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Prenatal MDMA exposure delays postnatal development in the rat a preliminary study.

3,4-methylenedioxymethamphetamine or MDMA (ecstasy) is a synthetic illicit drug which is widely consumed throughout the world. Drug abuse during pregnancy may have an impairing effect on the progeny of drug-abusing mothers. The purpose of the present study was to assess the effect of prenatal MDMA exposure on the progeny development, using a rat model. Pregnant animals were injected daily with MDMA (10 mgkg) between the 13th and 20th days of gestation. Male and female pups were then tested throughout the lactation period on the appearance and improvement of physical and sensory motor parameters. Appearance of some physical features (eyes opening and incisor eruption) and neurological reflexes as well as improving performances in negative geotaxis, gait and inclined board tests were delayed in pups prenatally exposed to MDMA compared to saline-treated pups. In contrast, functions that are necessary for survival such as forelimb reflex (that enables suckling) were present in both groups. At four weeks of age, MDMA animals recovered to normal level in all studied parameters. The delay in physical and neurological reflex development could be interpreted as alterations in maturation of some neuronal circuitries induced by prenatal MDMA exposure.

Interaction of 18-methoxycoronaridine with nicotinic acetylcholine receptors in different conformational states.

The interaction of 18-methoxycoronaridine (18-MC) with nicotinic acetylcholine receptors (AChRs) was compared with that for ibogaine and phencyclidine (PCP). The results established that 18-MC (a) is more potent than ibogaine and PCP inhibiting (-)-epibatidine-induced AChR Ca(2) influx. The potency of 18-MC is increased after longer pre-incubation periods, which is in agreement with the enhancement of (3)Hcytisine binding to resting but activatable Torpedo AChRs, (b) binds to a single site in the Torpedo AChR with high affinity and inhibits (3)HTCP binding to desensitized AChRs in a steric fashion, suggesting the existence of overlapping sites. This is supported by our docking results indicating that 18-MC interacts with a domain located between the serine (position 6) and valine (position 13) rings, and (c) inhibits (3)HTCP, (3)Hibogaine, and (3)H18-MC binding to desensitized AChRs with higher affinity compared to resting AChRs. This can be partially attributed to a slower dissociation rate from the desensitized AChR compared to that from the resting AChR. The enthalpic contribution is more important than the entropic contribution when 18-MC binds to the desensitized AChR compared to that for the resting AChR, and vice versa. Ibogaine analogs inhibit the AChR by interacting with a luminal domain that is shared with PCP, and by inducing desensitization.

Direct quantification of 11-nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid in urine by liquid chromatographytandem mass spectrometry in relation to doping control analysis.

An accurate and precise method for the quantification of 11-nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid (THCA) in urine by liquid chromatographytandem mass spectrometry (LCMSMS) for doping analysis purposes has been developed. The method involves the use of only 200 microL of urine and the use of D(9)-THCA as internal standard. No extraction procedure is used. The urine samples are hydrolysed using sodium hydroxide and diluted with a mixture of methanolglacial acetic acid (11). Chromatographic separation is achieved using a C8 column with gradient elution. All MS and MSMS parameters were optimised in both positive and negative electrospray ionisation modes. For the identification and the quantification of THCA three product ions are monitored in both ionisation modes. The method is linear over the studied range (5-40 ngmL), with satisfactory intra-and inter-assay precision, and the relative standard deviations (RSDs) are lower than 15%. Good accuracy is achieved with bias less than 10% at all levels tested. No significant matrix effects are observed. The selectivity and specificity are satisfactory, and no interferences are detected. The LCMSMS method was applied for the analysis of 48 real urine samples previously analysed with a routine gas chromatographymass spectrometry (GCMS) method. A good correlation between the two methods was obtained (r(2) > 0.98) with a slope close to 1.

Determination of a new designer drug, N-hydroxy-3,4-methylenedioxymethamphetamine and its metabolites in rats using ultra-performance liquid chromatography-tandem mass spectrometry.

An N-hydroxy analogue of 3,4-methylendioxymethamphetamine (MDMA), N-hydroxy MDMA (N-OH MDMA), has recently been distributed as a new designer drug in some drug markets. Very little data is available to the metabolic and pharmacological properties of N-OH MDMA, although it has been reported that the N-demethyl analogue, N-hydroxy-3,4-methylenedioxyamphetamine (N-OH MDA), is mainly metabolized to MDA in rats. In this study, an analytical method for the determination of N-OH MDMA and its metabolites in biological samples was developed, and the metabolic properties of N-OH MDMA in rats were investigated. After the i.p. administration of N-OH MDMA to pigmented hairy rats (5mgkgday, 10 days), N-OH MDMA and its N-dehydroxy and N-demethyl metabolites (MDMA, N-OH MDA and MDA) in rat plasma, urine and hair samples were determined by ultra-performance LC (UPLC)-MSMS. The hair sample was extracted by 1-h sonication and overnight soaking in 5M hydrochloric acid-methanol (120). The plasma, urine, and hair extract samples were purified using a solid-phase extraction procedure. N-OH MDMA in the samples could be precisely analyzed by avoiding an alkaline environment. The parent compound very rapidly disappeared from the rat plasma (<15min) and urine (<10h), and most of the N-OH MDMA was excreted in the rat urine as MDMA and MDA in 72h. In the rat hair samples collected 4 weeks after the first administration, N-OH MDMA (0.03ngmg) and N-OH MDA (0.13ngmg) were clearly detected as well as MDMA (149ngmg) and MDA (52ngmg). This analytical method will be useful for the analysis of N-OH MDMA and its metabolites in biological samples.

MDMA causes a redistribution of serotonin transporter from the cell surface to the intracellular compartment by a mechanism independent of phospho-p38-mitogen activated protein kinase activation.

3,4-methylenedioxymethamphetamine (MDMA) causes long-term serotonin depletion and reduced serotonin transporter (SERT) function in humans and in animal models. Using quantitative Western blotting and real-time PCR, we have shown that total SERT protein in the striatum and nucleus accumbens and mRNA levels in the dorsal raphe nucleus were not significantly changed following MDMA exposure in rats (4 x 2 h i.p. injections, 10 mgkg each). In mouse neuroblastoma (N(2)A) cells transiently expressing green fluorescent protein-tagged human SERT (GFP-hSERT), we have shown redistribution of SERT from the cell surface to intracellular vesicles on exposure to MDMA using cell surface biotinylation, total internal reflection fluorescence microscopy (TIRFM) and live-cell confocal microscopy. To investigate the mechanism responsible for SERT redistribution, we used specific antibodies to phospho-p38-mitogen activated protein kinase (p38 MAPK), a known signalling pathway involved in SERT membrane expression. We found that p38 MAPK activation was not involved in the MDMA-induced redistribution of SERT from the cell-surface to the cell interior. A loss of SERT from the cell surface on acute exposure to MDMA may contribute to the decreased SERT function seen in rats exposed to MDMA.

Phencyclidine withdrawal disrupts episodic-like memory in rats reversal by donepezil but not clozapine.

Episodic memory is the capacity to recall an event in time and place (What Where When). Impaired episodic memory is a debilitating cognitive symptom in schizophrenia but is poorly controlled by currently available antipsychotic drugs. Consistent with glutamatergic abnormality in schizophrenia, the NDMA receptor antagonist, phencyclidine (PCP), induces persistent schizophrenia-like symptoms including memory deficits in humans and rodents and is widely used as an animal model of the disorder. However, in contrast to humans, PCP and PCP withdrawal-induced memory deficits in rodents are reversed by antipsychotic drugs such as clozapine. One possible explanation is that the memory tasks used in animal studies do not simultaneously test the What Where When components that characterize episodic memory in human tasks. We investigated whether subchronic PCP withdrawal disrupts memory in rats in a task that requires simultaneous integration of memory for object, place and context. Rats learn to discriminate objects under specific spatial and contextual conditions analogous to the What Where When components of human episodic memory. We found that PCP withdrawal impaired performance on this task and that the atypical antipsychotic drug clozapine did not reverse this impairment. However the acetylcholinesterase inhibitor (AChEI) donepezil, which has been shown to improve episodic memory in humans did reverse the effect of PCP. This suggests that PCP withdrawal disruption of object-place-context recognition in rats may prove to be a useful model to investigate episodic memory impairment in schizophrenia and supports the suggestion that AChEIs could prove to be a useful pharmacological strategy to specifically treat episodic memory problems in schizophrenia.

Cannabis and Ecstasy MDMA empirical measures of creativity in recreational users.

This study investigated the associations between chronic cannabis and EcstasyMDMA use and one objective and two subjective measure of creativity. Fifteen abstinent Ecstasy users, 15 abstinent cannabis users, and 15 nondrug-user controls, completed three measures of creativity the Consequences behavioral test of creativity, self-assessed performance on the Consequences test, and Goughs Trait Self-Report Creative Adjective Checklist. The Consequences test involved five scenarios where possible consequences had to be devised scoring was conducted by the standard blind rating (by two independent judges) for remoteness and rarity, and by a frequency and rarity of responses method. Cannabis users had significantly more rare-creative responses than controls (Tukey, p < 0.05) this effect remained significant with gender as a covariate. There were no significant differences between the groups on the number of standard scoring remote-creative ideas or for fluency of responses. On self-rated creativity, there was a significant ANOVA group difference (p < 0.05), with Ecstasy users tending to rate their answers as more creative than controls (Tukey comparison p 0.058, two-tailed). Ecstasy users did not differ from controls on the behavioral measures of creativity, although there was a borderline trend for self-assessment of greater creativity. Cannabis users produced significantly more rare-creative responses, but did not rate themselves as more creative.

Alter ego representations in San Agustin monolithic sculptures possible plant hallucinogenic influences.

This article examines the evidence for plant hallucinogenic use (possibly Brugmansia, Brunfelsia chiricaspi, Desfontainia R., Anadenanthera peregrina, Banisteriopsis sps, Psychotropia viridis and Virola theidora) by the San Agustin culture, an extinct peoples who resided in the Magdelena River area of Colombia from the third century B.C. until the sixteenth century A.D. Based on thematic materials gathered from a cross-cultural survey of plant hallucinogens, the author examines themes in the monolithic sculptures of this culture in light of man-animal transformations and shamanic themes linked to plant hallucinogenic ingestion.

Experiences of encounters with ayahuasca--the vine of the soul.

Ayahuasca is a psychoactive brew used by the indigenous populations of the Amazon. The aim of this qualitative study was to gain insight into the experiences of western users of ayahuasca, as well as to ascertain the experienced meaning that participants felt by their participation. Twenty-five people from Northern Europe with experiences of group sessions with ayahuasca wrote anonymous descriptions of their experiences. The Empirical Phenomenological Psychological method was used for this analysis. The analysis resulted in 33 categories which were assembled into six general themes (a) motivation and aim, (b) contractile frightening state (c) sudden transformation of the experience, (d) limitless expansive states with transcendental experiences, (f) reflections, and (g) changed worldview and new orientation to life. These themes provided a new structure, called the transcendental circle. Participants reported many positive psychological and physical improvements that indicate that ayahuasca could be of potential interest in the development of new medicines and therapies.

3,4-methylenedioxyamfetamine (ecstasy) use reduces cognition.

3,4-methylenedioxyamfetamine (MDMA, ecstasy) use reduces cognition by reducing levels of dopamine and serotonin in the central nervous system. This results in dose-related cognition impairment, particularly in complex cognitive skills, as well as causing disorders such as mood changes, hallucinations, altered perception amd memory loss. MDMA reduces the level of these neurotransmitters within the neural synapses by reducing the number of intraneural transporters to the synaptic clefts, increasing deactivation with the synapse and or increasing degradation with the pre- and postsynaptic neurons. Users may have varied reasons for MDMA use and therefore require help and support from their families or friends, and knowledgeable and well-skilled healthcare professionals for successful abstinence, avoidance of further psychological damage and a reversal of adverse health effects or reduction in their severity.

Pharmacological characterization of cannabinoid receptor activity in the rat-isolated ileum myenteric plexus-longitudinal muscle preparation.

Cannabinoid effects on intestinal transit are commonly evaluated in rats. We characterized the cannabinoid receptors mediating the inhibitory effect of 5-(1,1-dimethylheptyl)-2-5-hydroxy-2-(3-hydroxypropyl)-cyclohexyl-phenol (CP 55,940), (R)-()-2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo1,2,3-de-1,4-benzoxazin-6-yl-1-naphthalenylmethanone mesylate (WIN 55,212-2), arachidonylethanolamide (AEA) and Delta(9)-tetrahydrocannabinol (Delta(9)-THC) on contractions of the rat ileum myenteric plexus-longitudinal muscle (MPLM) preparation. The interaction of each agonist was examined with the CB(1) and CB(2) receptor antagonist rimonabant and SR 144,528 respectively, on contractions elicited by electrical field stimulation (EFS) or exogenous ACh. The interaction of AEA with capsazepine, a TRPV(1) receptor antagonist, was also investigated. EFS with single and trains of pulses evoked neurogenic ACh-mediated twitch and rebound contractions respectively. The rank order of potency for inhibition was CP 55,940 WIN 55,212-2 > AEA > Delta(9)-THC and AEA > WIN 55,212-2 Delta(9)-THC CP 55,940 respectively. The stereoisomer WIN 55,212-3 was without effect. Rimonabant antagonized the inhibition of the twitches with pK(B) values of around 8.60, but only antagonized rebound contractions induced by WIN 55,212-2, AEA and Delta(9)-THC, with pA(2) values of around 6.80. Rimonabant increased the twitches but inhibited the rebound contractions. Contractions to exogenous ACh were not altered. These observations extended to the guinea pig ileum MPLM. The rat MPLM contains CB(1) receptors and at least two non-CB(1)-non-CB(2)-non-TRPV(1) receptors attenuating EFS-evoked ACh-mediated contractions in an EFS frequency-dependent pre-synaptic and stereo-specific manner. Augmentation of the twitches by rimonabant may be through antagonism of an endocannabinoid tone or inverse agonism, whereas inhibition of the rebound contractions involved partial agonism.

The effects of kappa-opioid receptor ligands on prepulse inhibition and CRF-induced prepulse inhibition deficits in the rat.

Kappa-opioid receptor (KOR) agonists produce dysphoria and psychotomimesis in humans. KORs are enriched in the prefrontal cortex and other brain regions that regulate mood and cognitive function. Dysregulation of the dynorphinKOR system has been implicated in the pathogenesis of schizophrenia, depression, and bipolar disorder. Prepulse inhibition of the acoustic startle reflex (PPI), a sensorimotor gating process, is disrupted in many psychiatric disorders. The present study determined whether KOR ligands alter PPI in rats. Utilizing a range of doses of the synthetic KOR agonists (-) U50,488, (-) U50,488, and U69,593 and the naturally occurring KOR agonist, Salvinorin A, we demonstrate that KOR activation does not alter PPI or startle reactivity in rats. Similarly, selective KOR blockade using the long-acting antagonist nor-binaltorphimine (nor-BNI) was without effect. In contrast to KOR ligands, MK-801 and quinpirole produced deficits in PPI. Stress and corticotropin-releasing factor (CRF) decrease PPI levels. The dynorphinKOR system has been suggested to be a key mediator of various behavioral effects produced by stress and CRF. We therefore examined the contribution of KORs to CRF-induced alterations in PPI. Intracerebroventricular infusion of CRF decreased PPI. Administration of nor-BNI failed to affect the CRF-evoked disruption in PPI. Together, these results provide no evidence of a link between the dynorphinKOR system and deficits in sensory gating processes. Additional studies, however, examining whether dysregulation of this opioid system contributes to cognitive deficits and other behavioral abnormalities associated with psychiatric disorders are warranted.

Toxicity of ecstasy (MDMA) towards embryonic stem cell-derived cardiac and neural cells.

Ecstasy or methylenedioxymethamphetamine (MDMA) is primarily a recreational drug commonly used during the child bearing period, thus, there is a major concern regarding the embryonic and fetal toxicity of this drug. Here, we report the cardio- and neuro-toxic effects of MDMA on beating embryoid bodies (EBs) and neural cell-containing EBs derived from mouse embryonic stem cell (ESCs). Based on our linear discriminate function, MDMA is considered to be a moderate or weak teratogen. Moreover, the generation of EBs with neural cell morphology and the expression of MAP2, a mature neuron marker, decrease more when MDMA is administered during the EB formation stage rather than post-plated EBs. In addition, the ID50 (inhibition of differentiation) of EBs with neural cell morphology is less than cardiac cells. In conclusion, MDMA causes a marked reduction in beating cardiomyocytes and neurons in ESC cultures, and this drug has a more potent toxicity on neural rather than cardiac cell differentiation.

Induction of p53-independent apoptosis by a novel synthetic hexahydrocannabinol analog is mediated via Sp1-dependent NSAID-activated gene-1 in colon cancer cells.

Nonsteroidal anti-inflammatory drug (NSAID)-activated gene-1 (NAG-1) has received greater attention as a novel molecular target for anti-cancer therapeutics in recent years. We identified a novel synthetic hexahydrocannabinol analog, LYR-8 (1-((9S)-1-hydroxy-6,6,9-trimethyl-6a,7,8,9,10,10a-hexahydro-6H-benzocchromen-2-yl)ethanone), as a potent NAG-1 and apoptosis inducer in a panel of human cancer cells. LYR-8 did not possess any affinity for cannabinoid receptor CB(1) or CB(2), which eliminates the concern about potential psychoactive side effects. LYR-8 dramatically induced NAG-1 expression and apoptosis in HCT116 (wild-type p53) and HT29 (mutant p53) colon cancer cells. The NAG-1 expression by LYR-8 was not blocked by pifithrin-alpha, a specific p53 inhibitor, which was different from doxorubicin that induced p53-dependent NAG-1 transcriptional activity. The induction of NAG-1 promoter activity by LYR-8 was strongly correlated with increased Sp1 activation as noted in various luc-promoter activities. Furthermore, pretreatment with the specific Sp1 inhibitor mithramycin A completely reversed the LYR-8-induced NAG-1 expression in both HCT116 and HT29 cells. Knockdown of NAG-1 using siRNA significantly reversed LYR-8-induced cell death in both wild-type and mutant p53-expressing colon cancer cells. Furthermore, sensitization with NAG-1 inducer sulindac sulfide synergized LYR-8-induced cell death in both colon cancer cells. These results suggest that induction of NAG-1 via Sp1 activation is a promising therapeutic approach in cancer treatment, and that a novel compound like LYR-8 could be a potent chemotherapeutic agent for colon cancers including p53-mutated cancer.

Tocolytic effect of delta9-tetrahydrocannabinol in mice model of lipopolysaccharide--induced preterm delivery role of nitric oxide.

In this study, we explained that exogenous cannabinoid, Delta(9)-tetrahydrocannabinol (THC), has a preventive effect in a murine model of lipopolysaccharide (LPS)-induced preterm delivery and the contribution of nitric oxide (NO) pathway as a mechanism involved in this process. Preterm delivery was induced by double dose of 35 microgkg LPS with 3-hour interval on gestational day (gd) 15. Delta(9)-tetrahydrocannabinol was administered with (a) double dose (0.02, 0.05, 0.1, 0.5, 1, and 5 mgkg) 1 hour before each LPS injection, on gd 15 and (b) single administration (0.05, 0.1, and 0.5 mgkg,) on gds 13 and 14, and the double administration, 1 hour before each LPS injection. To assess the involved mechanism, either AM281 (CB1 receptor antagonist, 2 mgkg) and AM630 (CB2 receptor antagonist, 5 mgkg) or N(omega)-nitro-L-arginine methyl ester (L-NAME, 2 mgkg) was administered 1 hour before each THC injection on gds 13, 14, and 15. The main outcome measurement was the incidence of preterm delivery after injection of last LPS dose. Any interaction in the incidence and time of preterm delivery was ruled out by administration of AM281, AM630, or L-NAME alone. Chronic THC treatment (0.5 mgkg) significantly decreased the incidence of LPS-induced premature labor and increased the delivery time. Both AM281 and L-NAME reversed THC-induced attenuation of preterm delivery rate and pregnancy duration. Unlike AM281, AM630 did not influence the rate of preterm delivery in THC-treated mice. Delta(9)-Tetrahydrocannabinol contributes to the regulation of gestational duration in LPS-induced preterm delivery probably by NO coupling through the CB1 receptor.

A long hangover from party drugs residual proteomic changes in the hippocampus of rats 8 weeks after γ-hydroxybutyrate (GHB), 3,4-methylenedioxymethamphetamine (MDMA) or their combination.

3,4-Methylenedioxymethamphetamine (MDMA) and gamma-hydroxybutyrate (GHB) are popular party drugs that are used for their euphoric and prosocial effects, and sometimes in combination. Both drugs increase markers of oxidative stress in the hippocampus and can cause lasting impairments in hippocampal-dependent forms of memory. To gain further information on the biochemical mechanisms underlying these effects, the current study examined residual changes in hippocampal protein expression measured 8 weeks after chronic administration of GHB (500mgkg), MDMA (5mgkg) or their combination (GHBMDMA). The drugs were administered once a day for 10 days in an environment with an elevated ambient temperature of 28 degrees C. Results showed significant changes in protein expression, relative to controls, in all three groups MDMA and GHB given alone caused residual changes in 8 and 5 proteins respectively, while the GHBMDMA combination significantly changed 6 proteins. The altered proteins had roles in neuroplasticity, neuroprotection, intracellular signalling and cytoskeletal function. The largest change (-4.3-fold) was seen in the MDMA group with the protein C-crk a protein implicated in learning-related neuroplasticity. The second largest change (3.0-fold) was seen in the GHB group in Glutathione-S-transferase (GST), a protein that protects against oxidative stress. Two cytoskeletal proteins (Tubulin Folding Cofactor B and Tropomyosin-alpha-3 chain) and one plasticity related protein (Neuronal Pentraxin-1 NP1) were similarly changed in both the MDMA and the GHB groups, while two intracellular signalling proteins (alpha-soluble NSF-attachment protein and subunits of the V-type proton ATPase) were changed in both the MDMAGHB and the MDMA groups. These results provide some insight into the molecular pathways possibly underlying the lasting cognitive deficits arising from GHB andor MDMA use.

Receptor antagonist of NMDA and animal models of schizophrenia.

Schizophrenia is one of the common mental diseases. Because the mechanism of the schizophrenia is significantly complicated, the cause is still unknown. N-methyl-D-aspartate receptor antagonist can simulate the positive and negative symptoms, as well as the cognitive disorder of schizophrenia. Thus it has been widely used to establish the animal models of schizophrenia. The relationship of the three blocking agents of ion channels (phencyclidine, MK-801, ketamine) and the establishment of schizophrenia animal models is reviewed in this article.

Composition, standardization and chemical profiling of Banisteriopsis caapi, a plant for the treatment of neurodegenerative disorders relevant to Parkinsons disease.

Banisteriopsis caapi, a woody vine from the Amazonian basin, is popularly known as an ingredient of a sacred drink ayahuasca, widely used throughout the Amazon as a medicinal tea for healing and spiritual exploration. The usefulness of Banisteriopsis caapi has been established for alleviating symptoms of neurological disorders including Parkinsons disease. Primary objective of this study was to develop the process for preparing standardized extracts of Banisteriopsis caapi to achieve high potency for inhibition of human monoamine oxidases (MAO) and antioxidant properties. The aqueous extracts prepared from different parts of the plant collected from different geographical locations and seasons were analyzed by HPLC for principal bioactive markers. The extracts were simultaneously tested in vitro for inhibition of human MAOs and antioxidant activity for analysis of correlation between phytochemical composition of the extracts and bioactivities. Reversed-phase HPLC with photodiode array detection was employed to profile the alkaloidal and non-alkaloidal components of the aqueous extract of Banisteriopsis caapi. The Banisteriopsis caapi extracts and standardized compositions were tested in vitro for inhibition of recombinant preparations of human MAO-A and MAO-B. In vitro cell-based assays were employed for evaluation of antioxidant property and mammalian cell cytotoxicity of these preparations. Among the different aerial parts, leaves, stemslarge branches and stem bark of Banisteriopsis caapi, HPLC analysis revealed that most of the dominant chemical and bioactive markers (1, 2, 5, 7-9) were present in high concentrations in dried bark of large branch. A library of HPLC chromatograms has also been generated as a tool for fingerprinting and authentication of the studied Banisteriopsis caapi species. The correlation between potency of MAO inhibition and antioxidant activity with the content of the main active constituents of the aqueous Banisteriopsis caapi extracts and standardized compositions was established. Phytochemical analysis of regularcommercial Banisteriopsis caapi dried stems, obtained from different sources, showed a similar qualitative HPLC profile, but relatively low content of dominant markers 1, 2, 7, and 9, which led to decreased MAO inhibitory and antioxidant potency compared to Banisteriopsis caapi Da Vine. The ethnopharmacological use of bark of matured stemlarge branch of Banisteriopsis caapi as well as whole matured stem is supported by the results obtained in this investigation. Among various constituents of Banisteriopsis caapi, harmine (7), harmaline (6) and tetrahydroharmine (5) are responsible for MAO-A inhibition, while two major proanthocyanidines, epicatechin (8) and procyanidine B2 (9) produce antioxidant effects. The compounds 1-9 can serve as reliable markers for identification and standardization of Banisteriopsis caapi aerial parts, collected in different seasons andor from different geographical regions.

A new digitized method of the compulsive gnawing test revealed dopaminergic activity of salvinorin A in vivo.

The compulsive gnawing (CG) test has been used for numerous years as an assay to determine the dopaminergic activity of various compounds. We developed a new method of quantification via a digitization step which allowed a more precise measurement of the gnawing activity. It was the aim of the present study to explore possible dopaminergic effects of salvinorin A (SA), the major active compound of Salvia divinorum, using the new digitized CG test. A group of experiments using male C57BL6 mice were performed to validate the new method of quantification showing only significant increases of gnawing when the dopamine reuptake inhibitors buproprion (20 mgkg, p.0.) and nomifensine (10 mgkg, i.p.) were given concomitantly with apomorphine (10 mgkg, i.p.). Different concentrations of the SA (1.0, 2.5, 5, and 10 mgkg, i.p.) were tested with positive dopaminergic activity when administered with apomorphine which differed from the semisynthetic counterpart U-69593. Furthermore, the activity observed with SA was unsuccessfully antagonized by the κ-opioid receptor antagonist norbinaltorphimine (NorBNI 10 and 20 mgkg, i.p.), while the dopamine antagonist haloperidol did successfully block (0.06 mgkg, i.p.) the gnawing activity seen with SA. Our data further strengthen the argument that salvinorin A is not a selective κ-opioid receptor agonist and is the first in vivo study that veers from salvinorin A acting solely like its synthetic counterparts. Furthermore, the digitized CG test system used in this study provides a new computational method to accurately detect behavior associated with dopaminergic neurotransmission.

Prevalence and risk factors of ecstasy use among college students in Astara, Islamic Republic of Iran.

We determined the prevalence and risk factors for 3,4-methylenedioxy-methamphetamine (MDMA, ecstasy) use among college students in Astara, a northern border city of Iran. In a cross-sectional questionnaire survey of 1226 students, the lifetime prevalence of ecstasy use was 5.6%. The lifetime prevalence of use of other drugs, mostly cannabis and opium, was 4.6%. A fifth of students (21.8%) were current cigarette smokers and 24.8% had ever used alcohol. After logistic regression, the factors influencing ever use of ecstasy were ever use of other drugs, ever use of alcohol, current cigarette smoking and living alone or with friends. Targeted prevention programmes should be conducted in all colleges.

Simultaneous screening for and determination of 128 date-rape drugs in urine by gas chromatography-electron ionization-mass spectrometry.

Date-rape drugs (DRDs) are used for the purpose of drugging unsuspected victims and raping or robbing them while under the influence of the drug. The wide variety of substances used for criminal purposes, their low concentrations in body fluids and, often, a long time delay between the event and clinical examination make comprehensive screening analysis of biological materials collected from crime victims for the presence of these drugs very difficult. Detection of a drug used to facilitate sexual assault in biological fluids can be very important evidence of a committed crime. The purpose of this study was to develop a simple GC-EI-MS screening procedure for date-rape drugs in urine. Target analytes were isolated by solid-phase extraction. 2-mL urine samples were extracted and then derivatized by using BSTFA1%TMCS reagent. Detection of all compounds was based on full-scan mass spectra and for each compound one ion was chosen for further quantification. The method allowed the simultaneous screening, detection and quantification of 128 compounds from different groups (number of compounds) opioids (20), amphetamines (11), GHB and related products (3), hallucinogens (9), benzodiazepines (18), antihistamines (9), antidepressants (14), selective serotonin-reuptake inhibitors (4), antipsychotics (7), barbiturates (7), other sedatives (5), muscle relaxants (2) and other drugs (19). The procedure can easily be expanded to encompass more substances. The developed method appeared to be suitable for screening for the target DRDs. The procedure was successfully applied to the analysis of authentic urine samples collected from victims of rapes and other crimes in routine casework.

The measurement of ecstasy in human hair by triple phase directly suspended droplet microextraction prior to HPLC-DAD analysis.

New pre-concentration technique, triple phase suspended droplet microextraction (SD-LPME) and liquid chromatography-photodiode array detection was applied to determine ecstasy, MDMA (3,4-methylendioxy-N-methylamphetamine) in hair samples. In this research MDMA in hair was digested and after treatment extracted. The effective parameters were investigated and method was evaluated. Under the optimal conditions, the MDMA was enriched by factor 98.11. Linearity (r0.9921), was obtained in the range of 10-15,000 ng mL(-1) and detection limit was 0.1 ng mL(-1).

Effects of monoamine oxidase inhibitor and cytochrome P450 2D6 status on 5-methoxy-N,N-dimethyltryptamine metabolism and pharmacokinetics.

5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is a natural psychoactive indolealkylamine drug that has been used for recreational purpose. Our previous study revealed that polymorphic cytochrome P450 2D6 (CYP2D6) catalyzed 5-MeO-DMT O-demethylation to produce active metabolite bufotenine, while 5-MeO-DMT is mainly inactivated through deamination pathway mediated by monoamine oxidase (MAO). This study, therefore, aimed to investigate the impact of CYP2D6 genotypephenotype status and MAO inhibitor (MAOI) on 5-MeO-DMT metabolism and pharmacokinetics. Enzyme kinetic studies using recombinant CYP2D6 allelic isozymes showed that CYP2D6.2 and CYP2D6.10 exhibited 2.6- and 40-fold lower catalytic efficiency (V(max)K(m)), respectively, in producing bufotenine from 5-MeO-DMT, compared with wild-type CYP2D6.1. When co-incubated with MAOI pargyline, 5-MeO-DMT O-demethylation in 10 human liver microsomes showed significantly strong correlation with bufuralol 1-hydroxylase activities (R(2)0.98 P<0.0001) and CYP2D6 contents (R(2)0.77 P0.0007), whereas no appreciable correlations with enzymatic activities of other P450 enzymes. Furthermore, concurrent MAOI harmaline sharply reduced 5-MeO-DMT depletion and increased bufotenine formation in human CYP2D6 extensive metabolizer hepatocytes. In vivo studies in wild-type and CYP2D6-humanized (Tg-CYP2D6) mouse models showed that Tg-CYP2D6 mice receiving the same dose of 5-MeO-DMT (20mgkg, i.p.) had 60% higher systemic exposure to metabolite bufotenine. In addition, pretreatment of harmaline (5mgkg, i.p.) led to 3.6- and 4.4-fold higher systemic exposure to 5-MeO-DMT (2mgkg, i.p.), and 9.9- and 6.1-fold higher systemic exposure to bufotenine in Tg-CYP2D6 and wild-type mice, respectively. These findings indicate that MAOI largely affects 5-MeO-DMT metabolism and pharmacokinetics, as well as bufotenine formation that is mediated by CYP2D6.

Dynamic planar solid phase microextraction-ion mobility spectrometry for rapid field air sampling and analysis of illicit drugs and explosives.

A preconcentration device that targets the volatile chemical signatures associated with illicit drugs and explosives (high and low) has been designed to fit in the inlet of an ion mobility spectrometer (IMS). This is the first reporting of a fast and sensitive method for dynamic sampling of large volumes of air using planar solid phase microextraction (PSPME) incorporating a high surface area for absorption of analytes onto a sol-gel polydimethylsiloxane (PDMS) coating for direct thermal desorption into an IMS. This device affords high extraction efficiencies due to strong retention properties at ambient temperature, resulting in the detection of analyte concentrations in the parts per trillion range when as low as 3.5 L of air are sampled over the course of 10 s (absolute mass detection of less than a nanogram). Dynamic PSPME was used to sample the headspace over the following 3,4-methylenedioxymethamphetamine (MDMA) tablets resulting in the detection of 12-40 ng of piperonal, high explosives (Pentolite) resulting in the detection of 0.6 ng of 2,4,6-trinitrotoluene (TNT), and low explosives (several smokeless powders) resulting in the detection of 26-35 ng of 2,4-dinitrotoluene (2,4-DNT) and 11-74 ng of diphenylamine (DPA).

Amphetamine toxicities classical and emerging mechanisms.

The drugs of abuse, methamphetamine and MDMA, produce long-term decreases in markers of biogenic amine neurotransmission. These decreases have been traditionally linked to nerve terminals and are evident in a variety of species, including rodents, nonhuman primates, and humans. Recent studies indicate that the damage produced by these drugs may be more widespread than originally believed. Changes indicative of damage to cell bodies of biogenic and nonbiogenic amine-containing neurons in several brain areas and endothelial cells that make up the blood-brain barrier have been reported. The processes that mediate this damage involve not only oxidative stress but also include excitotoxic mechanisms, neuroinflammation, the ubiquitin proteasome system, as well as mitochondrial and neurotrophic factor dysfunction. These mechanisms also underlie the toxicity associated with chronic stress and human immunodeficiency virus (HIV) infection, both of which have been shown to augment the toxicity to methamphetamine. Overall, multiple mechanisms are involved and interact to promote neurotoxicity to methamphetamine and MDMA. Moreover, the high coincidence of substituted amphetamine abuse by humans with HIV andor chronic stress exposure suggests a potential enhanced vulnerability of these individuals to the neurotoxic actions of the amphetamines.

Cannabinoid CB1 receptor antagonism prevents neurochemical and behavioural deficits induced by chronic phencyclidine.

Clinical and laboratory studies suggest that the endocannabinoid system is involved in schizophrenia disorders. Recent evidence indicates that cannabinoid receptor (CB1) antagonists have a pharmacological profile similar to antipsychotic drugs. We investigated the behavioural and biochemical effects of the CB1 antagonist AM251 in a phencyclidine (PCP) animal paradigm modelling the cognitive deficit and some negative symptoms of schizophrenia. Chronic AM251 (0.5 mgkg for 3 wk) improved the PCP-altered recognition memory, as indicated by a significant amelioration of the discrimination index compared to chronic PCP alone (2.58 mgkg for 1 month). AM251 also reversed the PCP-induced increase in immobility in the forced swim test resembling avolition, a negative sign of schizophrenia. In order to analyse the mechanisms underlying these behaviours, we studied the effects of AM251 on the endocannabinoid system (in terms of CB1 receptor density and functional activity and endocannabinoid levels) and c-Fos protein expression. The antagonist counteracted the alterations in CB1 receptor function induced by PCP in selected cerebral regions involved in schizophrenia. In addition, in the prefrontal cortex, the key region in the integration of cognitive and negative functions, AM251 markedly raised anandamide levels and reversed the PCP-induced increase of 2-arachidonoylglycerol concentrations. Finally, chronic AM251 fully reversed the PCP-elicited expression of c-Fos protein in the prefrontal cortical region. These findings suggest an antipsychotic-like profile of the CB1 cannabinoid receptor antagonist which, by restoring the function of the endocannabinoid system, might directly or indirectly normalize some of the neurochemical maladaptations present in this schizophrenia-like animal model.

Δ⁹-Tetrahydrocannabivarin suppresses in vitro epileptiform and in vivo seizure activity in adult rats.

We assessed the anticonvulsant potential of the phytocannabinoid Δ⁹-tetrahydrocannabivarin (Δ⁹-THCV) by investigating its effects in an in vitro piriform cortex (PC) brain slice model of epileptiform activity, on cannabinoid CB1 receptor radioligand-binding assays and in a generalized seizure model in rats. Δ⁹-THCV was applied before (10 μm Δ⁹-THCV) or during (10-50 μm Δ⁹-THCV) epileptiform activity induced by Mg²() -free extracellular media in adult rat PC slices and measured using multielectrode array (MEA) extracellular electrophysiologic techniques. The actions of Δ⁹-THCV on CB1 receptors were examined using ³HSR141716A competition binding and ³⁵SGTPγS assays in rat cortical membranes. Effects of Δ⁹-HCV (0.025-2.5 mgkg) on pentylenetetrazole (PTZ)-induced seizures in adult rats were also assessed. After induction of stable spontaneous epileptiform activity, acute Δ⁹ -THCV application (≥ 20 μm) significantly reduced burst complex incidence and the amplitude and frequency of paroxysmal depolarizing shifts (PDSs). Furthermore, slices pretreated with 10 μm Δ⁹-THCV prior to induction of epileptiform activity exhibited significantly reduced burst complex incidence and PDS peak amplitude. In radioligand-binding experiments, Δ⁹-THCV acted as a CB1 receptor ligand, displacing 0.5 nm ³HSR141716A with a Ki∼290 nm, but exerted no agonist stimulation of ³⁵SGTPγS binding. In PTZ-induced seizures in vivo, 0.25 mgkg Δ⁹-THCV significantly reduced seizure incidence. These data demonstrate that Δ⁹-THCV exerts antiepileptiform and anticonvulsant properties, actions that are consistent with a CB1 receptor-mediated mechanism and suggest possible therapeutic application in the treatment of pathophysiologic hyperexcitability states.

Differential effects of serotonin (5-HT)2 receptor-targeting ligands on locomotor responses to nicotine-repeated treatment.

We verified the hypothesis that serotonin (5-HT)(2) receptors control the locomotor effects of nicotine (0.4 mg kg(-1)) in rats by using the 5-HT(2A) receptor antagonist M100907, the preferential 5-HT(2A) receptor agonist DOI, the 5-HT(2C) receptor antagonist SB 242084, and the 5-HT(2C) receptor agonists Ro 60-0175 and WAY 163909. Repeated pairings of a test environment with nicotine for 5 days, on Day 10 significantly augmented the locomotor activity following nicotine administration. Of the investigated 5-HT(2) receptor ligands, M100907 (2 mg kg(-1)) or DOI (1 mg kg(-1)) administered during the first 5 days in combination with nicotine attenuated or enhanced, respectively, the development of nicotine sensitization. Given acutely on Day 10, M100907 (2 mg kg(-1)), Ro 60-0175 (1 mg kg(-1)), and WAY 163909 (1.5 mg kg(-1)) decreased the expression of nicotine sensitization. In another set of experiments, where the nicotine challenge test was performed on Day 15 in animals treated repeatedly (Days 1-5, 10) with nicotine, none of 5-HT(2) receptor ligands administered during the second withdrawal period (Days 11-14) to nicotine-treated rats altered the sensitizing effect of nicotine given on Day 15. Our data indicate that 5-HT(2A) receptors (but not 5-HT(2C) receptors) play a permissive role in the sensitizing effects of nicotine, while stimulation of 5-HT(2A) receptors enhances the development of nicotine sensitization and activation of 5-HT(2C) receptors is essential for the expression of nicotine sensitization. Repeated treatment with the 5-HT(2) receptor ligands within the second nicotine withdrawal does not inhibit previously established sensitization.

Increased effects of 3,4-methylenedioxymethamphetamine (ecstasy) in a rat model of depression.

3,4-Methylenedioxymethamphetamine (MDMA, ecstasy) is associated with increases in core body temperature (T(C)) and depressive mood states in users. Flinders Sensitive Line (FSL) rats represent a rat model of depression originally bred from Sprague-Dawley (SD) rats. They are more sensitive to both muscarinic and serotonergic agonists and have altered thermoregulatory responses to various drugs. To examine the link between MDMA and depression, eight FSL and eight SD rats were administered saline and 5 and 7.5 mgkg MDMA. Immediately following administration, rats were confined to an area with an ambient temperature (T(A)) of 30 ± 1°C for 30 minutes before being allowed access to a thermal gradient for four hours. The brains were removed one week after final dose of MDMA and concentrations of serotonin and dopamine were measured. Treatment with MDMA at both doses led to a higher T(C) in the FSL rats than the SD rats at high T(A) (P < 0.01). Fatalities due to hyperthermia occurred in the FSL rats after both doses, whereas all but one of the SD rats recovered well. Heart rate was also much higher after MDMA in the FSL rats throughout the experiments. The FSL rats showed significant decreases in all transmitters measured (P < 0.05). These differences between strains were not accounted for by altered blood or brain concentrations of MDMA. The results indicate that the FSL rats may be more susceptible to developing MDMA-induced hyperthermia and possible damage to the brain. These findings may be of importance to human users of MDMA who also have depression.

Cannabinoids as novel anti-inflammatory drugs.

Cannabinoids are a group of compounds that mediate their effects through cannabinoid receptors. The discovery of Δ9-tetrahydrocannabinol (THC) as the major psychoactive principle in marijuana, as well as the identification of cannabinoid receptors and their endogenous ligands, has led to a significant growth in research aimed at understanding the physiological functions of cannabinoids. Cannabinoid receptors include CB1, which is predominantly expressed in the brain, and CB2, which is primarily found on the cells of the immune system. The fact that both CB1 and CB2 receptors have been found on immune cells suggests that cannabinoids play an important role in the regulation of the immune system. Recent studies demonstrated that administration of THC into mice triggered marked apoptosis in T cells and dendritic cells, resulting in immunosuppression. In addition, several studies showed that cannabinoids downregulate cytokine and chemokine production and, in some models, upregulate T-regulatory cells (Tregs) as a mechanism to suppress inflammatory responses. The endocannabinoid system is also involved in immunoregulation. For example, administration of endocannabinoids or use of inhibitors of enzymes that break down the endocannabinoids, led to immunosuppression and recovery from immune-mediated injury to organs such as the liver. Manipulation of endocannabinoids andor use of exogenous cannabinoids in vivo can constitute a potent treatment modality against inflammatory disorders. This review will focus on the potential use of cannabinoids as a new class of anti-inflammatory agents against a number of inflammatory and autoimmune diseases that are primarily triggered by activated T cells or other cellular immune components.

Establishing derived textual control in activity schedules with children with autism.

Activity schedules are often used to facilitate task engagement and transition for children with autism. This study evaluated whether conditional discrimination training would serve to transfer the control from activity-schedule pictures to printed words (i.e., derived textual control). Two preschoolers with autism were taught to select pictures and printed words given their dictated names. Following training, participants could respond to printed words by completing the depicted task, match printed words to pictures, and read printed words without explicit training (i.e., emergent relations).

Involvement of dopamine D1D2 receptors on harmane-induced amnesia in the step-down passive avoidance test.

Ingestion of harmane and other alkaloids derived from plant Peganum harmala has been shown to elicit profound behavioural and toxic effects in humans, including hallucinations, excitation, feelings of elation, and euphoria. These alkaloids in the high doses can cause a toxic syndrome characterized by tremors and convulsions. Harmane has also been shown to act on a variety of receptor systems in the mammalian brain, including those for serotonin, dopamine and benzodiazepines. In animals, it has been reported to affect short and long term memory. In the present study, effects of dopamine D1 and D2 receptor antagonists on the harmane (HA)-induced amnesia and exploratory behaviors were examined in mice. One-trial step-down and hole-board paradigms were used for the assessment of memory retention and exploratory behaviors in adult male NMRI mice respectively. Intraperitoneal (i.p.) administration of HA (5 and 10 mgkg) immediately after training decreased memory consolidation, while had no effect on anxiety-like behavior. Memory retrieval was not altered by 15- or 30 min pre-testing administration of the D1 (SCH23390, 0.025, 0.05 and 0.1 mgkg) or D2 (sulpiride 12.5, 25 and 50 mgkg) receptor antagonists, respectively. In contrast, SCH23390 (0.05 and 0.1 mgkg) or sulpiride (25 and 50 mgkg) pre-test administration fully reversed HA-induced impairment of memory consolidation. Finally, neither D1 nor D2 receptor blockade affected exploratory behaviors in the hole-board paradigm. Altogether, these findings strongly suggest an involvement of D1 and D2 receptors modulation in the HA-induced impairment of memory consolidation.

Identification of post-generation effect of 3,4-methylenedioxymethamphetamine on the mouse brain by large-scale gene expression analysis.

The compound 3,4-methylenedioxymethamphetamine (MDMA or ecstasy) is a synthetic, psychoactive drug chemically similar to the stimulant methamphetamine and the hallucinogen mescaline. Accumulated data has revealed potential toxic effects associated with MDMA on brain serotonin and dopamine neurons in animal models. However, the relevance of these adverse effects on prenatal exposure to this drug remains unknown. In this study, we demonstrated that prenatal (F0) exposure to MDMA caused permanent large-scale transcriptional changes in the brains of the offspring (F1), especially in the cerebral cortex, by gene expression profiling analysis. The expression analysis of the brain of F1 pups, after maternal ingestion of MDMA (20 mgkg MDMA), revealed significant transcriptional changes in both male and female pups. Supervised analysis resulted in the identification of 804 outlier genes in males and 1784 outlier genes in females as MDMA-associated genes in the F1 generation. Most of the functional categories of genes, among the outlier genes, were intracellular signaling pathways, including the MAPK signaling pathway, Wnt signaling pathway, and neuroactive ligand-receptor interaction pathway. Although these genes were affected by MDMA exposure in utero, their association with brain dysfunction requires further investigation. The results of this study suggest that prenatal MDMA exposure may affect the developing brain.

Multiorgan failure from 1-benzylpiperazine ingestion--legal high or lethal high

1-benzylpiperazine (BZP) is synthetic stimulant. It was legal and openly sold in New Zealand before October 1, 2008. Two cases of life-threatening toxicity associated with BZP use are reported in detail in this article. Case one describes an adult female who developed status epilepticus, hyperthermia, disseminated intravascular coagulation, rhabdomyolysis, and renal failure associated with a BZP ingestion. Case two developed a similar pattern of toxicity from the combined use of BZP and 3,4-methylenedioxy-N-methylamphetamine. Both cases required prolonged hospital care but survived. There have been reports of deaths associated with the combined ingestion of BZP and 3,4-methylenedioxy-N-methylamphetamine. The effects may possibly be synergistic when co-ingested. Case one suggests that BZP alone has the potential to cause serious sympathomimetic toxicity.

Synthesis and determination of acute and chronic pain activities of 1-1-(3-methylphenyl) (tetralyl)piperidine as a new derivative of phencyclidine via tail immersion and formalin tests.

Phencyclidine (1-(1-phenylcyclohexyl)piperidine, CAS 956-90-1, PCP, 1) and ketamine (2-O-chlorophenyl-2-methylaminocyclohexan, CAS 1867-66-9, II) revealed some analgesic effects. Some of their derivatives have been synthesized for biological properties studies. Utilizing 1-tetralone as a starting material, 1-1-(3-methylphenyl)(tetralyl)piperidine, (PCP-CH3-tetralyl, III) was synthesized and its analgesic effects were studied on rats via tail immersion (as a model of acute thermal pain) and formalin (as a model of acute chemical and chronic pain) tests and compared with those of ketamine and PCP. The results indicated a marked anti-nociception 2-25 min after ketamine injection, but this analgesic effect lasted for 40 min following PCP-CH3-tetralyl application in the tail immersion test. However, the data obtained from the formalin test showed that chronic pain could be significantly attenuated by ketamine, PCP and PCP-CH3-tetralyl.

Dopaminergic augmentation of delta-9-tetrahydrocannabinol (THC) discrimination possible involvement of D(2)-induced formation of anandamide.

Although delta-9-tetreahydrocannabinol (THC)-induced elevations in accumbal dopamine levels are believed to play an important role in the abuse-related effects of cannabis, little direct evidence has been provided that the dopaminergic system is involved in the psychotropic effects of THC. The objective of this study is to investigate whether drugs activating or blocking the dopaminergic system modulate the discriminative effects of THC. In rats that had learned to discriminate 3 mgkg of THC from vehicle injections, the indirect dopaminergic agonists cocaine and amphetamine, the D(1)-receptor agonist SKF-38393, and the D(2)-receptor agonists quinpirole and apomorphine did not produce significant THC-like discriminative effects. However, both cocaine and amphetamine and D(2)-, but not the D(1)-, receptor agonists, augmented THC discrimination. Neither the D(1)-receptor antagonist SCH-23390 nor the D(2)-receptor antagonist raclopride reduced the discriminative effects of THC, even at doses that significantly depressed baseline operant responding. However, the D(2)-, but not the D(1)-, antagonist counteracted the augmentation of THCs discriminative effects produced by cocaine and amphetamine. We hypothesized that release of anandamide by activation of D(2) receptors was responsible for the observed augmentation of THC discrimination. This hypothesis was supported by two findings. First, the cannabinoid CB(1)-receptor antagonist rimonabant blocked quinpirole-induced augmentation of THC discrimination. Second, inhibition of anandamide degradation by blockade of fatty acid amide hydrolase augmented the THC-like effects of quinpirole. Dopamine does not play a major role in THC discrimination. However, activation of the dopaminergic system positively modulates the discriminative effects of THC, possibly through D(2)-induced elevations in brain levels of anandamide.

Acute psychotropic effects of oral cannabis extract with a defined content of Delta9-tetrahydrocannabinol (THC) in healthy volunteers.

The medical use of cannabinoids is limited mainly by their undesirable effects. With respect to acute psychotropic effects, the aim of this study is the comparison of an oral cannabis extract and low-dose diazepam in a cross-over experiment in drug-naïve healthy women. Sixteen healthy females participated in this randomized, double-blind, active comparator-controlled, single-dose, balanced 2-way cross-over study. Cannabis extract with standardised Delta (9)-tetrahydrocannabinol (THC) content (20 mg) or active placebo (5 mg diazepam) was administered orally. Subjects were assessed by self- and observer-rated visual analogue scales (VAS), the BRIEF PSYCHIATRIC RATING SCALE (BPRS) and three psychomotor tests up to 6 h after administration. VAS showed significantly elevated fatigue, drowsiness, dizziness, and feeling high after cannabis as compared to baseline and diazepam. BPRS scores were significantly higher after cannabis intake. Only in one psychomotor test a decrease of psychomotor activity after cannabis was evident. One subject in the cannabis condition experienced severe transient psychotic symptoms. Orally administered cannabis produced significant central depressant side-effects compared to diazepam, mostly subjective effects (VAS) but marginal effects in psychomotor performance in 15 healthy females. Regarding the medical use of cannabis, a rigorous benefit-risk analysis and an exact psychiatric assessment before and during treatment are necessary.

Acquisition session length modulates consolidation effects produced by 5-HT2C ligands in a mouse autoshaping-operant procedure.

Although the neurotransmitter, 5-hydroxytryptamine (serotonin, 5-HT), has been implicated as a mediator of learning and memory, the specific role of 5-HT receptors in rodents requires further delineation. In this study, 5-HT2C receptor ligands of varying relative intrinsic efficacies were tested in a mouse learning and memory model called autoshaping-operant. On day 1, mice were placed in experimental chambers and presented with a tone on a variable interval schedule. The tone remained on for 6 s or until a nose-poke response occurred to produce a dipper with Ensure solution. Mice were then injected with saline, MK212 (full agonist), m-chlorophenylpiperazine (partial agonist), mianserin, and SB206 553 (inverse agonists), and methysergide and ()-2-bromo lysergic acid diethylamide ()-hydrogen tartrate (neutral antagonists). Each compound was injected after either 1 or 2-h acquisition sessions on day 1 to investigate the role of acquisition session length on consolidation. Day 1 injection of the 5-HT2C inverse agonist mianserin produced greater retrieval impairments of the autoshaped operant response on day 2 than any other agent tested. Furthermore, decreasing the length of the acquisition session to 1h significantly increased the difficulty of the autoshaping task further modulating the consolidation effects produced by the 5-HT2C ligands tested.

THC Prevents MDMA Neurotoxicity in Mice.

The majority of MDMA (ecstasy) recreational users also consume cannabis. Despite the rewarding effects that both drugs have, they induce several opposite pharmacological responses. MDMA causes hyperthermia, oxidative stress and neuronal damage, especially at warm ambient temperature. However, THC, the main psychoactive compound of cannabis, produces hypothermic, anti-inflammatory and antioxidant effects. Therefore, THC may have a neuroprotective effect against MDMA-induced neurotoxicity. Mice receiving a neurotoxic regimen of MDMA (20 mgkg x 4) were pretreated with THC (3 mgkg x 4) at room (21 degrees C) and at warm (26 degrees C) temperature, and body temperature, striatal glial activation and DA terminal loss were assessed. To find out the mechanisms by which THC may prevent MDMA hyperthermia and neurotoxicity, the same procedure was carried out in animals pretreated with the CB(1) receptor antagonist AM251 and the CB(2) receptor antagonist AM630, as well as in CB(1), CB(2) and CB(1)CB(2) deficient mice. THC prevented MDMA-induced-hyperthermia and glial activation in animals housed at both room and warm temperature. Surprisingly, MDMA-induced DA terminal loss was only observed in animals housed at warm but not at room temperature, and this neurotoxic effect was reversed by THC administration. However, THC did not prevent MDMA-induced hyperthermia, glial activation, and DA terminal loss in animals treated with the CB(1) receptor antagonist AM251, neither in CB(1) and CB(1)CB(2) knockout mice. On the other hand, THC prevented MDMA-induced hyperthermia and DA terminal loss, but only partially suppressed glial activation in animals treated with the CB(2) cannabinoid antagonist and in CB(2) knockout animals. Our results indicate that THC protects against MDMA neurotoxicity, and suggest that these neuroprotective actions are primarily mediated by the reduction of hyperthermia through the activation of CB(1) receptor, although CB(2) receptors may also contribute to attenuate neuroinflammation in this process.

Deaths involving serotonergic drugs.

Serotonin-active drugs are detected relatively frequently in Victorian deaths. During 2002-2008, there were 1123 fatalities where one or more of the serotonin-active drugs tramadol, venlafaxine, fluoxetine, sertraline, citalopram, paroxetine and MDMA, were detected. These deaths were reviewed using pathology, toxicology and police reports, to determine the contribution of these drugs to the cause of death, particularly if serotonin toxicity was the mechanism of death. There were 28 cases of most interest to this research because of the presence of the target drugs and the circumstances suggesting the likelihood of serotonin toxicity involvement in death. There were 5 cases of reported serotonin toxicity and 23 other deaths suspected to have involved this form of toxicity. Tramadol featured most commonly out of the seven target drugs and was frequently detected in combination with serotonergic antidepressants. MDMA was also detected relatively commonly and was associated with moclobemide in 4 cases of confirmed serotonin toxicity. There were an additional 1095 cases where natural disease, external injury or the misuse of other drugs caused death, of which 2 reported the incidental contribution of serotonin toxicity.

Associations between duration of illicit drug use and health conditions results from the 2005-2007 national surveys on drug use and health.

To estimate and compare prevalence rates of lifetime health conditions by inferred duration of illicit drug use among the general U.S. adult population and to investigate associations between duration of use of each specific illicit drug (marijuana, cocaine, heroin, hallucinogens, or inhalant) and each lifetime health condition after controlling for potential confounding factors. Data from respondents aged 35 to 49 (N 29,195) from the 2005-2007 National Surveys on Drug Use and Health (NSDUH) were analyzed. The prevalence rates of a broad range of health conditions by duration of use of specific illicit drug among persons 35 to 49 years of age in the United States were estimated and compared. After adjustment for potential confounding factors, the results of 20 multivariate logistic regression models indicated positive associations between duration of marijuana use and anxiety, depression, sexually transmitted disease (STD), bronchitis, and lung cancer between duration of cocaine use and anxiety and pancreatitis between duration of heroin use and anxiety, hepatitis, and tuberculosis between duration of hallucinogen use and tinnitus and STD and between duration of inhalant use and anxiety, depression, HIVAIDS, STD, tuberculosis, bronchitis, asthma, sinusitis, and tinnitus. This study provides initial analyses on the relationships between illicit drug use and health conditions based on a large nationally representative sample. These results can help prepare for treating health problems among former and continuing illicit drug users.

Metabolic interactions between ethanol and MDMA in primary cultured rat hepatocytes.

3,4-Methylenedioxymethamphetamine (MDMA ecstasy), a drug of abuse commonly consumed at rave parties, is often taken in a polydrug abuse scenario, ethanol being one of the most associated drugs. Both MDMA and ethanol are mainly metabolized in the liver with formation of toxic metabolites. Our working hypothesis is that ethanol can modify the metabolism of MDMA through the cytochrome P450 system, and that this effect may be further potentiated by hyperthermia, a well-known consequence of MDMA abuse. To investigate these putative interactions we used primary rat hepatocyte cultures, which were exposed to 300 mM ethanol, 1.6 mM MDMA and the combination of both, at normothermic (36.5 degrees C) and hyperthermic (40.5 degrees C) conditions. After 24 h, the levels of MDA, HMA and HMMA in the cell culture medium were quantified by GCMS. In addition, we repeated the same experimental design preceded by 1h incubation with 0.18 microM ketoconazole or 150 microM diallyl sulphide (CYP3A and CYP2E1 inhibitors, respectively), to evaluate the putative role of these isoenzymes in the observed effects. The results obtained showed that ethanol exposure increases the formation of some MDMA metabolites such as HMA (1.8 times increase) and MDA (1.5 times increase). This effect was markedly increased under hyperthermic conditions (HMA, MDA and HMMA formation increased 10, 6 and 16 times, respectively) and is mediated, at least partially, by CYP3A and CYP2E1.

Peyote and mescaline exposures a 12-year review of a statewide poison center database.

Peyote, a cactus containing the hallucinogen mescaline, has been used by Native Americans for thousands of years. Illicit use is also known to occur, but reports in the medical literature consist only of isolated case reports. We sought to identify characteristics of patients with reported exposure to peyote or mescaline. We performed a retrospective review of the California Poison Control System database for the years 1997-2008 for all cases of single-substance human exposure using the search terms peyote and mescaline. There were a total of 31 single-substance exposures to peyote or mescaline. Thirty (97%) exposures were intentional 30 (97%) exposures were through the oral route, whereas one patient (3%) insufflated mescaline powder. Five patients (16%) were managed at home, whereas the remainder patients were managed in a healthcare facility. Commonly reported effects included hallucinations, tachycardia, agitation, and mydriasis. Vomiting was reported in only one case. Although uncommonly encountered, use of peyote and mescaline was associated with clinically significant effects requiring treatment in a substantial number of patients. Clinical effects were usually mild or moderate, and life-threatening toxicity was not reported in this case series.

Ecstasy as harmful as heroin

There is evidence that the use of MDMA (methylenedioxymethamphetamine), colloquially known as ecstasy particularly among late adolescents and young adults is increasing in Australia. Despite recent government-sponsored public education programs, there is a perception that recreational use of MDMA is much less harmful than other illicit substances like heroin. Recent seizures by police in Australia underline the extent of the demand for MDMA and how lucrative trafficking in MDMA has become. In two recent Australian cases, appellate courts considered the legislative intent of both State and Commonwealth legislation and held that a quantity-based penalty regime applied which distinguished between traffickable and commercial quantities of illicit drugs and that no distinction turned on the relative harmfulness of MDMA. In examining the question of harmfulness, this column summarises the pharmacology and morbidity of MDMA and considers the links between MDMA and other substances of abuse and the implications for further prevention programs.

Repeated intermittent methylenedioxymethamphetamine exposure protects against the behavioral and neurotoxic, but not hyperthermic, effects of an MDMA binge in adult rats.

We have recently shown that chronic intermittent exposure of adolescent rats to 3,4-methylenedioxymethamphetamine (MDMA or Ecstasy) completely blocks the reduction in serotonin transporter (SERT) binding and the hypoactivity seen following a subsequent MDMA binge treatment. The present study determined whether a similar neuroprotective effect also occurs in rats given the same intermittent MDMA exposure in adulthood. Adult male Sprague-Dawley rats were given either MDMA (10 mgkg x 2) or saline, every fifth day, from postnatal day (PD) 60 to PD 85. The MDMA-induced latency until seminal plug production was reduced over the course of intermittent treatments. After a 1-week wash-out period, animals received either a low- or high-dose MDMA binge (2.5 or 5.0 mgkg x 4). Core body temperature was measured during and after the binge to determine the effects of MDMA pretreatment on MDMA-induced hyperthermia. Spontaneous motor activity was determined the next day, and cortical and hippocampal samples were collected at 1 week postbinge to measure serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) concentrations as well as 3Hcitalopram binding to SERT. Hyperthermia occurred more rapidly and seminal discharge was more common in the MDMA-pretreated group compared to the MDMA-naïve group in animals given the low-dose binge. MDMA preexposure protected animals from the reductions in cortical 5-HT levels and SERT binding produced by the high-dose binge and blocked the postbinge hypoactivity. These findings indicate that chronic, intermittent MDMA exposure in adulthood induces neuroprotective effects similar to those seen with adolescent treatment. However, there was also evidence for drug-induced sensitization in adults that was not observed in adolescents. Thus, altered drug sensitivity in chronic Ecstasy users may depend not only on the frequency and pattern of use but also on the age of the user.

The cannabinoid 1-receptor silent antagonist O-2050 attenuates preference for high-fat diet and activated astrocytes in mice.

Endocannabinoids have been shown to activate reward-related feeding and to promote astrocytic differentiation. We investigated whether high-fat diet (HFD) intake produced a preference for HFD via an endocannabinoid-dependent mechanism. In the conditioned place preference test, the 2-week HFD-intake group showed preference for HFD and had increased expression of a marker for reactive astrocytes, glial fibrillary acid protein (GFAP), in the hypothalamus. The cannabinoid CB(1)-receptor antagonist O-2050 reduced the preference for HFD and expression of GFAP in the hypothalamus. These results suggested that HFD intake led to the development of a preference for HFD via astrocytic CB(1) receptors in the hypothalamus.

Gas chromatography-ion trap mass spectrometry method for the simultaneous measurement of MDMA (ecstasy) and its metabolites, MDA, HMA, and HMMA in plasma and urine.

The investigation of 3,4-methylenedioxymethamphetamine (MDMA ecstasy) abuse requires very robust methods with high sensitivity and wide linearity ranges for the quantification of this drug of abuse and its main metabolites in body fluids. An optimized gas chromatography-ion trap mass spectrometry (GC-ITMS) methodology with electron impact ionization addressing these issues is presented. The sample preparation involves an enzymatic hydrolysis of urine and plasma for conjugate cleavage, a SPE extraction, and a derivatization process. The method was fully validated in rat plasma and urine. Linearity for a wide concentration range was achieved for MDMA, and the metabolites 3,4-methylenedioxyamphetamine (MDA), 4-hydroxy-3-methoxyamphetamine (HMA) and 4-hydroxy-3-methoxymethamphetamine (HMMA). Limits of quantification were 2 ngmL in plasma and 3.5 ngmL in urine using a Selected Ion Monitoring detection mode. Selectivity, accuracy, precision, and recovery met the required criteria for the method validation. This GC-ITMS method provides high sensitivity and adequate performance characteristics for the simultaneous quantification of MDMA, MDA, HMA and HMMA in the studied matrices.

Receptors and channels targeted by synthetic cannabinoid receptor agonists and antagonists.

It is widely accepted that non-endogenous compounds that target CB(1) andor CB(2) receptors possess therapeutic potential for the clinical management of an ever growing number of disorders. Just a few of these disorders are already treated with Delta(9)-tetrahydrocannabinol or nabilone, both CB(1)CB(2) receptor agonists, and there is now considerable interest in expanding the clinical applications of such agonists and also in exploiting CB(2)-selective agonists, peripherally restricted CB(1)CB(2) receptor agonists and CB(1)CB(2) antagonists and inverse agonists as medicines. Already, numerous cannabinoid receptor ligands have been developed and their interactions with CB(1) and CB(2) receptors well characterized. This review describes what is currently known about the ability of such compounds to bind to, activate, inhibit or block non-CB(1), non- CB(2) G protein-coupled receptors such as GPR55, transmitter gated channels, ion channels and nuclear receptors in an orthosteric or allosteric manner. It begins with a brief description of how each of these ligands interacts with CB(1) andor CB(2) receptors.

The serotonin 2C receptor potently modulates the head-twitch response in mice induced by a phenethylamine hallucinogen.

Hallucinogenic serotonin 2A (5-HT(2A)) receptor partial agonists, such as ( or -)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI), induce a frontal cortex-dependent head-twitch response (HTR) in rodents, a behavioral proxy of a hallucinogenic response that is blocked by 5-HT(2A) receptor antagonists. In addition to 5-HT(2A) receptors, DOI and most other serotonin-like hallucinogens have high affinity and potency as partial agonists at 5-HT(2C) receptors. We tested for involvement of 5-HT(2C) receptors in the HTR induced by DOI. Comparison of 5-HT(2C) receptor knockout and wild-type littermates revealed an approximately 50% reduction in DOI-induced HTR in knockout mice. Also, pretreatment with either the 5-HT(2C) receptor antagonist SB206553 or SB242084 eradicated a twofold difference in DOI-induced HTR between the standard inbred mouse strains C57BL6J and DBA2J, and decreased the DOI-induced HTR by at least 50% in both strains. None of several measures of 5-HT(2A) receptors in frontal cortex explained the strain difference, including 5-HT(2A) receptor density, Galpha(q) or Galpha(io) protein levels, phospholipase C activity, or DOI-induced expression of Egr1 and Egr2. 5-HT(2C) receptor density in the brains of C57BL6J and DBA2J was also equivalent, suggesting that 5-HT(2C) receptor-mediated intracellular signaling or other physiological modulators of the HTR may explain the strain difference in response to DOI. We conclude that the HTR to DOI in mice is strongly modulated by 5-HT(2C) receptor activity. This novel finding invites reassessment of hallucinogenic mechanisms involving 5-HT(2) receptors.

Transmembrane domain 6 of the human serotonin transporter contributes to an aqueously accessible binding pocket for serotonin and the psychostimulant 3,4-methylene dioxymethamphetamine.

The plasma membrane serotonin (5-HT) transporter (SERT, SLC6A4) clears 5-HT after release at nerve termini and is targeted by both antidepressant medications and psychostimulants (e.g. MDMA, cocaine). Homology modeling of human SERT (hSERT), based on high resolution structures of the microbial SLC6 family member LeuT(Aa), along with biochemical studies of wild type and mutant transporters, predicts transmembrane (TM) domains 1, 3, 6, and 8 comprise the 5-HT-binding pocket. We utilized the substituted cysteine accessibility method along with surface and site-specific biotinylation to probe TM6 for aqueous accessibility and differential interactions with 5-HT and psychostimulants. Our results are consistent with TM6 being composed of an aqueous-accessible, alpha-helical extracellular domain (TM6a) that is separated by a central, unwound section from a cytoplasmically localized domain (TM6b) with limited aqueous accessibility. The substitution G338C appears to lock hSERT in an outward-facing conformation that, although accessible to aminoethylmethanethiosulfonate-biotin, 5-HT, and citalopram, is incapable of inward 5-HT transport. Transport of 5-HT by G338C can be partially restored by the TM1 mutation Y95F. With regard to methanethiosulfonate (MTS) inactivation of uptake, TM6a Cys mutants demonstrate Na()-dependent 2-(trimethylammonium)ethyl-MTS sensitivity. Studies with the centrally located substitution S336C reveal features of a common binding pocket for 5-HT and 3,4-methylenedioxymethamphetamine (MDMA). Interestingly, the substitution I333C reveals an MDMA-induced conformation not observed with 5-HT. In the context of prior studies on TM1, our findings document shared and unique features of TM6 contributing to hSERT aqueous accessibility, ligand recognition, and conformational dynamics.

Differential effects of ecstasy on short-term and working memory a meta-analysis.

Quantitative analysis of studies examining the effect of ecstasy on short-term and working memory in the verbal and visuo-spatial domain was undertaken. Thirty verbal short-term memory, 22 verbal working memory, 12 visuospatial short-term memory and 9 visuospatial working memory studies met inclusion criteria. Ecstasy users performed significantly worse in all memory domains, both in studies using drug-naïve controls and studies using polydrug controls. These results are consistent with previous meta-analytic findings that ecstasy use is associated with impaired short-term memory function. Lifetime ecstasy consumption predicted effect size in working memory but not in short-term memory. The current meta-analysis adds to the literature by showing that ecstasy use in humans is also associated with impaired working memory, and that this impairment is related to total lifetime ecstasy consumption. These findings highlight the long-term, cumulative behavioral consequences associated with ecstasy use in humans.

Sex differences in the effects of N,N-diethyllysergamide (LSD) on behavioural activity and prepulse inhibition.

The aim of this study was to describe sex differences in the behavioural effects of N,N-diethyllysergamide (LSD) (locomotor activity and other behavioural repertoire in the open field) and its effects on sensorimotor gating in rats (prepulse inhibition (PPI) of the acoustic startle reaction). Three groups of animals were analysed males, oestral and pro-oestral phase females (EP females), and metoestral and dioestral phase females (MD females). LSD (5, 50 and 200 microgkg subcutaneously) attenuated locomotor activity and normal behavioural repertoire, and induced flat body posture, wet dog shakes and disrupted PPI. The most prominent behavioural findings of LSD were for LSD 200 microgkg which suppressed almost all behavioural activity. LSD had mainly inhibitory locomotor effects in males and MD females, yet in EP female rats LSD increased locomotion during the second half of testing period. The main sex differences were observed in locomotor and exploratory behaviour. Both EP and MD females were less sensitive to hypolocomotor effects of LSD and had less pronounced thigmotaxis than males. Further EP females had increased rearing after LSD 5microgkg. On the contrary although LSD disrupted PPI in males and MD female rats, EP females were protected from this disruptive effect. Thus, EP females seem to have a lower sensitivity to LSD behavioural actions.

Normal thermoregulatory responses to 3-iodothyronamine, trace amines and amphetamine-like psychostimulants in trace amine associated receptor 1 knockout mice.

3-Iodothyronamine (T1AM) is a metabolite of thyroid hormone. It is an agonist at trace amine-associated receptor 1 (TAAR1), a recently identified receptor involved in monoaminergic regulation and a potential novel therapeutic target. Here, T1AM was studied using rhesus monkey TAAR1 andor human dopamine transporter (DAT) co-transfected cells, and wild-type (WT) and TAAR1 knock-out (KO) mice. The IC(50) of T1AM competition for binding of the DAT-specific radio-ligand (3)HCFT was highly similar in DAT cells, WT striatal synaptosomes and KO striatal synaptosomes (0.72-0.81 microM). T1AM inhibition of 10 nM (3)Hdopamine uptake (IC(50) WT, 1.4 or - 0.5 microM KO, 1.2 or - 0.4 microM) or 50 nM (3)Hserotonin uptake (IC(50) WT, 4.5 or - 0.6 microM KO, 4.7 or - 1.1 microM) in WT and KO synaptosomes was also highly similar. Unlike other TAAR1 agonists that are DAT substrates, TAAR1 signaling in response to T1AM was not enhanced in the presence of DAT as determined by CRE-luciferase assay. In vivo, T1AM induced robust hypothermia in WT and KO mice equivalently and dose dependently (maximum change degrees Celsius 50 mgkg at 60 min WT -6.0 or - 0.4, KO -5.6 or - 1.0 and 25 mgkg at 30 min WT -2.7 or - 0.4, KO -3.0 or - 0.2). Other TAAR1 agonists including beta-phenylethylamine (beta-PEA), MDMA (3,4-methylenedioxymethamphetamine) and methamphetamine also induced significant, time-dependent thermoregulatory responses that were alike in WT and KO mice. Therefore, TAAR1 co-expression does not alter T1AM binding to DAT in vitro nor T1AM inhibition of (3)Hmonoamine uptake ex vivo, and TAAR1 agonist-induced thermoregulatory responses are TAAR1-independent. Accordingly, TAAR1-directed compounds will likely not affect thermoregulation nor are they likely to be cryogens.

Elevated urine levels of bufotenine in patients with autistic spectrum disorders and schizophrenia.

Previous studies have suggested that the endogeneous psychotomimetic molecule bufotenine (N-N-dimethyl-5-idroxytryptamine) may play a role in the pathogenesis of severe mental disorders. The potential association of bufotenine with the clinical features of autism and schizophrenia is not entirely understood. In this study, we measured urinary levels of bufotenine in subjects with autistic spectrum disorder (ASD), schizophrenia and healthy comparison subjects free of psychiatric symptoms. We also sought to assess whether urine concentrations of this molecule may be associated with the clinical characteristics of psychiatric patients. Urine bufotenine levels were measured using a high-performance liquid chromatography-mass spectrometry (HPLC-MS) assay in young adults with severe ASD (n15), patients with schizophrenia (n15), and healthy control subjects (n18). The Vineland Adaptive Behavior Scale was used to measure adaptive behaviors in ASD individuals. The Brief Psychiatric Rating Scale (BPRS) was used for patients with schizophrenia. Urine bufotenine levels were significantly higher in ASD subjects (3.30 - 0.49 microgL, p<0.05) and patients with schizophrenia (4.39 - 0.43 microgL, p<0.001) compared with controls (1.53 - 0.30 microgL). Among patients with ASD, there was a significant positive correlation between urine bufotenine and hyperactivity scores on the Vineland Adaptive Behavior Scale (r0.479, p<0.05). No other associations were detected. Our results indicate that elevated urine levels of the endogeneous psychotomimetic molecule bufotenine may play a role in ASD and schizophrenia, and can be correlated with hyperactivity scores in autism.

Drug classification science, politics, both or neither

Governments currently classify illicit drugs for various purposes to guide courts in the sentencing of convicted violators of drug control laws, to prioritize targets of prevention measures and to educate the public about relative risks of the various drugs. It has been proposed that classification should be conducted by scientists and drug experts rather than by politicians, so that it will reflect only accurate factual knowledge of drug effects and risks rather than political biases. Although this is an appealing goal, it is inherently impossible because rank-ordering of the drugs inevitably requires value judgements concerning the different types of harm. Such judgements, even by scientists, depend upon subjective personal criteria and not only upon scientific facts. Moreover, classification that is meant to guide the legal system in controlling dangerous drug use can function only if it is in harmony with the values and sentiments of the public. In some respects, politicians may be better attuned to public attitudes and wishes, and to what policies the public will support, than are scientific experts. The problems inherent in such drug classification are illustrated by the examples of cannabis and of salvinorin A. They raise the question as to whether the classification process really serves any socially beneficial purpose.

Analysis of licking microstructure provides no evidence for a reduction in reward value following acute or sub-chronic phencyclidine administration.

The N-methyl D-aspartate antagonist phencyclidine (PCP) is purported to mimic the negative, cognitive and positive symptoms of schizophrenia. Thus, acute and sub-chronic PCP treatment in rodents might produce anhedonia, a decrease in the pleasure produced by rewards. Experiment 1 investigated whether acute PCP treatment changes the value of sucrose. A comparison was made to ()MK-801, a drug often used interchangeably with PCP in preclinical studies. Experiment 2 assessed the effects of withdrawal from sub-chronic PCP treatment on the value of sucrose. Experiment 1 examined the dose-response effects of PCP and ()MK-801 on licking microstructure during sucrose consumption. Experiment 2 assessed the effects of withdrawal from sub-chronic PCP treatment (5 mgkg twice daily for 7 days), on licking microstructure during sucrose consumption. Locomotor activity testing was carried out in experiment 2 to confirm the sensitisation effect of the PCP regimen on amphetamine-induced hyperlocomotion. Low to moderate acute doses of PCP and ()MK-801 increased the amount of sucrose consumed. Higher doses decreased consumption and the number of licks per cluster (cluster size) but also increased the average inter-lick interval, which may indicate motor impairment. There was no evidence that withdrawal from sub-chronic PCP treatment produced decreases in consumption or lick cluster size. Following acute PCP treatment, we found no evidence of reduced reward value without the presence of confounding motor deficits. Sub-chronic PCP withdrawal also produced no decrease in reward value. Therefore, the current results indicate that neither acute PCP treatment nor sub-chronic PCP withdrawal produce consummatory anhedonia.

The psychedelic genes of maize redundantly promote carbohydrate export from leaves.

Whole-plant carbohydrate partitioning involves the assimilation of carbon in leaves and its translocation to nonphotosynthetic tissues. This process is fundamental to plant growth and development, but its regulation is poorly understood. To identify genes controlling carbohydrate partitioning, we isolated mutants that are defective in exporting fixed carbon from leaves. Here we describe psychedelic (psc), a new mutant of maize (Zea mays) that is perturbed in carbohydrate partitioning. psc mutants exhibit stable, discrete chlorotic and green regions within their leaves. psc chlorotic tissues hyperaccumulate starch and soluble sugars, while psc green tissues appear comparable to wild-type leaves. The psc chlorotic and green tissue boundaries are usually delineated by larger veins, suggesting that translocation of a mobile compound through the veins may influence the tissue phenotype. psc mutants display altered biomass partitioning, which is consistent with reduced carbohydrate export from leaves to developing tissues. We determined that the psc mutation is unlinked to previously characterized maize leaf carbohydrate hyperaccumulation mutants. Additionally, we found that the psc mutant phenotype is inherited as a recessive, duplicate-factor trait in some inbred lines. Genetic analyses with other maize mutants with variegated leaves and impaired carbohydrate partitioning suggest that Psc defines an independent pathway. Therefore, investigations into the psc mutation have uncovered two previously unknown genes that redundantly function to regulate carbohydrate partitioning in maize.

The effects of the glycine reuptake inhibitor R213129 on the central nervous system and on scopolamine-induced impairments in psychomotor and cognitive function in healthy subjects.

In this study the effects of R213129, a selective glycine transporter 1 inhibitor, on central nervous system function were investigated in healthy males in the absence and presence of scopolamine. This was a double-blind, placebo-controlled, 4-period crossover ascending dose study evaluating the following endpoints body sway, saccadic and smooth pursuit eye movements, pupillometry, electroencephalography, visual analogue scales for alertness, mood, calmness and psychedelic effects, adaptive tracking, finger tapping, Visual and Verbal Learning Task, Stroop test, hormone levels and pharmacokinetics. R213129 dose levels were selected based on exposure levels that blocked the GlyT1 sites >50% in preclinical experiments. Forty-three of the 45 included subjects completed the study. Scopolamine significantly affected almost every central nervous system parameter measured in this study. R213129 alone compared with placebo did not elicit pharmacodynamic changes. R213129 had some small effects on scopolamine-induced central nervous system impairments. Scopolamine-induced finger tapping impairment was further enhanced by 3 mg R213129 with 2.0 taps10 seconds (95% CI -4.0, -0.1), electroencephalography alpha power was increased by 10 mg R213129 with respectively 12.9% (0.7, 26.6%), scopolamine-induced impairment of the Stroop test was partly reversed by 10 mg R213129 with 59 milliseconds (-110, -7). Scopolamine produced robust and consistent effects in psychomotor and cognitive function in healthy volunteers. The most logical reason for the lack of R213129 effects seems to be that the central nervous system concentrations were too low. The effects of higher doses in healthy volunteers and the clinical efficacy in patients remain to be established.

Central nervous system effects of haloperidol on THC in healthy male volunteers.

In this study, the hypothesis that haloperidol would lead to an amelioration of Δ9-tetrahydrocannabinol (THC)-induced psychotomimetic effects was investigated. In a double-blind, placebo-controlled, partial three-way crossover ascending dose study the effects of THC, haloperidol and their combination were investigated in 35 healthy, male mild cannabis users, measuring Positive and Negative Syndrome Scale, Visual Analogue Scales for alertness, mood, calmness and psychedelic effects, saccadic and smooth pursuit eye measurements, electroencephalography, Body Sway, Stroop test, Visual and Verbal Learning Task, hormone levels and pharmacokinetics. Compared with placebo, THC significantly decreased smooth pursuit, Visual Analogue Scales alertness, Stroop test performance, immediate and delayed word recall and prolactin concentrations, and significantly increased positive and general Positive and Negative Syndrome Scale score, Visual Analogue Scales feeling high, Body Sway and electroencephalography alpha. Haloperidol reversed the THC-induced positive Positive and Negative Syndrome Scale increase to levels observed with haloperidol alone, but not THC-induced high feelings. Compared with placebo, haloperidol significantly decreased saccadic peak velocity, smooth pursuit, Visual Analogue Scales mood and immediate and delayed word recall and significantly increased Body Sway, electroencephalography theta and prolactin levels. THC-induced increases in positive Positive and Negative Syndrome Scale but not in Visual Analogue Scales feeling high were reversed by haloperidol. This indicates that psychotic-like effects induced by THC are mediated by dopaminergic systems, but that other systems are involved in feeling high. Additionally, the clear reductions of psychotic-like symptoms by a clinically relevant dose of haloperidol suggest that THC administration may be a useful pharmacological cannabinoid model for psychotic effects in healthy volunteers.

Comparing probability and non-probability sampling methods in Ecstasy research implications for the internet as a research tool.

The usage of Ecstasy and related drug (ERD) has increasingly been the focus of epidemiological and other public health-related research. One of the more promising methods is the use of the Internet as a recruitment and survey tool. However, there remain methodological concerns and questions about representativeness. Three samples of ERD users in Melbourne, Australia surveyed in 2004 are compared in terms of a number of key demographic and drug use variables. The Internet, face-to-face, and probability sampling methods appear to access similar but not identical groups of ERD users. Implications and limitations of the study are noted and future research is recommended.

Precipitated withdrawal counters the adverse effects of subchronic cannabinoid administration on male rat sexual behavior.

In the present study, sexual behavior of male rats was assessed following prolonged treatment with the CB(1) receptor agonist, HU-210 (0.1mgmgday for 10 days) under conditions of drug maintenance, spontaneous withdrawal and precipitated withdrawal (induced via administration of the CB(1) receptor antagonist AM251 1mgkg). Following subchronic cannabinoid treatment, sexual activity in male rats was impaired under both the drug maintenance and spontaneous withdrawal conditions, as revealed by a reduction in frequency of both intromissions and ejaculations. Notably, the induction of precipitated drug withdrawal reversed the negative effects of subchronic HU-210 treatment on sexual activity as seen by a reversal of the suppression of ejaculations. These data illustrate that, contrary to expectations, the impairments in male sexual activity following protracted cannabinoid administration are not due to drug withdrawal, per se, but are likely mediated by neuroadaptive changes provoked by repeated drug exposure.

Comparative effects of 3,4-methylenedioxymethamphetamine and 4-methylthioamphetamine on rat liver mitochondrial function.

Ecstasy, which is used as a recreational drug, is a common street name for 3, 4-methylenedioxymethamphetamine (MDMA). Another drug of abuse chemically related, though less common than MDMA, is the amphetamine derivative 4-methylthioamphetamine (MTA). MDMA and MTA induce different systemic and organ-specific effects, including neurotoxicity, hyperthermia, nephrotoxicity, cardiotoxicity and hepatotoxicity. Therefore, it is clear that MDMA and MTA are responsible for inducing organ toxicity. The mechanisms underlying MDMA and MTA-induced hepatotoxicity are multifactorial, and therefore not completely understood. Recent findings indicate interference with cellular bioenergetics as an important toxicological feature of ecstasy. However, less is known about the involvement of mitochondria in MTA-induced hepatotoxicity. Thus, we compared the direct influence of MDMA and MTA on rat liver mitochondrial function mitochondrial permeability transition (MPT), mitochondrial oxidative stress, and mitochondrial bioenergetics. It was shown that MTA (from 0.025 up to 0.1mM) was more potent than MDMA (from 0.2 up to 0.5mM) in decreasing the sensitivity of rat liver mitochondria to MPT. However, higher concentrations of MTA (from 0.5 up to 2mM) were highly toxic to mitochondria. MTA simultaneously increased H(2)O(2) production in a monoamine oxidase (MAO)-dependent way, and uncoupled and inhibited mitochondrial respiration. In contrast, MDMA had only limited or no effects on these mitochondrial parameters. According to these results, it is possible to postulate that, depending on the concentration, MTA can potentially be more efficient in its effects on liver mitochondria than MDMA and, also, that its harmful effects may contribute to its hepatotoxicity.

The effects of 3,4-methylenedioxymethamphetamine (MDMA) on nicotinic receptors intracellular calcium increase, calpaincaspase 3 activation, and functional upregulation.

Previous work by our group demonstrated that homomeric alpha7 nicotinic acetylcholine receptors (nAChR) play a role in the neurotoxicity induced by 3,4-methylenedioxymethamphetamine (MDMA), as well as the binding affinity of this drug to these receptors. Here we studied the effect of MDMA on the activation of nAChR subtypes, the consequent calcium mobilization, and calpaincaspase 3 activation because prolonged Ca(2) increase could contribute to cytotoxicity. As techniques, we used fluorimetry in Fluo-4-loaded PC12 cells and electrophysiology in Xenopus oocytes. MDMA produced a rapid and sustained increase in calcium without reaching the maximum effect induced by ACh. It also concentration-dependently inhibited the response induced by ACh, nicotine, and the specific alpha7 agonist PNU 282987 with IC(50) values in the low micromolar range. Similarly, MDMA induced inward currents in Xenopus oocytes transfected with human alpha7 but not with alpha4beta2 nAChR and inhibited ACh-induced currents in both receptors in a concentration-dependent manner. The calcium response was inhibited by methyllycaconitine (MLA) and alpha-bungarotoxin but not by dihydro-beta-erythroidine. These results therefore indicate that MDMA acts as a partial agonist on alpha7 nAChRs and as an antagonist on the heteromeric subtypes. Subsequently, calcium-induced Ca(2) release from the endoplasmic reticulum and entry through voltage-operated calcium channels are also implicated as proved using specific antagonists. In addition, treatment with MDMA for 24 h significantly increased basal Ca(2) levels and induced an increase in alpha-spectrin breakdown products, which indicates that calpain and caspase 3 were activated. These effects were inhibited by pretreatment with MLA. Moreover, pretreatment with MDMA induced functional upregulation of calcium responses to specific agonists of both heteromeric and alpha7 nAChR. Sustained calcium entry and calpain activation could favor the activation of Ca(2)-dependent enzymes such as protein kinase C and nitric oxide synthase, which are involved in the generation of ROS and the blockade of the dopamine transporter. This, together with caspase 3 activation, must play a role in MDMA-induced cytotoxicity.

Jimson weed abuse in an Oklahoma teen.

Jimson weed, a plant often abused by teenagers and young adults, grows wild throughout Oklahoma. It is best known for its hallucinogenic properties however intoxication can lead to anticholinergic manifestations that are potentially dangerous. Over the past six years, sixty-three individuals in Oklahoma have been hospitalized for jimson weed intoxication, including this Oklahoma teen. Importance lies in proper identification, understanding, and management in persons presenting with jimson weed poisoning.

A case of cannabis-induced mania.

We present the course of a mania most likely induced by cannabis in a young cannabis-dependent patient, but apart from this, healthy male adult. Mania developed in parallel to increasing cannabis inhalation and remitted quickly and completely within one week of abstinence without any antimanic medication in a protective inpatient setting. Simultaneously, the THC-COOH level decreased in serum and urine.

Hair analysis for Delta9-tetrahydrocannabinolic acid A--new insights into the mechanism of drug incorporation of cannabinoids into hair.

Differentiation between external contamination and incorporation of drugs or their metabolites from inside the body via blood, sweat or sebum is a general issue in hair analysis and of high concern when interpreting analytical results. In hair analysis for cannabinoids the most common target is Delta9-tetrahydrocannabinol (THC), sometimes cannabidiol (CBD) and cannabinol (CBN) are determined additionally. After repeated external contamination by cannabis smoke these analytes are known to be found in hair even after performing multiple washing steps. A widely accepted strategy to unequivocally prove active cannabis consumption is the analysis of hair extracts for the oxidative metabolite 11-nor-9-carboxy-THC (THC-COOH). Although the acidic nature of this metabolite suggests a lower rate of incorporation into the hair matrix compared to THC, it is not fully understood up to now why hair concentrations of THC-COOH are generally found to be much lower (mostly <10 pgmg) than the corresponding THC concentrations. Delta9-Tetrahydrocannabinolic acid A (THCA A) is the preliminary end product of the THC biosynthesis in the cannabis plant. Unlike THC it is non-psychoactive and can be regarded as a precursor of THC being largely decarboxylated when heated or smoked. The presented work shows for the first time that THCA A is not only detectable in blood and urine of cannabis consumers but also in THC positive hair samples. A pilot experiment performed within this study showed that after oral intake of THCA A on a regular basis no relevant incorporation into hair occurred. It can be concluded that THCA A in hair almost exclusively derives from external contamination e.g. by side stream smoke. Elevated temperatures during the analytical procedure, particularly under alkaline conditions, can lead to decarboxylation of THCA A and accordingly increase THC concentrations in hair. Additionally, it has to be kept in mind that in hair samples tested positive for THCA A at least a part of the non-artefact THC probably derives from external contamination as well, because in condensate of cannabis smoke both THC and THCA A are present in relevant amounts. External contamination by side stream smoke could therefore explain the great differences in THC and THC-COOH hair concentrations commonly found in cannabis users.

Auditory mismatch negativity deficits in long-term heavy cannabis users.

Mismatch negativity (MMN) is an auditory event-related potential indicating auditory sensory memory and information processing. The present study tested the hypothesis that chronic cannabis use is associated with deficient MMN generation. MMN was investigated in age- and gender-matched chronic cannabis users (n 30) and nonuser controls (n 30). The cannabis users were divided into two groups according to duration and quantity of cannabis consumption. The MMNs resulting from a pseudorandomized sequence of 2 × 900 auditory stimuli were recorded by 32-channel EEG. The standard stimuli were 1,000 Hz, 80 dB SPL and 90 ms duration. The deviant stimuli differed in duration (50 ms) or frequency (1,200 Hz). There were no significant differences in MMN values between cannabis users and nonuser controls in both deviance conditions. With regard to subgroups, reduced amplitudes of frequency MMN at frontal electrodes were found in long-term (≥8 years of use) and heavy (≥15 jointsweek) users compared to short-term and light users. The results indicate that chronic cannabis use may cause a specific impairment of auditory information processing. In particular, duration and quantity of cannabis use could be identified as important factors of deficient MMN generation.

Psychedelics and the human receptorome.

We currently understand the mental effects of psychedelics to be caused by agonism or partial agonism of 5-HT(2A) (and possibly 5-HT(2C)) receptors, and we understand that psychedelic drugs, especially phenylalkylamines, are fairly selective for these two receptors. This manuscript is a reference work on the receptor affinity pharmacology of psychedelic drugs. New data is presented on the affinity of twenty-five psychedelic drugs at fifty-one receptors, transporters, and ion channels, assayed by the National Institute of Mental Health-Psychoactive Drug Screening Program (NIMH-PDSP). In addition, comparable data gathered from the literature on ten additional drugs is also presented (mostly assayed by the NIMH-PDSP). A new method is introduced for normalizing affinity (K(i)) data that factors out potency so that the multi-receptor affinity profiles of different drugs can be directly compared and contrasted. The method is then used to compare the thirty-five drugs in graphical and tabular form. It is shown that psychedelic drugs, especially phenylalkylamines, are not as selective as generally believed, interacting with forty-two of forty-nine broadly assayed sites. The thirty-five drugs of the study have very diverse patterns of interaction with different classes of receptors, emphasizing eighteen different receptors. This diversity of receptor interaction may underlie the qualitative diversity of these drugs. It should be possible to use this diverse set of drugs as probes into the roles played by the various receptor systems in the human mind.

Everyday and prospective memory deficits in ecstasypolydrug users.

The impact of ecstasypolydrug use on real-world memory (i.e. everyday memory, cognitive failures and prospective memory PM) was investigated in a sample of 42 ecstasypolydrug users and 31 non-ecstasy users. Laboratory-based PM tasks were administered along with self-reported measures of PM to test whether any ecstasypolydrug-related impairment on the different aspects of PM was present. Self-reported measures of everyday memory and cognitive failures were also administered. Ecstasypolydrug associated deficits were observed on both laboratory and self-reported measures of PM and everyday memory. The present study extends previous research by demonstrating that deficits in PM are real and cannot be simply attributed to self-misperceptions. The deficits observed reflect some general capacity underpinning both time- and event-based PM contexts and are not task specific. Among this group of ecstasypolydrug users recreational use of cocaine was also prominently associated with PM deficits. Further research might explore the differential effects of individual illicit drugs on real-world memory.

Pharmacological characterization of the discriminative stimulus of inhaled 1,1,1-trichloroethane.

The present study examined the involvement of the GABAA, N-methyl-D-aspartate (NMDA), nicotinic acetylcholine, and mu-opioid receptor systems in the transduction of the discriminative stimulus effects of the abused inhalant 1,1,1-trichloroethane (TCE). Sixteen B6SJLF1J mice were trained to discriminate 10 min of exposure to 12,000-ppm inhaled TCE vapor from air. Substitution and antagonism tests and TCE blood concentration analysis were subsequently conducted. TCE blood concentrations decreased rapidly after cessation of exposure, falling by 66% within 5 min. TCE vapor concentration-dependently substituted for the 12,000-ppm training stimulus. The volatile anesthetic halothane concentration-dependently and fully substituted for TCE. The benzodiazepine midazolam partially substituted for TCE, producing a maximum of 68% TCE-lever selection. The benzodiazepine antagonist flumazenil attenuated midazolam substitution for TCE, but not the discriminative stimulus effects of TCE itself. The noncompetitive NDMA receptor antagonists phencyclidine and dizocilpine failed to substitute for TCE. Nicotine and the central nicotinic receptor antagonist mecamylamine also failed to produce any TCE-lever selection, nor did they antagonize the discriminative stimulus of TCE. The mu-opioid receptor agonist morphine did not substitute for TCE. The opioid antagonist naltrexone failed to antagonize the discriminative stimulus of TCE. Overall, the present results, combined with previous studies, suggest that the discriminative stimulus effects of TCE are mediated primarily by positive GABAA receptor modulatory effects though a mechanism distinct from the benzodiazepine binding site.

Analysis of cannabis in oral fluid specimens by GC-MS with automatic SPE.

Methamphetamine (MA) is the most commonly abused drug in Korea, followed by cannabis. Traditionally, MA analysis is carried out on both urine and hair samples and cannabis analysis in urine samples only. Despite the fact that oral fluid has become increasingly popular as an alternative specimen in the field of driving under the influence of drugs (DUID) and work place drug testing, its application has not been expanded to drug analysis in Korea. Oral fluid is easy to collect and handle and can provide an indication of recent drug abuse. In this study, we present an analytical method using GC-MS to determine tetrahydrocannabinol (THC) and its main metabolite 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in oral fluid. The validated method was applied to oral fluid samples collected from drug abuse suspects and the results were compared with those in urine. The stability of THC and THC-COOH in oral fluid stored in different containers was also investigated. Oral fluid specimens from 12 drug abuse suspects, submitted by the police, were collected by direct expectoration. The samples were screened with microplate ELISA. For confirmation they were extracted using automated SPE with mixed-mode cation exchange cartridge, derivatized and analyzed by GC-MS using selective ion monitoring (SIM). The concentrations ofTHC and THC-COOH in oral fluid showed a large variation and the results from oral fluid and urine samples from cannabis abusers did not show any correlation. Thus, detailed information about time interval between drug use and sample collection is needed to interpret the oral fluid results properly. In addition, further investigation about the detection time window ofTHC and THC-COOH in oral fluid is required to substitute oral fluid for urine in drug testing.

Deficits of long-term memory in ecstasy users are related to cognitive complexity of the task.

Despite animal evidence that methylenedioxymethamphetamine (ecstasy) causes lasting damage in brain regions related to long-term memory, results regarding human memory performance have been variable. This variability may reflect the cognitive complexity of the memory tasks. However, previous studies have tested only a limited range of cognitive complexity. Furthermore, comparisons across different studies are made difficult by regional variations in ecstasy composition and patterns of use. The objective of this study is to evaluate ecstasy-related deficits in human verbal memory over a wide range of cognitive complexity using subjects drawn from a single geographical population. Ecstasy users were compared to non-drug using controls on verbal tasks with low cognitive complexity (stem completion), moderate cognitive complexity (stem-cued recall and word list learning) and high cognitive complexity (California Verbal Learning Test, Verbal Paired Associates and a novel Verbal Triplet Associates test). Where significant differences were found, both groups were also compared to cannabis users. More cognitively complex memory tasks were associated with clearer ecstasy-related deficits than low complexity tasks. In the most cognitively demanding task, ecstasy-related deficits remained even after multiple learning opportunities, whereas the performance of cannabis users approached that of non-drug using controls. Ecstasy users also had weaker deliberate strategy use than both non-drug and cannabis controls. Results were consistent with the proposal that ecstasy-related memory deficits are more reliable on tasks with greater cognitive complexity. This could arise either because such tasks require a greater contribution from the frontal lobe or because they require greater interaction between multiple brain regions.

Cannabinoid receptor 1 binding activity and quantitative analysis of Cannabis sativa L. smoke and vapor.

Cannabis sativa L. (cannabis) extracts, vapor produced by the Volcano vaporizer and smoke made from burning cannabis joints were analyzed by GC-flame ionization detecter (FID), GC-MS and HPLC. Three different medicinal cannabis varieties were investigated Bedrocan, Bedrobinol and Bediol. Cannabinoids plus other components such as terpenoids and pyrolytic by-products were identified and quantified in all samples. Cannabis vapor and smoke was tested for cannabinoid receptor 1 (CB1) binding activity and compared to pure Delta(9)-tetrahydrocannabinol (Delta(9)-THC). The top five major compounds in Bedrocan extracts were Delta(9)-THC, cannabigerol (CBG), terpinolene, myrcene, and cis-ocimene in Bedrobinol Delta(9)-THC, myrcene, CBG, cannabichromene (CBC), and camphene in Bediol cannabidiol (CBD), Delta(9)-THC, myrcene, CBC, and CBG. The major components in Bedrocan vapor (>1.0 mgg) were Delta(9)-THC, terpinolene, myrcene, CBG, cis-ocimene and CBD in Bedrobinol Delta(9)-THC, myrcene and CBD in Bediol CBD, Delta(9)-THC, myrcene, CBC and terpinolene. The major components in Bedrocan smoke (>1.0 mgg) were Delta(9)-THC, cannabinol (CBN), terpinolene, CBG, myrcene and cis-ocimene in Bedrobinol Delta(9)-THC, CBN and myrcene in Bediol CBD, Delta(9)-THC, CBN, myrcene, CBC and terpinolene. There was no statistically significant difference between CB1 binding of pure Delta(9)-THC compared to cannabis smoke and vapor at an equivalent concentration of Delta(9)-THC.

Amphetamines induce tissue factor and impair tissue factor pathway inhibitor role of dopamine receptor type 4.

Amphetamine intake is associated with acute vascular syndromes. Since these events are caused by arterial thrombosis and this in turn is triggered by tissue factor (TF), this study examines whether amphetamines regulate TF in human endothelial cells. Amphetamine (10(-7)-10(-4) molL) enhanced thrombin- and tumour necrosis factor (TNF)-alpha-induced as well as basal TF expression (P 0.029, 0.0003, and 0.003 at maximal concentration), and TNF-alpha-induced plasminogen activator inhibitor (PAI)-1 expression (P 0.003), whereas tissue factor pathway inhibitor expression was impaired (P 0.008). Similarly, 3,4-methylenedioxymethamphetamine (10(-7)-10(-4) molL) enhanced TF expression (P 0.046). These effects were paralleled by an increased TF activity (P 0.002) moreover, clotting time of human plasma was accelerated by supernatant from amphetamine-treated cells (P 0.03). Amphetamine enhanced TF mRNA expression via phosphorylation of the mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase (ERK) and p38 (P 0.03 and 0.033), but not c-Jun NH(2)-terminal kinase (JNK P 0.81). The effect of amphetamine on TF expression was abrogated by the dopamine D4 receptor antagonists L-745,870 and L-750,667, but not D2 or D3 receptor antagonists furthermore, L-745,870 blunted the amphetamine-induced activation of ERK and p38, but not JNK. Amphetamines induce endothelial TF expression via stimulation of dopamine D4 receptor and activation of the MAPKs p38 and ERK. These effects occur at clinically relevant amphetamine concentrations and may account for the increased incidence of acute vascular syndromes after amphetamine consumption.

Agonist binding fraction of dopamine D23 receptors in rat brain a quantitative autoradiographic study.

There has arisen considerable interest in the study of dopamine D(23) agonist binding sites by positron emission tomography (PET), based on the claim that agonist sites represent a functional subset of the total number of sites labeled by more conventional antagonist ligands. To test the basis of this claim, we used quantitative autoradiography to measure the abundance of binding sites of a dopamine D(23) agonist ((3)HNPA) and an antagonist ((3)Hraclopride) in cryosections of rat brain. Saturation binding studies revealed that the B(max) for (3)HNPA was nearly identical to that of (3)Hraclopride in dorsal brain regions, but was 25% less in the ventral striatum and 56% less in the olfactory tubercle. We also tested the displacement of the two ligands by the hallucinogen LSD, which is known to have dopamine agonist properties. Whereas displacement of (3)Hraclopride by increasing LSD concentrations was monophasic, displacement of (3)HNPA was biphasic, suggesting an action of LSD via a subset of dopamine D(23) agonist binding sites. Addition of the stable GTP analogue Gpp(NH)p to the medium abolished 90% of the (3)HNPA binding, and increased (3)Hraclopride binding by 10%, with a shift to the right in the LSD competition curve, suggesting retention of endogenous dopamine in washed cryostat sections. Thus (3)HNPA and (3)Hraclopride binding sites have nearly identical abundances in rat dorsal striatum, but are distinct in the ventral striatum, and with respect to their displacement by LSD.

Cannabinoids and the gut new developments and emerging concepts.

Cannabis has been used to treat gastrointestinal (GI) conditions that range from enteric infections and inflammatory conditions to disorders of motility, emesis and abdominal pain. The mechanistic basis of these treatments emerged after the discovery of Delta(9)-tetrahydrocannabinol as the major constituent of Cannabis. Further progress was made when the receptors for Delta(9)-tetrahydrocannabinol were identified as part of an endocannabinoid system, that consists of specific cannabinoid receptors, endogenous ligands and their biosynthetic and degradative enzymes. Anatomical, physiological and pharmacological studies have shown that the endocannabinoid system is widely distributed throughout the gut, with regional variation and organ-specific actions. It is involved in the regulation of food intake, nausea and emesis, gastric secretion and gastroprotection, GI motility, ion transport, visceral sensation, intestinal inflammation and cell proliferation in the gut. Cellular targets have been defined that include the enteric nervous system, epithelial and immune cells. Molecular targets of the endocannabinoid system include, in addition to the cannabinoid receptors, transient receptor potential vanilloid 1 receptors, peroxisome proliferator-activated receptor alpha receptors and the orphan G-protein coupled receptors, GPR55 and GPR119. Pharmacological agents that act on these targets have been shown in preclinical models to have therapeutic potential. Here, we discuss cannabinoid receptors and their localization in the gut, the proteins involved in endocannabinoid synthesis and degradation and the presence of endocannabinoids in the gut in health and disease. We focus on the pharmacological actions of cannabinoids in relation to GI disorders, highlighting recent data on genetic mutations in the endocannabinoid system in GI disease.

Monolithic silica spin column extraction and simultaneous derivatization of amphetamines and 3,4-methylenedioxyamphetamines in human urine for gas chromatographic-mass spectrometric detection.

A simple, sensitive, and specific method with gas chromatography-mass spectrometry was developed for simultaneous extraction and derivatization of amphetamines (APs) and 3,4-methylenedioxyamphetamines (MDAs) in human urine by using a monolithic silica spin column. All the procedures, such as sample loading, washing, and elution were performed by centrifugation. APs and MDAs in urine were adsorbed on the monolithic silica and derivatized with propyl chloroformate in the column. Methamphetamine-d(5) was used as an internal standard. The linear ranges were 0.01-5.0 microg mL(-1) for methamphetamine (MA) and 3,4-methylenedioxymethamphetamine (MDMA) and 0.02-5.0 microg mL(-1) for amphetamine (AP) and 3,4-methylenedioxyamphetamine (MDA) (coefficient of correlation > or 0.995). The recovery of APs and MDAs in urine was 84-94%, and the relative standard deviation of the intra- and interday reproducibility for urine samples containing 0.1, 1.0, and 4.0 microg mL(-1) of APs and MDAs ranged from 1.4% to 13.6%. The lowest detection limit (signal-to-noise ratio > or 3) in urine was 5 ng mL(-1) for MA and MDMA and 10 ng mL(-1) for AP and MDA. The proposed method can be used to perform simultaneous extraction and derivatization on spin columns that have been loaded with a small quantity of solvent by using centrifugation.

Cannabis and caries--does regular cannabis use increase the risk of caries in cigarette smokers

The use of cannabis by adolescents in Switzerland has almost doubled in the past decade. Empirical observations in private dental practices indicate that cannabis users have more carious lesions than those who do not use cannabis. The aim of this study was to investigate the hypothesis that regular cannabis use increases the risk of caries because of hyposalivation or lifestyle. Forty-three regular cannabis users were enrolled in the test group and 42 tobacco smokers were used as a negative control group. All subjects were 18-25 years old. Data were obtained using a standardized questionnaire and a clinical examination. There was no significant difference between groups in decayed and filled surfaces (DFS), saliva flow rate and plaque and gingival indices. The cannabis group had, however, significantly higher DS (decayed surface) values (p 0.0001) and significantly lower frequencies of daily tooth brushing and dental control visits (p < 0.0001) than the control group. Additionally, the cannabis group reported a significantly higher consumption of sugar-containing beverages than the control group (p 0.0078). To obtain more objective data relations, the DS values of male cannabis users were also compared with those of Swiss military recruits found in another study. The cannabis users had more caries on smooth surfaces than the military recruits. Although comparison with epidemiological data suggested that the prevalence of caries on smooth surfaces is elevated in cannabis users, DFS data indicated that cannabis users do not have an increased risk of caries. Lifestyle combined with short-term hyposalivation after delta-9-tetrahydrocannabinol consumption is the most probable cause of the high prevalence of caries on smooth surfaces in cannabis users. Further studies are needed to investigate the effects of cannabis use on oral health.

Plasma and urine profiles of Delta9-tetrahydrocannabinol and its metabolites 11-hydroxy-Delta9-tetrahydrocannabinol and 11-nor-9-carboxy-Delta9-tetrahydrocannabinol after cannabis smoking by male volunteers to estimate recent consumption by athletes.

Since 2004, cannabis has been prohibited by the World Anti-Doping Agency for all sports competitions. In the years since then, about half of all positive doping cases in Switzerland have been related to cannabis consumption. In doping urine analysis, the target analyte is 11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol (THC-COOH), the cutoff being 15 ngmL. However, the wide urinary detection window of the long-term metabolite of Delta(9)-tetrahydrocannabinol (THC) does not allow a conclusion to be drawn regarding the time of consumption or the impact on the physical performance. The purpose of the present study on light cannabis smokers was to evaluate target analytes with shorter urinary excretion times. Twelve male volunteers smoked a cannabis cigarette standardized to 70 mg THC per cigarette. Plasma and urine were collected up to 8 h and 11 days, respectively. Total THC, 11-hydroxy-Delta(9)-tetrahydrocannabinol (THC-OH), and THC-COOH were determined after hydrolysis followed by solid-phase extraction and gas chromatographymass spectrometry. The limits of quantitation were 0.1-1.0 ngmL. Eight puffs delivered a mean THC dose of 45 mg. Plasma levels of total THC, THC-OH, and THC-COOH were measured in the ranges 0.2-59.1, 0.1-3.9, and 0.4-16.4 ngmL, respectively. Peak concentrations were observed at 5, 5-20, and 20-180 min. Urine levels were measured in the ranges 0.1-1.3, 0.1-14.4, and 0.5-38.2 ngmL, peaking at 2, 2, and 6-24 h, respectively. The times of the last detectable levels were 2-8, 6-96, and 48-120 h. Besides high to very high THC-COOH levels (245 - 1,111 ngmL), THC (3 - 8 ngmL) and THC-OH (51 - 246 ngmL) were found in 65 and 98% of cannabis-positive athletes urine samples, respectively. In conclusion, in addition to THC-COOH, the pharmacologically active THC and THC-OH should be used as target analytes for doping urine analysis. In the case of light cannabis use, this may allow the estimation of more recent consumption, probably influencing performance during competitions. However, it is not possible to discriminate the intention of cannabis use, i.e., for recreational or doping purposes. Additionally, pharmacokinetic data of female volunteers are needed to interpret cannabis-positive doping cases of female athletes.

An in vivo microdialysis assessment of concurrent MDMA and cocaine administration in Sprague-Dawley rats.

Despite the popularity of polysubstance abuse among recreational methylendioxymethamphetamine (MDMA) users, relatively few controlled experimental studies have documented the neurobehavioral effects of MDMA in combination with other abused substances. In this study, the combined acute effects of MDMA and cocaine were examined by conducting in vivo microdialysis in the rat nucleus accumbens while simultaneously monitoring locomotor activity. Male Sprague-Dawley rats were administered cocaine (10 or 20 mgkg, i.p.), MDMA (1.5 or 3.0 mgkg, i.p.), or one of four combinations of cocaine and MDMA during microdialysis experiments. Locomotor activity was monitored, and dialysis samples were collected every 30 min for 3 h prior to injections, for one 30-min period following saline injections, and for an additional 3-h period following drug injections. Samples were analyzed for dopamine content by high-performance liquid chromatography with electrochemical detection. Significant differences in locomotor activity and dopamine efflux were found among treatment groups, with some MDMAcocaine combinations producing significantly greater increases compared to single doses of cocaine or MDMA within the first 30 min after injection. Considering the popularity of polysubstance use among recreational MDMA users, the clinical implications of the current findings warrant further investigation.

Antagonism at serotonin 5-HT(2A) receptors modulates functional activity of frontohippocampal circuit.

Several second-generation antipsychotics are characterised by a significant antagonistic effect at serotonin 5-HT(2A) receptors (5-HT(2A)R), a feature that has been associated with lower incidence of extra-pyramidal symptoms and a putative amelioration of positive and negative symptoms experienced by schizophrenic patients. However, the neurofunctional substrate of 5-HT(2A) antagonism and its exact contribution to the complex pharmacological profile of these drugs remain to be elucidated. Here, we used pharmacological magnetic resonance imaging to map the modulatory effects of the selective 5-HT(2A)R antagonist Ml00907 on the spatiotemporal patterns of brain activity elicited by acute phencyclidine (PCP) challenge in the rat. PCP is a non-competitive NMDA receptor antagonist that induces dysregulation of corticolimbic glutamatergic neurotransmission and produces cognitive impairment and psychotic-like symptoms reminiscent of those observed in schizophrenia. Pre-administration of M100907 produced focal and region-dependent attenuation of PCP-induced response in frontoseptohippocampal areas. As early studies highlighted a permissive role of 5-HT(2A)R on frontal dopamine release, the role of post-synaptic dopamine D(1) receptors on PCP-induced response was examined by using the potent antagonist SCH23390. Interestingly, SCH23390 did not affect PCPs response in any of the regions examined. This finding rules out a significant contribution of dopamine in the functional changes mapped and, indirectly, the inhibitory effect of M100907, in favour of a glutamatergic origin. Our data expand recent evidence suggesting a key role of 5-HT(2A)R in modulating glutamate-mediated cognitive performance in the prefrontal cortex and highlight the whole frontoseptohippocampal circuit as a key functional substrate of 5-HT(2A)R antagonism in normal and disease states.

Tumour necrosis factor alpha suppression by MDMA is mediated by peripheral heteromeric nicotinic receptors.

MDMA is an illegal drug widely used by young people. The present study aimed to determine the involvement of different nicotinic acetylcholine receptor (nAChR) subtypes in the suppressive effect of MDMA in TNF-alpha production. Dihydrobetaerythroidine (antagonist of heteromeric nAChR), and hexamethonium (antagonist of peripheral nAChR), fully antagonized the effect of MDMA. Conversely, methyllycaconitine (antagonist of homomeric nAChR), did not modify it. From in vitro experiments, a direct effect was ruled out. In this study we provide the first evidence that in rodents MDMA impairs the production of TNF-alpha by activation of heteromeric nAChR expressing beta-2 subunits located in the periphery.

Direct detection of Delta9-tetrahydrocannabinol in saliva using a novel homogeneous competitive immunoassay with fluorescence quenching.

For the detection of the major active component of cannabis, Delta9-tetrahydrocannabinol (THC) in aqueous samples, a homogeneous competitive immunoassay based on fluorescence quenching induced by fluorescence resonance energy transfer (FRET) has been developed. The fluorescence of anti-THC-antibody, labeled with fluorescence dye DY-481XL, can be quenched after its binding to THC-BSA-quencher conjugate (bovine serum albumin coupled with THC and another fluorescence dye, DYQ-661, as quencher). This quenching effect is inhibited when the antibodies bind to free THC in aqueous sample, thus competing for binding sites with the THC-BSA-quencher conjugate. The extent of the inhibition corresponds to the concentration of THC in the samples. The assay principle is simple and the test duration is within 10 min. The detection limit for THC in buffer was 2 ng mL(-1). In pooled saliva samples a detection limit of 50 ng mL(-1) was achieved.

Depressive and anxiety symptomatology in ecstasy users the relative contribution of genes, trauma, life stress and drug use.

Previous research has identified elevated rates of depressive and anxiety symptoms amongst ecstasy users however, few studies have examined which factors increase the likelihood of experiencing such symptoms. The current study aimed to determine the relationship between ecstasy use and depressiveanxiety symptomatology after controlling for known environmental and genetic (polymorphism of the serotonin transporter gene) risk factors for depression and anxiety disorders. Participants consisted of a community sample of 184 18-35-year olds who had taken ecstasy at least once in the past 12 months. Participants completed an interview and questionnaires and provided a saliva sample. Mood symptoms were assessed using the Mood and Anxiety Symptom Questionnaire. Timeline methods were used to collect information on lifetime and recent ecstasy use, as well as recent other drug use and life stress. Trauma exposure was measured using the Composite International Diagnostic Interview--Trauma List. Genomic DNA was extracted from participant saliva samples. Neither lifetime nor recent ecstasy use was associated with the severity of current mood symptoms, either alone or in combination with genetic risk factors. Rather, lifetime trauma, recent stressful life events, the frequency of tobacco use and recent polydrug use significantly predicted the severity of depressive and anxiety symptoms. These results highlight the need to consider the role of environmental factors when examining the relationship between ecstasy use and mood symptoms. Whether ecstasy exacerbates such symptoms in vulnerable individuals requires further investigation using prospective designs.

JWH018, a common constituent of Spice herbal blends, is a potent and efficacious cannabinoid CB receptor agonist.

Spice is an herbal blend primarily marketed in Europe as a mild hallucinogen with prominent cannabis-like effects and as a legal alternative to cannabis. However, a recent report identified a number of synthetic additives in samples of Spice. One of these, the indole derivative JWH018, is a ligand for the cannabinoid receptor 1 (CB(1)) cannabinoid receptor and inhibits cAMP production in CB(1) receptor-expressing CHO cells. Other effects of JWH018 on CB(1) receptor-mediated signalling are not known, particularly in neurons. Here we have evaluated the signalling pathways activated by JWH018 at CB(1) receptors. We investigated the effects of JWH018 on neurotransmission in cultured autaptic hippocampal neurons. We further analysed its activation of ERK12 mitogen activated protein kinase (MAPK) and internalization of CB(1) receptors in HEK293 cells stably expressing this receptor. In cultured autaptic hippocampal neurons, JWH018 potently inhibited excitatory postsynaptic currents (IC(50) 14.9 nM) in a concentration- and CB(1) receptor-dependent manner. Furthermore, it increased ERK12 MAPK phosphorylation (EC(50) 4.4 nM). We also found that JWH018 potently induced rapid and robust CB(1) receptor internalization (EC(50) 2.8 nM t(12) 17.3 min). JWH018, a prominent component of several herbal preparations marketed for their psychoactivity, is a potent and effective CB(1) receptor agonist that activates multiple CB(1) receptor signalling pathways. Thus, it is likely that the subjective effects of Spice are due to activation of cannabinoid CB(1) receptors by JWH018, added to this herbal preparation.

Effect of subchronic phencyclidine administration on sucrose preference and hippocampal parvalbumin immunoreactivity in the rat.

Persistent blockade of NMDA receptor function by repeated phencyclidine dosing produces pathophysiological changes that model deficits observed in schizophrenia. The present study investigates the effects of subchronic phencyclidine administration (PCP 2 or 5mgkg bi-daily for 7 days followed by a drug-free period) on sucrose choice, a measure of anhedonia. Sucrose preference in a two-bottle sucrose-water choice test was assessed 1 and 2 weeks after PCP. Results showed no differences in sucrose intake between PCP rats and controls, nor a difference in water intake or total volume of liquid consumed at either time-point. Six weeks post-PCP, analysis of brains showed a reduction in expression of parvalbumin immunoreactive neurons in the hippocampus with significant reductions localised to the CA1 and CA23 regions. These results demonstrate that while subchronic PCP may not be a valid model for the negative symptom of anhedonia observed in schizophrenia, it induces pathology in the brain in hippocampal subregions that are reminiscent of changes observed in schizophrenia.

c-Fos identification of neuroanatomical sites associated with haloperidol and clozapine disruption of maternal behavior in the rat.

Rat maternal behavior is a complex social behavior. Most antipsychotic drugs disrupt active maternal responses (e.g., pup retrieval, pup licking and nest building). Our previous work shows that typical antipsychotic haloperidol disrupts maternal behavior by blocking dopamine D(2) receptors, whereas atypical clozapine works by blocking 5-HT(2A2C) receptors. The present study used c-Fos immunohistochemistry technique, together with pharmacological tools and behavioral observations, and delineated the neuroanatomical bases of the disruptive effects of haloperidol and clozapine. Postpartum female rats were treated with haloperidol (0.2 mgkg sc) or clozapine (10.0 mgkg sc), with or without pretreatment of quinpirole (a selective dopamine D(2)D(3) agonist, 1.0 mgkg sc) or 2,5-dimethoxy-4-iodo-amphetamine (DOI, a selective 5-HT(2A2C) agonist, 2.5 mgkg sc). They were then sacrificed 2 h later after a maternal behavior test was conducted. Brain regions that have been previously implicated in the regulation of rat maternal behavior andor in the antipsychotic action were examined. Behaviorally, both haloperidol and clozapine disrupted pup retrieval, pup licking and nest building. Pretreatment of quinpirole, but not DOI, reversed the haloperidol-induced disruptions. In contrast, pretreatment of DOI, but not quinpirole, reversed the clozapine-induced deficits. Neuroanatomically, the nucleus accumbens (both the shell and core), dorsolateral striatum and lateral septum showed increased c-Fos expression to the treatment of haloperidol. In contrast, the nucleus accumbens shell showed increased expression of c-Fos to the treatment of clozapine. More importantly, pretreatment of quinpirole and DOI produced opposite response profiles in the brain regions where haloperidol and clozapine had an effect. Based on these findings, we concluded that haloperidol disrupts active maternal behavior primarily by blocking dopamine D(2) receptors in a neural circuitry involving the nucleus accumbens, dorsolateral striatum and lateral septum. In contrast, clozapine appears to disrupt maternal behavior mainly by blocking serotonin 5-HT(2A2C) receptors in the nucleus accumbens shell.

Multiple hits during postnatal and early adulthood periods disrupt the normal development of sensorimotor gating ability in rats.

In the present study, we evaluated a multiple-hit animal model of schizophrenia in an attempt to capture the complex interactions among various adverse developmental factors in schizophrenia. Sprague-Dawley rats were assigned to receive either repeated daily 3-h maternal separation for eight days (first hit) on postnatal days (PND) 3 to 10, andor avoidance conditioning for six days (second hit) on PND 49-56, andor repeated phencyclidine treatment (third hit, 3.0 mgkg, sc) immediately after each daily avoidance conditioning. Prepulse inhibition (PPI) of acoustic startle reflex was assessed at late adolescence (PND 41-43) and early adulthood (PND 62-63). The change in %PPI from the adolescence phase to adulthood phase was used to index the maturation-related improvement of sensorimotor gating ability. Maternal separation, avoidance conditioning and PCP treatment had a complex three-way interaction on the functional improvement of sensorimotor gating. Maternal separation impaired PPI improvement preferentially in the saline rats that were not subjected to avoidance conditioning. Avoidance conditioning had no effect on PPI improvement in the non-maternally separated rats, but restored the maternal separation-induced disruption. However, this restoration effect was abolished by PCP treatment. The present study also identified a number of behavioral, emotional and learning abnormalities caused by these three developmental insults which may precede their interactive disruption of normal development of sensorimotor gating ability.

Cannabinoid inhibition of macrophage migration to the trans-activating (Tat) protein of HIV-1 is linked to the CB(2) cannabinoid receptor.

Macrophages and macrophage-like cells are important targets of HIV-1 infection at peripheral sites and in the central nervous system. After infection, these cells secrete a plethora of toxic factors, including the viral regulatory trans-activating protein (Tat). This protein is highly immunogenic and also serves as a potent chemoattractant for monocytes. In the present study, the exogenous cannabinoids delta-9-tetrahydrocannabinol (THC) and (-)-cis-3-2-hydroxy-4-(1,1-dimethylheptyl)phenyl-trans-4-(3-hydroxypropyl) cyclohexanol (CP55940) were shown to significantly inhibit migration of human U937 macrophage-like cells to the Tat protein in a concentration-related manner. The CB(1) receptor-selective agonist N-(2-chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide (ACEA) had no effect on Tat-mediated migration. In contrast, the CB(2) receptor-selective agonist (1R,3R)-1-4-(1,1-dimethylheptyl)-2,6-dimethoxyphenyl-3-methylcyclohexanol (O-2137) exerted a concentration-related inhibition of U937 cell migration in response to Tat. Pharmacological blockage of CB(1) receptor signaling using the antagonist 5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-N-(1-piperidyl)pyrazole-3-carboxamide hydrochloride (SR141716A) had no effect on CP55940-mediated inhibition of macrophage migration to Tat, whereas treatment with the CB(2) receptor antagonist (1S-endo)-5-(4-chloro-3-methylphenyl)-1-((4-methylphenyl)methyl)-N-(1,3,3-trimethylbicyclo(2.2.1)hept-2-yl)-1H-pyrazole-3-carboxamide (SR144528) reversed the CP55940-mediated inhibition of migration. In addition, THC had no inhibitory effect on U937 migration to Tat after small interfering RNA knockdown of the CB(2) receptor. Collectively, the pharmacological and biochemical knockdown data indicate that cannabinoid-mediated modulation of macrophage migration to the HIV-1 Tat protein is linked to the CB(2) cannabinoid receptor. Furthermore, these results suggest that the CB(2) cannabinoid receptor has potential to serve as a therapeutic target for ablation of HIV-1-associated untoward inflammatory response.

Rapid assembly of the salvileucalin B norcaradiene core.

Preparation of the polycyclic core of the cytotoxic natural product salvileucalin B is described. The key feature of this synthetic strategy is a copper-catalyzed intramolecular arene cyclopropanation to provide the central norcaradiene. These studies lay the foundation for continued investigations toward an enantioselective total synthesis of 1.

Dose-related effects of MDMA on psychomotor function and mood before, during, and after a night of sleep loss.

3,4-methylenedioxymethamphetamine (MDMA) is known to improve psychomotor function and mood when measured during daytime. However, MDMA users tend to take this drug at dance parties while staying awake for prolonged periods of time. This study was designed to assess dose-related residual effects of MDMA on psychomotor function and mood after a night without sleep. Sixteen recreational MDMA users received single doses of 25, 50, and 100 mg MDMA in a randomized, double-blind, placebo-controlled cross-over study. Results showed that sleep loss significantly impaired psychomotor function. MDMA generally did not affect performance but did improve rapid information processing at the highest dose in the morning after administration. In the evening, MDMA also increased subjective ratings of positive mood at every dose and subjective arousal at the highest dose. These subjective effects were no longer present after a night of sleep loss. It is concluded that sleep deprivation impairs psychomotor function and that stimulant effects of MDMA are not sufficient to compensate for this impairment.

The discriminative effects of the kappa-opioid hallucinogen salvinorin A in nonhuman primates dissociation from classic hallucinogen effects.

The widely available hallucinogen salvinorin A is a unique example of a plant-derived compound selective for kappa-opioid receptors and may produce effects distinct from those of other compounds with classic hallucinogenic or dissociative properties which are also abused in humans. The objective of this study is to characterize the salvinorin A discriminative cue in nonhuman primates with high kappa-receptor genetic homology to humans. Adult rhesus monkeys (n 3) were trained to discriminate salvinorin A (0.015 mgkg, s.c.) from vehicle, in a food-reinforced operant discrimination assay. Parallel studies, using unconditioned behavioral endpoints (facial relaxation and ptosis) also evaluated the kappa-opioid receptor mediation of salvinorin A in vivo function. Monkeys trained to discriminate salvinorin A generalized structurally diverse, centrally penetrating kappa-agonists (bremazocine, U69,593, and U50,488). By contrast, mu- and delta-opioid agonists (fentanyl and SNC80, respectively) were not generalized, nor were the serotonergic 5HT2 hallucinogen psilocybin or the dissociative N-methyl-D-aspartic acid antagonist, ketamine. The discriminative effects of salvinorin A were blocked by the opioid antagonist quadazocine (0.32 mgkg), but not by the 5HT2 antagonist ketanserin (0.1 mgkg). Consistent with these findings, salvinorin and kappa-agonists (e.g., U69,593) produce effects in the unconditioned endpoints (e.g., ptosis), whereas psilocybin was inactive. These findings support the conclusion that the interoceptivediscriminative cue produced by salvinorin A is mediated by agonism at kappa-receptors and is mechanistically distinct from that produced by a classic serotonergic hallucinogen.

CB1 cannabinoid receptors increase neuronal precursor proliferation through AKTglycogen synthase kinase-3betabeta-catenin signaling.

The endocannabinoid system is involved in the regulation of many physiological effects in the central and peripheral nervous system. Recent findings have demonstrated the presence of a functional endocannabinoid system within neuronal progenitors located in the hippocampus and ventricularsubventricular zone that participates in the regulation of cell proliferation. It is presently unknown whether the endocannabinoid system exerts a widespread effect on neuronal precursors from different neurogenic regions, and very little is known about the signaling by which it regulates neuronal precursor proliferation. Herein, we demonstrate the presence of cannabinoid CB(1) receptors in granule cell precursors (GCPs) during early cerebellar development. Activation of CB(1) receptors by HU-210 promoted GCP proliferation in vitro, an effect that was prevented by a selective CB(1) antagonist. Accordingly, in vivo experiments showed that GCP proliferation was increased by chronic HU-210 treatment and that in CB(1)-deficient mice cell proliferation was significantly lower than in wild-type littermates, indicating that the endocannabinoid system is physiologically involved in regulation of GCP proliferation. The pro-proliferative effect of cannabinoids in GCPs was mediated through the CB(1)AKTglycogen synthase kinase-3betabeta-catenin pathway. Involvement of this pathway was also observed in cultures of neuronal precursors from the subventricular zone, suggesting that this pathway may be a general mechanism by which endocannabinoids regulate proliferation of neuronal precursors. These observations suggest that endocannabinoids constitute a new family of lipid signaling cues that may exert a widespread effect on neuronal precursor proliferation during brain development.

CoMFA analyses of C-2 position salvinorin A analogs at the kappa-opioid receptor provides insights into epimer selectivity.

The highly potent and kappa-opioid (KOP) receptor-selective hallucinogen Salvinorin A and selected analogs have been analyzed using the 3D quantitative structure-affinity relationship technique Comparative Molecular Field Analysis (CoMFA) in an effort to derive a statistically significant and predictive model of salvinorin affinity at the KOP receptor and to provide additional statistical support for the validity of previously proposed structure-based interaction models. Two CoMFA models of Salvinorin A analogs substituted at the C-2 position are presented. Separate models were developed based on the radioligand used in the kappa-opioid binding assay, (3)Hdiprenorphine or (125)I6 beta-iodo-3,14-dihydroxy-17-cyclopropylmethyl-4,5 alpha-epoxymorphinan ((125)IIOXY). For each dataset, three methods of alignment were employed a receptor-docked alignment derived from the structure-based docking algorithm GOLD, another from the ligand-based alignment algorithm FlexS, and a rigid realignment of the poses from the receptor-docked alignment. The receptor-docked alignment produced statistically superior results compared to either the FlexS alignment or the realignment in both datasets. The (125)IIOXY set (Model 1) and (3)Hdiprenorphine set (Model 2) gave q(2) values of 0.592 and 0.620, respectively, using the receptor-docked alignment, and both models produced similar CoMFA contour maps that reflected the stereoelectronic features of the receptor model from which they were derived. Each model gave significantly predictive CoMFA statistics (Model 1 PSET r(2)0.833 Model 2 PSET r(2)0.813). Based on the CoMFA contour maps, a binding mode was proposed for amine-containing Salvinorin A analogs that provides a rationale for the observation that the beta-epimers (R-configuration) of protonated amines at the C-2 position have a higher affinity than the corresponding alpha-epimers (S-configuration).

Simultaneous quantification of cannabinoids and metabolites in oral fluid by two-dimensional gas chromatography mass spectrometry.

Development and validation of a method for simultaneous identification and quantification of Delta9-tetrahydrocannabinol (THC), cannabidiol (CBD), cannabinol (CBN), and metabolites 11-hydroxy-THC (11-OH-THC) and 11-nor-9-carboxy-THC (THCCOOH) in oral fluid. Simultaneous analysis was problematic due to different physicochemical characteristics and concentration ranges. Neutral analytes, such as THC and CBD, are present in ngmL, rather than pgmL concentrations, as observed for the acidic THCCOOH biomarker in oral fluid. THCCOOH is not present in cannabis smoke, definitively differentiating cannabis use from passive smoke exposure. THC, 11-OH-THC, THCCOOH, CBD, and CBN quantification was achieved in a single oral fluid specimen collected with the Quantisal device. One mL oral fluidbuffer solution (0.25 mL oral fluid and 0.75 mL buffer) was applied to conditioned CEREX Polycrom THC solid-phase extraction (SPE) columns. After washing, THC, 11-OH-THC, CBD, and CBN were eluted with hexaneacetoneethyl acetate (603020, vvv), derivatized with N,O-bis-(trimethylsilyl)trifluoroacetamide and quantified by two-dimensional gas chromatography electron ionization mass spectrometry (2D-GCMS) with cold trapping. Acidic THCCOOH was separately eluted with hexaneethyl acetateacetic acid (75252.5, vvv), derivatized with trifluoroacetic anhydride and hexafluoroisopropanol, and quantified by the more sensitive 2D-GCMS-electron capture negative chemical ionization (NCI-MS). Linearity was 0.5-50 ngmL for THC, 11-OH-THC, CBD and 1-50 ngmL for CBN. The linear dynamic range for THCCOOH was 7.5-500 pgmL. Intra- and inter-assay imprecision as percent RSD at three concentrations across the linear dynamic range were 0.3-6.6%. Analytical recovery was within 13.8% of target. This new SPE 2D-GCMS assay achieved efficient quantification of five cannabinoids in oral fluid, including pgmL concentrations of THCCOOH by combining differential elution, 2D-GCMS with electron ionization and negative chemical ionization. This method will be applied to quantification of cannabinoids in oral fluid specimens from individuals participating in controlled cannabis and Sativex (50% THC and 50% CBD) administration studies, and during cannabis withdrawal.

ERK signalling pathway is not involved in PSA-NCAM-dependent alterations of hippocampal plasticity evoked by CB1 receptor activation.

The present study investigated the potential role of the extracellular signal-regulated kinase (ERK) pathway in the alternation of polysialylated neural cell adhesion molecule (PSA-NCAM) expression and proliferation rates in the dentate gyrus (DG) evoked by activation of the CB1 receptor. When given at a dose of 0.1 mgkg, the CB1 receptor agonist, 3-(1,1-dimethylheptyl)-11-hydroxy-Delta(8)-tetrahydrocannabinol (HU-210), increased the levels of the phosphorylated forms of ERK (pERK1 and pERK2) in the hippocampus when measured 30 min after injection. This HU-210-induced effect was inhibited by alpha-amino(4-aminophenyl)thiomethylene-2-(trifluoromethyl) benzeneacetonitrile (SL327, 30 mgkg) - an inhibitor of mitogen-activated protein kinase kinase (MEK12), the upstream kinase of ERK - given 1 h before HU-210 administration. Additionally, SL327 alone significantly attenuated the basal level of both pERK1 and pERK2. HU-210 (0.1 mgkg) decreased the number of PSA-NCAM-immunoreactive (IR) cells but did not affect the rate of proliferation, which was analyzed as the number of Ki-67-IR cells measured in the DG 2 days after HU-210 administration. The data indicated that SL327 (30 mgkg) alone decreased the number of PSA-NCAM-IR cells 2 days after treatment. Joint administration of SL327 and HU-210 decreased the number of PSA-NCAM cells more robustly than did the administration of either alone. In addition, SL327 did not decrease the number of Ki-67-IR cells, while pretreatment with SL327 1 h before HU-210 administration did. These results suggest that stimulation of the ERK cascade caused by CB1 receptor activation is not involved in hippocampal plasticity governed by PSA-NCAM expression.

The influence of 5-HTT and COMT genotypes on verbal fluency in ecstasy users.

Deficits in verbal fluency associated with ecstasy use have been well established however, the mechanisms underlying this impairment have yet to be elucidated. In this study we investigated for the first time whether there was a disproportionate impairment in two cognitive subcomponents of verbal fluency clustering (ability to generate words within the same subcategory) and switching (ability to change the subcategory). We also investigated a possible association between ecstasy use and verbal fluency in subjects genotyped for 5-HTT (5-HTTLPR and 5-HTTVNTR) and COMT (val(108158)met, rs165599 and rs2097603) polymorphisms, in order to find a potential implication of genetic factors. Ecstasy polydrug users (n 30) and non-ecstasy users (n 41) were evaluated in both semantic and phonemic fluency. Results showed that ecstasy users had poorer semantic (but not phonemic) fluency performance than controls. Detailed analysis of clustering and switching performance revealed that this impairment was associated with poorer clustering mechanisms. Clustering was also modulated by the COMT rs165599 polymorphism independently of the group. A specific effect of the 5-HTTLPR polymorphism on switching performance was also found, with ss carriers performing significantly worse than ls and ll carriers, suggesting a serotonin modulation of frontal-executive flexibility. Based on the impaired clustering and switching strategies observed in ecstasy users, it might be proposed that both semantic knowledge and retrieval are impaired in this population. The verbal fluency deficit in ecstasy users may be attributable to a disruption of frontal-striatal circuits directly related with the serotonin function as well as a depletion of lexical-semantic stores mediated by temporal structures.

High throughput analysis of drugs of abuse in hair by combining purposely designed sample extraction compatible with immunometric methods used for drug testing in urine.

Drug testing in hair usually requires a rather complex sample treatment before drugs are amenable to analysis by either immunological andor chromatographic coupled to mass spectrometry methods. Immunological methods applied are usually dedicated to hair analysis as analytes present in this matrix are not always the same present in urine. Comedical s.a.s. laboratories recently commercialized reagents (VMA-T) purposely designed for hair sample treatment which are compatible with current immunometric methods used for urine drug testing. This is possible as some analytes (6-MAM and cocaine) present in hair after sample treatment are converted to those detected in urine (morphine and benzoylecgonine). A correlation study for several drug classes performed in two laboratories with 32 clinical and 12 spiked drug free (controls) hair samples shows that implementation of the method on clinical chemistry analyzers is easy and that results obtained by different operators and instruments are comparable and reproducible. The main advantage of VMA-T method is the possibility to simultaneously extract from hair main drug classes, in a period of time lower than 2h and its compatibility with immunological methods applied in urine drug testing.

Phencyclidine (PCP) produces sexually dimorphic effects on voluntary sucrose consumption and elevated plus maze behavior.

Previous research in our laboratory indicates that the psychotomimetic drug phencyclidine (PCP) reduces voluntary sucrose consumption in male rats, potentially modeling the schizophrenic symptom of anhedonia. Given reports from the clinical literature that schizophrenia has a later age of onset and more favorable outcome in females, PCP might be expected to have sexually dimorphic effects in animal models of schizophrenia such as PCP-induced decreases in voluntary sucrose consumption. Young adult (66 days old) and adult (109 days old) male and female rats were trained to drink sucrose during a 30 minday presentation protocol. On the day prior to the test day, animals were treated with PCP (15 mgkg) or saline four hours after the onset of the sucrose presentation (20 h prior to the sucrose on the test day). PCP decreased sucrose consumption on the test day similarly in adult males and females, although females also showed decreased water consumption. In young animals, PCP decreased sucrose consumption in males but not in females. These results are consistent with the prediction that females will be less sensitive to the schizophrenia-like behavioral effects of PCP. In a separate study, the same animals were tested in an elevated plus maze one to two months after testing for voluntary sucrose consumption. Significant sex x drug interaction effects on a number of measures in the elevated plus maze indicated that prior exposure to PCP had an anxiolytic effect in females and an anxiogenic effect in males. While unexpected, this finding indicates an additional sexually dimorphic effect of PCP on behavior and its potential relevance to the PCP model of schizophrenia is discussed.

Semi-quantitative analysis of drugs of abuse, including tetrahydrocannabinol in hair using aqueous extraction and immunoassay.

A semi-quantitative analytical screening procedure for the determination of cocaine, amphetamines, opiates, and delta-9-tetrahydrocannabinol in hair has been developed. The procedure employs an aqueous extraction buffer, uses only 10mg of hair, requires 2h of incubation for the extraction to occur, and multiple drug classes can be screened using enzyme linked immunosorbent assays. Hair calibration standards were prepared around the recommended cut-off concentrations of the Society of Hair Testing. All drug classes showed excellent linearity over the concentration range tested, indicating that immunochemical screening can be used in a semi-quantitative mode for hair analysis using an aqueous buffer, rapid extraction and a small amount of hair.