an inverse agonist at the cannabinoid receptor CB(1), and is shown here to inhibit agonist-induced receptor internalization in a heterologous cell model. Since this peptide occurs naturally in the rodent brain, we determined its effect on appetite, an established central target of cannabinoid signaling. Hemopressin dose-dependently decreases night-time food intake in normal male rats and mice, as well as in obese obob male mice, when administered centrally or systemically, without causing any obvious adverse side effects. The normal, behavioral satiety sequence is maintained in male mice fasted overnight, though refeeding is attenuated. The anorectic effect is absent in CB(1) receptor null mutant male mice, and hemopressin can block CB(1) agonist-induced hyperphagia in male rats, providing strong evidence for antagonism of the CB(1) receptor in vivo. We speculate that hemopressin may act as an endogenous functional antagonist at CB(1) receptors and modulate the activity of appetite pathways in the brain.

The history of ergot of rye (Claviceps purpurea) III 1940-80.

The period 1940-80 in the history of ergot was dominated by two investigators, Arthur Stoll and Albert Hofmann. There was great excitement when their group isolated from ergot preparations the powerful psychotropic agent lysergic acid diethylamide (LSD). It was thought that this substance would help to find the cause of schizophrenia and other psychotic disorders, but it would prove to be a great disappointment and Hofmann would say later, in private, that he regretted having spent so much time on the compound. By contrast, bromocriptine, derived from ergocriptine, would prove a pivotal substance in our knowledge of dopamine receptors in the central nervous system. It is widely used for the suppression of lactation, the treatment of prolactinomas and the management of Parkinsons disease.

Photosensitized cleavage of plasmidic DNA by norharmane, a naturally occurring beta-carboline.

UV-A radiation (320-400 nm) induces damages to the DNA molecule and its components through photosensitized reactions. Beta-carbolines (betaCs), heterocyclic compounds widespread in biological systems, participate in several biological processes and are able to act as photosensitizers. The photosensitization of plasmidic DNA by norharmane in aqueous solution under UV-A radiation was studied. The effect of pH was evaluated and the participation of reactive oxygen species (ROS), such as hydroxyl radical (HO), superoxide anion (O(2)(-)) and singlet oxygen ((1)O(2)) was investigated. A strong dependence of the photosensitized DNA relaxation on the pH was observed. The extent of the reaction was shown to be higher in the experiments performed at pH 4.7 than those performed at pH 10.2. As was expected, an intermediate extent of the reaction was observed at physiological pH (pH 7.4). Kinetic studies using ROS scavengers revealed that the chemical reactions between ROS and DNA are not the main pathways responsible for the damage of DNA. Consequently, the predominant mechanism yielding the DNA strand break takes place most probably via a type I mechanism (electron transfer) from the single excited state (S(1)) of the protonated form of norharmane ((1)nHoH()). Additional information about the nature of the norharmane electronic excited states involved in the photocleavage reaction was obtained by using the N-methyl derivative of norharmane (N-methyl-norharmane).

Tetrahydrocannabinol (THC) for cramps in amyotrophic lateral sclerosis a randomised, double-blind crossover trial.

Many patients with amyotrophic lateral sclerosis (ALS) experience cramps during the course of the disease but so far, none of the medications used has been of proven benefit. The objective was to determine the effect of orally administered tetrahydrocannabinol (THC) on cramps in ALS patients. The authors conducted a randomised, double-blind, placebo-controlled crossover trial in 27 ALS patients suffering from moderate to severe (visual analogue scale (VAS) VAS≥4) daily cramps. There were 7 women and 20 men with a mean age of 57 years and a mean functional ALS score (ALSFRS-R) of 38.4. Patients were randomly assigned to receive 5 mg THC twice daily followed by placebo or vice versa. Each treatment period lasted for 2 weeks and was preceded by a 2-week drug-free observation period (run-in, wash-out period respectively). The primary outcome measure was change in cramp intensity as assessed by a VAS. Secondary outcome measures included the number of cramps per day, number of cramps during daytime and bedtime, intensity of fasciculations (VAS) as well as validated measures of quality of life (ALSAQ-40), quality of sleep (SDQ), appetite (FAACT) and depression (HADS). Complete data were available from 22 patients. THC was well tolerated. There was no evidence for a treatment effect on cramp intensity, number of cramps, fasciculation intensity or any of the other secondary outcome measures. This interventional study with orally administered THC 5 mg twice daily did not demonstrate subjective improvement of cramp intensity in ALS patients.

A latent pharmacokinetic time profile to model dose-response survival data.

The accelerating rotarod test is a preclinical pharmacodynamic test to assess the effect of a treatment on an animals motor coordination. Two models are proposed to analyze the dose-response time-to-event data that typically result from such experiments (1) a linear regression model and (2) an E(max) model with latent drug concentration at the site of action. Both cope with the survival character of the data. The latter model allows a direct comparison of compounds, but raises the question of whether the study design would benefit from the inclusion of additional mice for plasma concentration sampling on the one hand or whether additional time-to-event data without plasma concentration sampling should be ascertained from these additional mice on the other hand. A simulation study explores the impact on operational characteristics of this change of study design.

Marijuana correlates with use of other illicit drugs in a pain patient population.

A significant number of chronic pain patients may use marijuana. Physicians treating those patients can benefit by knowing whether their patients using marijuana are at higher risk for using other illicit drugs such as cocaine andor methamphetamine. Our objective was to determine whether marijuana-using chronic pain patients have a higher incidence of cocaine andor methamphetamine use. A retrospective study of the incidence of pain patients using marijuana andor other illicit drugs such as methamphetamine and cocaine versus the incidence of pain patients not using marijuana but using methamphetamine andor cocaine. Urine specimens from chronic pain patients were analyzed by LC-MSMS to determine the co-occurrence of these abused substances. In this study 21,746 urine specimens were obtained from chronic pain patients. We found a 13.0% incidence of patients positive for the acid form of Tetrahydrocannabinol (THCA). The percentage of those positive for cocaine was 4.6%, those positive for methamphetamine totaled 1.07%. Using both chi-square and a Logistic Regression analysis, we determined that there was a correlation between marijuana use and the use of other illicit drugs. The odds ratio was > 3.7 for other illicit drug use. The study is limited in that we obtained no data as to the causal relationships of this type of drug use. Pain physicians should be aware that this relationship exists and marijuana-using patients are at greater risk for use of other illicit drugs although no causal relationship is implied. Increased monitoring of these patients may help minimize potential morbidity due to drug interactions as well as identify patients who may be diverting prescriptions in order to pay for illicit drugs.

Catharanthine alkaloids are noncompetitive antagonists of muscle-type nicotinic acetylcholine receptors.

We compared the interaction of several catharanthine alkaloids including, ibogaine, vincristine, and vinblastine, with that for the noncompetitive antagonist phencyclidine (PCP) at muscle nicotinic acetylcholine receptors (AChRs) in different conformational states. The results established that catharanthine alkaloids (a) inhibit, in a noncompetitive manner, (-)-epibatidine-induced Ca(2) influx in TE671-halpha1beta1gammadelta cells with similar potencies (IC(50)17-25microM), (b) inhibit (3)HTCP binding to the desensitized Torpedo AChR with higher affinity compared to the resting AChR, and (c) enhance (3)Hcytisine binding to resting but activatable Torpedo AChRs, suggesting desensitizing properties. Interestingly, PCP inhibits (3)Hibogaine binding to the AChR in a steric fashion. This is corroborated by additional docking experiments indicating that the amino groups of neutral ibogaine form hydrogen bonds with the serine ring (position 6), a location shared with PCP. Since protonated ibogaine forms a salt bridge with one of the acidic residues at the outer ring (position 20), this ligand could be first attracted to the entrance of the channel by electrostatic interactions. Our data indicate that the catharanthine moiety is a minimum structural requirement for AChR inhibition including, ion channel blocking and desensitization, and that ibogaine and PCP bind to overlapping sites in the desensitized AChR ion channel.

Salvinorin A fails to substitute for the discriminative stimulus effects of LSD or ketamine in Sprague-Dawley rats.

Salvia divinorum is a small perennial shrub that has gained recent popularity among the drug-using subculture as a legal alternative to hallucinogens. Salvinorin A, the main active compound found in the S.divinorum plant, is an atypical hallucinogen with pharmacological selectivity at kappa opioid (KOP) receptor sites and is a unique non-nitrogenous neoclerodane diterpene which is structurally distinct from other opioid compounds. The novel structure of salvinorin A and its specific binding affinity to KOP receptors provide a unique opportunity to investigate neurochemical mechanisms of hallucination and hallucinogenic compounds. The current investigation assessed the substitution of salvinorin A in 16 male Sprague-Dawley rats trained to discriminate either the prototypical serotonergic hallucinogen, LSD (0.08mgkg, S.C., n8) or the dissociative anesthetic and glutamatergic hallucinogen, ketamine (8.0mgkg, I.P., n8) from vehicle under a FR 20 schedule of food-reinforced responding. Results indicated that neither LSD nor ketamine discrimination generalized to salvinorin A. These findings are consistent with the growing body of evidence that salvinorin A is pharmacologically distinct from other traditional hallucinogenic compounds.

The neurobiology of the switch process in bipolar disorder a review.

The singular phenomenon of switching from depression to its opposite state of mania or hypomania, and vice versa, distinguishes bipolar disorder from all other psychiatric disorders. Despite the fact that it is a core aspect of the clinical presentation of bipolar disorder, the neurobiology of the switch process is still poorly understood. In this review, we summarize the clinical evidence regarding somatic interventions associated with switching, with a particular focus on the biologic underpinnings presumably involved in the switch process. Literature for this review was obtained through a search of the MEDLINE database (1966-2008) using the following keywords and phrases switch, bipolar disorder, bipolar depression, antidepressant, SSRIs, tricyclic antidepressants, norepinephrine, serotonin, treatment emergent affective switch, mania, hypomania, HPA-axis, glucocorticoids, amphetamine, dopamine, and sleep deprivation. All English-language, peer-reviewed, published literature, including randomized controlled studies, naturalistic and open-label studies, and case reports, were eligible for inclusion. Converging evidence suggests that certain pharmacologic and nonpharmacologic interventions with very different mechanisms of action, such as sleep deprivation, exogenous corticosteroids, and dopaminergic agonists, can trigger mood episode switches in patients with bipolar disorder. The switch-inducing potential of antidepressants is unclear, although tricyclic antidepressants, which confer higher risk of switching than other classes of antidepressants, are a possible exception. Several neurobiological factors appear to be associated with both spontaneous and treatment-emergent mood episode switches these include abnormalities in catecholamine levels, up-regulation of neurotrophic and neuroplastic factors, hypothalamic-pituitary-adrenal axis hyperactivity, and circadian rhythms. There is a clear need to improve our understanding of the neurobiology of the switch process research in this field would benefit from the systematic and integrated assessment of variables associated with switching.

Use of non-occupational post-exposure prophylaxis does not lead to an increase in high risk sex behaviors in men who have sex with men participating in the EXPLORE trial.

Non-occupational post-exposure prophylaxis (nPEP) use is an HIV prevention strategy that has been recommended by the CDC to prevent HIV infection after a high risk sexual exposure since 1997. In a behavioral intervention trial of 4,295 MSM we assessed perceptions and use of nPEP over 4 years in six cities across the United States. Overall, 1.9% of MSM reported use of nPEP prior to enrollment, and 6.3% at least once during the trial. Awareness of nPEP was reported by 47.5%, with higher awareness in two sites with funded nPEP programs. Three seroconversions occurred in the 384 visits where nPEP courses were reported, with no effect of nPEP on risk of HIV acquisition in this cohort (hazard ratio 0.91, 95% confidence interval 0.29, 2.86). NPEP users were a riskier group increased odds of nPEP use were observed in association with multiple partners and unprotected receptive and insertive anal sex with HIV infected partners and partners with unknown HIV status. NPEP use was also associated with use of illicit drugs (injection drugs, crack cocaine, hallucinogens, and amphetamines). Importantly, willingness to use nPEP after high risk sex was associated with lower odds of high risk sex. After an episode of nPEP use, nPEP users remained more likely to report high risk sex than those in this cohort who had not previously used nPEP. However, within the subset of people who had previously reported high risk sex, previous nPEP use was not associated with higher odds of high risk sex, thus allaying fears that availability of nPEP would lead to an increase in high risk sex.

Opioid antagonism enhances marijuanas effects in heavy marijuana smokers.

Studies in laboratory animals strongly suggest reciprocal modulation of the opioidergic and endocannabinoid systems, a relationship that has not been demonstrated in humans. This study sought to clarify this interaction by assessing how a range of naltrexone doses altered the subjective, cognitive, and cardiovascular effects of marijuana. Daily marijuana smokers (n 29) participated in this within-subject, randomized, double-blind, placebo-controlled study. Naltrexone (0, 12, 25, 50, or 100 mg) was administered before active or inactive marijuana (3.27 or 0% THC) was smoked. Active marijuana increased subjective ratings of marijuana Strength, High, and positive subjective ratings of marijuana quality and drug effect including Liking, Good, and Take Again compared to inactive marijuana. Naltrexone alone decreased ratings of Liking, Take Again, and Stimulated compared with placebo, but increased ratings of drug Strength, High, Good, Liking, Stimulated, and Take Again when administered under active marijuana conditions. Active marijuana did not affect performance on cognitive tasks relative to inactive marijuana, whereas naltrexone decreased performance when administered alone or in combination with active marijuana. Active marijuana increased heart rate compared to inactive marijuana under placebo naltrexone conditions. Although naltrexone alone decreased heart rate, it further increased marijuanas cardiovascular effect. In heavy marijuana smokers opioid-receptor blockade enhanced the subjective and cardiovascular effects of marijuana, suggesting that endogenous opioids dampen cannabinoid effects in this population. These findings demonstrate that a broad range of clinically used doses of naltrexone potentially increases the abuse liability and cardiovascular risks of cannabinoids.

Morbidity associated with MDMA (ecstasy) abuse a survey of emergency department admissions.

We conducted a prospective, representative-sample nationwide study on morbidity related to 3,4, methylenedioxymethamphetamine (MDMA ecstasy) as determined from admissions to 5 geographically representative emergency departments (EDs) and from data from the poison information center (PIC). MDMArelated ED admissions were analyzed over a 7-month period and the records of all PIC calls were reviewed. There were 52 (age 15-44 years, 32 males) ecstasy-related ED admissions during the study period. Most (68%) admissions presented to the ED at night, 52% on weekends and 44% consumed the drug at clubs and parties. Forty-six percent of the patients took between 12 to 3 tablets and 29 patients (56%) had taken ecstasy before. Twenty-two subjects (42%) reported poly-drug use. Fifteen subjects (29%) required hospitalization, six of them (11%) to the intensive care unit. The most common manifestations were restlessness, agitation, disorientation, shaking, high blood pressure, headache and loss of consciousness. More serious complications were hyperthermia, hyponatremia, rhabdomyolysis, brain edema and coma. The image of ecstasy as a safe party drug is spurious. The results of this study confirm that the drug bears real danger of physical harm and of behavioral, psychological and psychiatric disturbances.

F15599, a preferential post-synaptic 5-HT1A receptor agonist activity in models of cognition in comparison with reference 5-HT1A receptor agonists.

We assessed the activity of F15599, a selective and high efficacy 5-HT(1A) agonist that preferentially activates post- versus pre-synaptic receptors, in rat cognitionmemory models. F15599 (0.16 mgkg i.p.) partially alleviated detrimental effects of phencyclidine on working and reference memory deficit in a hole-board model. It also attenuated phencyclidine-induced deficit of cognitive flexibility in a reversal learning task, without effects of its own. F13714 (0.04 mgkg) a chemical congener of F15599, and 8-OH-DPAT (0.01 or 0.16), were inactive against these phencyclidine-induced deficits, andor even worsened basal performances. F15599 (0.04-2.5) was less disruptive than F13714 (0.005-0.16) or 8-OH-DPAT (0.01-0.63), on basal performance in models of attention (5-choice serial reaction time task) and working memory (delayed non-matching to position). Finally, unlike either comparator, F15599 reduced PPI with modest potency and only partially. To conclude, F15599, in models of memorycognition, has a more favourable profile than F13714 and 8-OH-DPAT. This suggests that preferential activation of post-synaptic 5-HT(1A) receptors could prove useful in pathologies characterized by cognitivememory deficiencies, such as schizophrenia and depression.

Disruption of performance in the five-choice serial reaction time task induced by administration of N-methyl-D-aspartate receptor antagonists relevance to cognitive dysfunction in schizophrenia.

Schizophrenia patients suffer from cognitive impairments that are not satisfactorily treated by currently available medications. Cognitive dysfunction in schizophrenia encompasses deficits in several cognitive modalities that can be differentially responsive to different medications and are likely to be mediated by different neurobiological substrates. Translational animal models of cognitive deficits with relevance to schizophrenia are critical for gaining insights into the mechanisms underlying these impairments and developing more effective treatments. The five-choice serial reaction time task (5-CSRTT) is a cognitive task used in rodents that allows simultaneous assessment of several cognitive modalities, including attention, response inhibition, cognitive flexibility, and processing speed. Administration of N-methyl-D-aspartate (NMDA) glutamate receptor antagonists disrupts multiple 5-CSRTT performance measures in a way that mirrors various cognitive deficits exhibited by schizophrenia patients. Some of these disruptions are partially attenuated by antipsychotic medications that exhibit partial effectiveness on cognitive dysfunction in schizophrenia, suggesting that the model has predictive validity. Examination of the effects of pharmacological manipulations on 5-CSRTT performance disruptions induced by NMDA antagonists have implicated a range of brain regions, neurotransmitter systems, and specific receptor subtypes in schizophrenia-like impairment of different cognitive modalities. Thus, disruption of 5-CSRTT performance by NMDA antagonists represents a valuable tool for exploring the neurobiological bases of cognitive dysfunction in schizophrenia.

Cyclooxygenase-2 and tissue inhibitor of matrix metalloproteinases-1 confer the antimigratory effect of cannabinoids on human trabecular meshwork cells.

Cannabinoids have received considerable attention as potential antiglaucomatous drugs. Recently, prostaglandins (PG) have been suggested to contribute to this effect. Within the factors conferring the development of glaucoma, depletion of the aqueous humor outflow-regulating trabecular meshwork (TM) cells elicited by migration from the outflow system is considered to play a pivotal role. This study therefore investigates the impact of two cannabinoids, Delta(9)-tetrahydrocannabinol (THC) and R()-methanandamide (MA), on the migration of human TM cells and the involvement of the PG-synthesizing enzyme cyclooxygenase-2 (COX-2) and one of its potential downstream targets, the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), to this response. Using Boyden chamber assays cannabinoids were shown to elicit an antimigratory effect that was reversed by antagonists for CB(1) as well as CB(2) receptors and accompanied by upregulation of COX-2 and TIMP-1 expression and PGE(2) synthesis. Knockdown of cannabinoid-induced COX-2 or TIMP-1 expression by siRNA or inhibition of COX-2 activity by NS-398 led to a significant suppression of this antimigratory action. Migration was also diminished by the major COX-2 product PGE(2) and by recombinant TIMP-1. Experiments using selective E prostanoid (EP) receptor agonists and antagonists revealed that decreased migration by PGE(2), THC and MA was mediated via EP(2) and EP(4) receptors. Finally, the cannabinoid-mediated increases of TIMP-1 levels were abolished by NS-398, and PGE(2) was shown to elicit a concentration-dependent increase of TIMP-1. Collectively, this data demonstrate a COX-2-dependent upregulation of TIMP-1 conferring the antimigratory action of cannabinoids. A decreased migration reducing TM cell loss in glaucoma might be involved in the antiglaucomatous action of cannabinoids.

Potency trends of Δ9-THC and other cannabinoids in confiscated cannabis preparations from 1993 to 2008.

The University of Mississippi has a contract with the National Institute on Drug Abuse (NIDA) to carry out a variety of research activities dealing with cannabis, including the Potency Monitoring (PM) program, which provides analytical potency data on cannabis preparations confiscated in the United States. This report provides data on 46,211 samples seized and analyzed by gas chromatography-flame ionization detection (GC-FID) during 1993-2008. The data showed an upward trend in the mean Δ(9)-tetrahydrocannabinol (Δ(9)-THC) content of all confiscated cannabis preparations, which increased from 3.4% in 1993 to 8.8% in 2008. Hashish potencies did not increase consistently during this period however, the mean yearly potency varied from 2.5-9.2% (1993-2003) to 12.0-29.3% (2004-2008). Hash oil potencies also varied considerably during this period (16.8 ± 16.3%). The increase in cannabis preparation potency is mainly due to the increase in the potency of nondomestic versus domestic samples.

Sustained exposure to 3,4-methylenedioxymethamphetamine induces the augmentation of exocytotic serotonin release in rat organotypic raphe slice cultures.

3,4-Methylenedioxymethamphetamine (MDMA) causes serotonin efflux via serotonin transporter. Recently, we have reported that sustained exposure to MDMA induced an augmentation of serotonin release in rat raphe serotonergic slice cultures. Here we investigated the mechanism of augmented serotonin release from the slice cultures. Sustained MDMA exposure had no effect on MDMA-induced serotonin efflux in the synaptosomal fraction, whereas either tetrodotoxin, calcium channel inhibitors, or AMPA-receptor antagonists significantly attenuated the augmented serotonin release. These results suggest that the increase in Ca(2)-dependent exocytotic serotonin release is mediated through activation of AMPA receptors and responsible for the sustained MDMA-induced augmentation of serotonin release.

Decreased cerebral cortical serotonin transporter binding in ecstasy users a positron emission tomography(11)CDASB and structural brain imaging study.

Animal data indicate that the recreational drug ecstasy (3,4-methylenedioxymethamphetamine) can damage brain serotonin neurons. However, human neuroimaging measurements of serotonin transporter binding, a serotonin neuron marker, remain contradictory, especially regarding brain areas affected and the possibility that structural brain differences might account for serotonin transporter binding changes has not been explored. We measured brain serotonin transporter binding using (11)C N,N-dimethyl-2-(2-amino-4-cyanophenylthio) benzylamine in 50 control subjects and in 49 chronic (mean 4 years) ecstasy users (typically one to two tablets bi-monthly) withdrawn from the drug (mean 45 days). A magnetic resonance image for positron emission tomography image co-registration and structural analyses was acquired. Hair toxicology confirmed group allocation but also indicated use of other psychoactive drugs in most users. Serotonin transporter binding in ecstasy users was significantly decreased throughout all cerebral cortices (range -19 to -46%) and hippocampus (-21%) and related to the extent of drug use (years, maximum dose), but was normal in basal ganglia and midbrain. Substantial overlap was observed between control and user values except for insular cortex, in which 51% of ecstasy user values fell below the lower limit of the control range. Voxel-based analyses confirmed a caudorostral gradient of cortical serotonin transporter binding loss with occipital cortex most severely affected. Magnetic resonance image measurement revealed no overall regional volume differences between groups however, a slight left-hemispheric biased cortical thinning was detected in methamphetamine-using ecstasy users. The serotonin transporter binding loss was not related to structural changes or partial volume effect, use of other stimulant drugs, blood testosterone or oestradiol levels, major serotonin transporter gene promoter polymorphisms, gender, psychiatric status, or self-reported hyperthermia or tolerance. The ecstasy group, although grossly behaviourally normal, reported subnormal mood and demonstrated generally modest deficits on some tests of attention, executive function and memory, with the latter associated with serotonin transporter decrease. Our findings suggest that the typicallow dose (one to two tabletssession) chronic ecstasy-polydrug user might display a highly selective mild to marked loss of serotonin transporter in cerebral cortexhippocampus in the range of that observed in Parkinsons disease, which is not gender-specific or completely accounted for by structural brain changes, recent use of other drugs (as assessed by hair analyses) or other potential confounds that we could address. The striking sparing of serotonin transporter-rich striatum (although possibly affected in heavier users) suggests that serotonergic neurons innervating cerebral cortex are more susceptible, for unknown reasons, to ecstasy than those innervating subcortical regions and that behavioural problems in some ecstasy users during abstinence might be related to serotonin transporter changes limited to cortical regions.

Detection of methamphetamine, methylenedioxymethamphetamine, and 3,4-methylenedioxy-N-ethylamphetamine in spiked plasma by HPLC and TLC.

HPLC and TLC methods were developed for separation and detection of some amphetamine analogs methamphetamine (MA) 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) and 3,4-methylenedioxy-N-ethylamphetamine (MDEA) in spiked plasma samples. The methods are based on purple chromogens formed by displacement reaction of these secondary aliphatic amine-bearing drugs with 7,7,8,8-tetracyanoquinodimethane at 80 degrees C for 25 min. For HPLC, both normal phase (silica gel) and RP (C18) columns were used. With the former, good detection limits in plasma were obtained with a 6 min run 70, 100, and 500 ngmL for MDMA, MA, and MDEA, respectively. For TLC, hexane-chloroform (1 9) and benzene-diethyl ether-petroleum ether (40-60 degrees)-acetonitrile-ethyl methyl ketone (2 3.5 3.5 0.5 0.5) were used as mobile phases for silica gel 60 TLC and cyano-bonded silica gel HPTLC plates, respectively. The former offered more sensitive results than the latter. Influence of evaporation steps on recovery and interferences for the HPLC and TLC methods were investigated. The developed methods are selective, simple, and easily applicable.

Reduced 3,4-methylenedioxymethamphetamine (MDMA, Ecstasy)-initiated oxidative DNA damage and neurodegeneration in prostaglandin H synthase-1 knockout mice.

The neurodegenerative potential of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) and underlying mechanisms are under debate. Here, we show that MDMA is a substrate for CNS prostaglandin H synthase (PHS)-catalyzed bioactivation to a free radical intermediate that causes reactive oxygen species (ROS) formation and neurodegenerative oxidative DNA damage. In vitro PHS-1-catalyzed bioactivation of MDMA stereoselectively produced free radical intermediate formation and oxidative DNA damage that was blocked by the PHS inhibitor eicosatetraynoic acid. In vivo, MDMA stereoselectively caused gender-independent DNA oxidation and dopaminergic nerve terminal degeneration in several brain regions, dependent on regional PHS-1 levels. Conversely, MDMA-initiated striatal DNA oxidation, nerve terminal degeneration, and motor coordination deficits were reduced in PHS-1 - and -- knockout mice in a gene dose-dependent fashion. These results confirm the neurodegenerative potential of MDMA and provide the first direct evidence for a novel molecular mechanism involving PHS-catalyzed formation of a neurotoxic MDMA free radical intermediate.

Endocannabinoids inhibit the growth of free-living amoebae.

The cannabinoid Delta(9)-tetrahydrocannabinol inhibits the growth of some pathogenic amoebae in vitro and exacerbates amoebic encephalitis in animal models. However, the effects of endogenous cannabinoids on amoebae remain unknown. Therefore, we tested several endocannabinoids (N-acyl ethanolamines and 2-O-acyl glycerol) on different genera of amoebae. The results showed that all of the endocannabinoids tested inhibit amoebic growth at subpharmacological doses, with 50% inhibitory concentrations ranging from 15 to 20 microM. A nonhydrolyzable endocannabinoid had similar effects, showing that the inhibition seen results from endocannabinoids per se rather than from a catabolic product.

Delayed pre-conditioning by 3-nitropropionic acid prevents 3,4-methylenedioxymetamphetamine-induced 5-HT deficits.

The aim of the present study was to investigate whether late pre-conditioning using 3-nitropropionic acid (3NP) prevents the 5-hydroxytryptamine (5-HT) deficits caused by the amphetamine derivative 3,4-methylenedioxymethamphetamine (MDMA) in the rat. For this purpose we administered 3NP 24 h before MDMA (3 x 5 mgkg i.p., every 2 h) and rats were killed 7 days later. Pre-treatment of 3NP afforded complete protection against MDMA-induced 5-HT deficits independent of any effect on MDMA-induced hyperthermia or 5-HT transporter activity. To identify the transductional mechanisms responsible for the neuroprotective effect of 3NP, we first examined the involvement of nitric oxide (NO) by using selective inhibitors of all three nitric oxide synthase isoforms. Inhibition of endothelial and neuronal nitric oxide synthase, but not inducible nitric oxide synthase, reversed 3NP-induced pre-conditioning. The NO donor S-Nitroso-N-acetylpenicilamine mimicked 3NP effects further suggesting the involvement of NO in mediating 3NP protection. To investigate the involvement of NOSsoluble guanylate cyclase (sGC)protein kinase Gmitochondrial ATP-sensitive potassium channels (mitoK(ATP)) signaling pathway we examined the effect of 5-hydroxydecanoate (5-HD), a selective mitoK(ATP) blocker, and 1H-(1,2,4)oxadiazolo4,3-aquinoxaline-1-one, a potent inhibitor of sGC, on 3NP-induced tolerance. 5-hydroxydecanoate, but not 1H-(1,2,4)oxadiazolo4,3-aquinoxaline-1-one, suppressed 3NP-mediated protection suggesting that mitoK(ATP) opening, but not NO-mediated activation of sGC, participates in the mechanism underlying tolerance to MDMA. Our data also showed that the protective effect of 3NP was abolished by cycloheximide, supporting the involvement of de novo protein synthesis. In conclusion, 3NP-induced delayed tolerance against 5-HT deficits caused by MDMA occurs via NO production.

Tolerance to 3,4-methylenedioxymethamphetamine is associated with impaired serotonin release.

Tolerance to the behavioural effects of 3,4-methylenedioxymethamphetamine (MDMA) following high dose exposure has been attributed to alterations in serotonergic systems. The present study aimed to determine whether decreased 5-HT release andor 5-HT(2AC) receptor desensitization might play a role in tolerance by measuring the response to selective ligands following MDMA exposure. To this end, the latency to nose poke and emerge from a hide box to an open field arena following administration of various ligands to MDMA pre-treated and control rats was measured. Acute exposure to MDMA (0.0-3.3 mgkg), the 5-HT releasing stimulant fenfluramine (0.0-2.0 mgkg) and the 5-HT(2) receptor agonist m-CPP (0.0-1.25 mgkg) increased nose poke and emergence latency. Following administration of doses that produce 5-HT(2A) receptor-mediated behaviours, the 5-HT(2) receptor agonist (-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane failed to alter nose poke and emergence latency, suggesting a limited role of this receptor subtype in these behaviours. Activation of 5-HT(2C) receptors was implicated in the behavioural response to both MDMA and m-CPP since the increased emergence latency was dose-dependently attenuated by pre-treatment with the selective 5-HT(2C) receptor antagonist RS102221 (0.0-1.0 mgkg). Tolerance to the behavioural effect of MDMA and fenfluramine but not m-CPP was produced by prior exposure to MDMA (10 mgkg administered at two-hour intervals, total 40 mgkg), and tissue levels of 5-HT and 5-HIAA were decreased. These findings suggest that tolerance to the increased nose poke and emergence latency produced by MDMA is due to impaired 5-HT release.

Ecstasy (MDMA)-addicted subjects show increased serum levels of brain-derived neurotrophic factor, independently from a rise of drug-induced psychotic symptoms.

The recreational drug ecstasy3,4-methylenedioxymethamphetamine (MDMA) exerts a potent action on central serotonergic and dopaminergic neurons. These neurons utilize neurotrophins for their survival and function. In order to explore MDMA effects on neurotrophins, we measured by enzyme-linked immunosorbent assay the serum levels of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in ecstasy-addicted, ecstasy-addicted with signs of psychosis and healthy subjects. We found that BDNF serum levels were significantly increased in both groups of ecstasy-addicted as compared with healthy subjects, supporting the hypothesis that BDNF is involved in MDMA action.

Activity-based anorexia in C57BL6 mice effects of the phytocannabinoid, Delta9-tetrahydrocannabinol (THC) and the anandamide analogue, OMDM-2.

The activity-based anorexia (ABA) paradigm is one of the few animal models of human anorexia nervosa. We present here the translation of this approach to C57BL6 mice, a common background for genetically modified mice, and investigate the effects of the cannabinoid agonist, Delta(9)-tetrahydrocannabinol (THC) and the endocannabinoid uptake inhibitor, OMDM-2 in this model. The ABA paradigm was optimised so that food-restricted wheel-running mice displayed anorexia, reduced body weight and disrupted activity and circadian cycles. These conditions produced a murine ABA model with a defined stage and stability to allow for pharmacological intervention. Daily Delta(9)-THC (0.5 mgkg) decreased survival in the ABA animals but increased feeding in the survivors, OMDM-2 (3 mgkg) increased food intake, but not sufficiently to reverse weight loss. The effects of this model on endocannabinoid tone in the brain remain to be determined. Since the endocannabinoid system may be implicated in anorexia nervosa and in view of the positive modulation by cannabinoids of some aspects of ABA in this study, further investigation of the effects of cannabinoids in ABA is warranted.

Modulation of adipocyte biology by δ(9)-tetrahydrocannabinol.

It is recognized that the endocannabinoid system (ECS) plays a crucial role in the modulation of food intake and other aspects of energy metabolism. In this study, we aimed to investigate the effects of Δ(9)-tetrahydrocannabinol (THC) on adipocyte biology. 3T3-L1 cells were used to evaluate proliferation by sulforhodamine B (SRB) staining and methyl-(3)H-thymidine incorporation after 48 or 72 h of treatment with THC (1-500 nmoll). Cells were differentiated in the presence or absence of the cannabinoid, and adipogenesis was determined by measuring lipid accumulation and peroxisome proliferator-activated receptor γ (PPARγ) transcription through reverse transcriptase-PCR (RT-PCR). Lipolysis was quantified under basal conditions or after isoproterenol (IP, 100 nmoll) or insulin (INS, 100 nmoll) treatment. Transforming growth factor β (TGFβ), diacylglycerol lipase α, and N-acylphosphatidylethanolamine-specific phospholipase D (NAPE-PLD) transcriptions were determined by RT-PCR in preadipocytes and adipocytes and adiponectin only in adipocytes. THC treatment increased culture protein content and reduced methyl-(3)H-thymidine incorporation. Cells treated with THC underwent adipogenesis shown by the expression of PPARγ and had increased lipid accumulation. Basal and IP-stimulated lipolyses were inhibited by THC and there was no effect on lipolysis of INS-treated adipocytes. The effects on methyl-(3)H-thymidine incorporation and lipolysis seem to be mediated through CB1- and CB2-dependent pathways. THC decreased NAPE-PLD in preadipocytes and increased adiponectin and TGFβ transcription in adipocytes. These results show that the ECS interferes with adipocyte biology and may contribute to adipose tissue (AT) remodeling. Although these observations point toward increased AT deposition, the stimulation of adiponectin production and inhibition of lipolysis may be in favor of improved INS sensitivity under cannabinoid influence.

Endocrine and neurochemical effects of 3,4-methylenedioxymethamphetamine and its stereoisomers in rhesus monkeys.

3,4-Methylenedioxymethamphetamine (MDMA) is an amphetamine derivative that elicits complex biological effects in humans. One plausible mechanism for this phenomenon is that racemic MDMA is composed of two stereoisomers that exhibit qualitatively different pharmacological effects. In support of this, studies have shown that R(-)-MDMA tends to have hallucinogen-like effects, whereas S()-MDMA tends to have psychomotor stimulant-like effects. However, relatively little is known about whether these stereoisomers engender different endocrine and neurochemical effects. In the present study, the endocrine and neurochemical effects of each stereoisomer and the racemate were assessed in four rhesus monkeys after intravenous delivery at doses (1-3 mgkg) that approximated voluntary self-administration by rhesus monkeys and human recreational users. Specifically, fluorescence-based enzyme-linked immunosorbent assay was used to assess plasma prolactin concentrations, and in vivo microdialysis was used to assess extracellular dopamine and serotonin concentrations in the dorsal striatum. R(-)-MDMA, but not S()-MDMA, significantly increased plasma prolactin levels and the effects of S,R(-)-MDMA were intermediate to each of its component stereoisomers. Although S()-MDMA did not alter prolactin levels, it did significantly increase extracellular serotonin concentrations. In addition, S()-MDMA, but not R(-)-MDMA, significantly increased dopamine concentrations. Furthermore, as in the prolactin experiment, the effects of the racemate were intermediate to each of the stereoisomers. These studies demonstrate the stereoisomers of MDMA engender qualitatively different endocrine and neurochemical effects, strengthening the inference that differences in these stereoisomers might be the mechanism producing the complex biological effects of the racemic mixture of MDMA in humans.

A high-throughput respirometric assay for mitochondrial biogenesis and toxicity.

Mitochondria are a common target of toxicity for drugs and other chemicals and result in decreased aerobic metabolism and cell death. In contrast, mitochondrial biogenesis restores cell vitality, and there is a need for new agents to induce biogenesis. Current cell-based models of mitochondrial biogenesis or toxicity are inadequate because cultured cell lines are highly glycolytic with minimal aerobic metabolism and altered mitochondrial physiology. In addition, there are no high-throughput real-time assays that assess mitochondrial function. We adapted primary cultures of renal proximal tubular cells (RPTCs) that exhibit in vivo levels of aerobic metabolism, are not glycolytic, and retain higher levels of differentiated functions and used the Seahorse Bioscience analyzer to measure mitochondrial function in real time in multiwell plates. Using uncoupled respiration as a marker of electron transport chain (ETC) integrity, the nephrotoxicants cisplatin, HgCl(2), and gentamicin exhibited mitochondrial toxicity prior to decreases in basal respiration and cell death. Conversely, using FCCP (carbonylcyanide p-trifluoromethoxyphenylhydrazone)-uncoupled respiration as a marker of maximal ETC activity, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), SRT1720, resveratrol, daidzein, and metformin produced mitochondrial biogenesis in RPTCs. The merger of the RPTC model and multiwell respirometry results in a single high-throughput assay to measure mitochondrial biogenesis and toxicity and nephrotoxic potential.

Pharmacokinetic properties of delta9-tetrahydrocannabinol in oral fluid of occasional and chronic users.

Saliva is of continuing interest in detecting the influence of drugs while driving. Commercial tests are currently available where cannabis detectability is a major challenge. The present study aids in the understanding of tetrahydrocannabinol (THC) pharmacokinetics in oral fluid. The oral fluid analyses exhibited no significant differences between 12 occasional users and 12 chronic users smoking a standardized cannabis joint, except for the maximum concentrations in the first samples (occasional users, 397- 6438 ngg chronic users 387-71,747 ngg). THC was detectable in all samples with medians in the last samples (8 h) of 6.3 and 11.3 ngg in occasional and chronic users, respectively. The elimination half-life in both groups was 1.6 - 0.4 h. A series of samples was obtained over a period of 8 h without actual drug use representing a later elimination phase. Of these oral fluid samples, only 4.3% were negative for THC despite positive serum, and 24.1% of serum samples were negative despite positive oral fluid. This confirms that THC is detectable for longer in oral fluid than in serum. The oral fluid-to-serum ratios were 0.3 to 425 (median 16.5) with no difference between chronic and occasional users. The large inter- and intraindividual variability observed precludes a reliable estimation of THC serum concentrations from oral fluid data using this collection device.