Discriminative stimulus properties of 1.25mg/kg clozapine in rats: Mediation by serotonin 5-HT2 and dopamine D4 receptors.

The atypical antipsychotic drug clozapine remains one of most effective treatments for schizophrenia, given a lack of extrapyramidal side effects, improvements in negative symptoms, cognitive impairment, and in symptoms in treatment-resistant schizophrenia. The adverse effects of clozapine, including agranulocytosis, make finding a safe clozapine-like a drug a goal for drug developers. The drug discrimination paradigm is a model of interoceptive stimulus that has been used in an effort to screen experimental drugs for clozapine-like atypical antipsychotic effects. The present study was conducted to elucidate the receptor-mediated stimulus properties that form this clozapine discriminative cue by testing selective receptor ligands in rats trained to discriminate a 1.25mg/kg dose of clozapine from vehicle in a two choice drug discrimination task. Full substitution occurred with the 5-HT2A inverse agonist M100907 and the two preferential D4/5-HT2/α1 receptor antagonists Lu 37-114 ((S)-1-(3-(2-(4-(1H-indol-5-yl)piperazin-1-yl)ethyl)indolin-1-yl)ethan-1-one) and Lu 37-254 (1-(3-(4-(1H-indol-5-yl)piperazin-1-yl)propyl)-3,4-dihydroquinolin-2(1H)-one). Partial substitution occurred with the D4 receptor antagonist Lu 38-012 and the α1 adrenoceptor antagonist prazosin. Drugs selective for 5-HT2C, 5-HT6 muscarinic, histamine H1, and benzodiazepine receptors did not substitute for clozapine. The present findings suggest that 5-HT2A inverse agonism and D4 receptor antagonism mediate the discriminative stimulus properties of 1.25mg/kg clozapine in rats, and further confirm that clozapine produces a complex compound discriminative stimulus.

Pharmacological characterization of repeated administration of the first generation abused synthetic cannabinoid CP47,497.

A series of in vivo and in vitro assays were conducted to characterize the pharmacological effects of the first generation abused synthetic cannabinoid CP47,497, a racemic bicyclic cannabinoid that is similar in structure to the potent, high-efficacy synthetic cannabinoid CP55,940. CP47,497 was less efficacious than CP55,940 in activating G-proteins and dose-dependently produced common CB1 receptor-dependent pharmacological effects (i.e. catalepsy, hypothermia, antinociception, and hypolocomotion). CP47,497 also substituted for Δ9-tetrahydrocannabinol (THC) in the mouse drug discrimination, indicating that both drugs elicited a similar interceptive stimulus. The pharmacological effects of CP47,497 underwent tolerance following repeated administration and showed cross-tolerance following repeated THC administration, further suggesting a common cannabimimetic mechanism of action. Finally, the CB1 receptor antagonist rimonabant precipitated similar magnitudes of somatic withdrawal responses in mice treated repeatedly with THC or CP47,497. Taken together, these data verify the acute cannabimimetic effects of CP47,497, and indicate tolerance and dependence following repeated administration. The assays used here provide a straightforward approach to characterize the emerging next generation of abused synthetic cannabinoids.

The Selective Monoacylglycerol Lipase Inhibitor MJN110 Produces Opioid-Sparing Effects in a Mouse Neuropathic Pain Model.

Serious clinical liabilities associated with the prescription of opiates for pain control include constipation, respiratory depression, pruritus, tolerance, abuse, and addiction. A recognized strategy to circumvent these side effects is to combine opioids with other antinociceptive agents. The combination of opiates with the primary active constituent of cannabis (Δ(9)-tetrahydrocannabinol) produces enhanced antinociceptive actions, suggesting that cannabinoid receptor agonists can be opioid sparing. Here, we tested whether elevating the endogenous cannabinoid 2-arachidonoylglycerol through the inhibition of its primary hydrolytic enzyme monoacylglycerol lipase (MAGL), will produce opioid-sparing effects in the mouse chronic constriction injury (CCI) of the sciatic nerve model of neuropathic pain. The dose-response relationships of i.p. administration of morphine and the selective MAGL inhibitor 2,5-dioxopyrrolidin-1-yl 4-(bis(4-chlorophenyl)methyl)piperazine-1-carboxylate (MJN110) were tested alone and in combination at equieffective doses for reversal of CCI-induced mechanical allodynia and thermal hyperalgesia. The respective ED50 doses (95% confidence interval) of morphine and MJN110 were 2.4 (1.9-3.0) mg/kg and 0.43 (0.23-0.79) mg/kg. Isobolographic analysis of these drugs in combination revealed synergistic antiallodynic effects. Acute antinociceptive effects of the combination of morphine and MJN110 required µ-opioid, CB1, and CB2 receptors. This combination did not reduce gastric motility or produce subjective cannabimimetic effects in the drug discrimination assay. Importantly, combinations of MJN110 and morphine given repeatedly (i.e., twice a day for 6 days) continued to produce antiallodynic effects with no evidence of tolerance. Taken together, these findings suggest that MAGL inhibition produces opiate-sparing events with diminished tolerance, constipation, and cannabimimetic side effects.

Early adolescent nicotine exposure affects later-life cocaine reward in mice.

Adolescence represents a unique developmental period associated with increased risk-taking behavior and experimentation with drugs of abuse, in particular nicotine. We hypothesized that exposure to nicotine during early adolescence might increase the risk for drug reward in adulthood. To test this hypothesis, male ICR mice were treated with a subchronic regimen of nicotine or saline during adolescence, and their preference for cocaine, morphine and amphetamine was examined using the conditioned place preference (CPP) test in adulthood. Long-term behavioral changes induced by nicotine suggested a possible role of altered gene transcription. Thus, immunoblot for ΔFosB, a member of the Fos family of transcription factors, was conducted in the nucleus accumbens of these mice. Mice treated with nicotine during early but not late adolescence showed an increase in CPP for cocaine, morphine and amphetamine later in adulthood. This effect was not seen in mice pretreated with a subchronic regimen of nicotine as adults, suggesting that exposure to nicotine specifically during early adolescence increases the rewarding effects of other drugs in adulthood. However, adolescent nicotine exposure did not alter highly palatable food conditioning in mice. The enhancement of cocaine CPP by nicotine was strain-dependent and was blocked by pretreatment with nicotinic antagonists. In addition, nicotine exposure during early adolescence induced ΔFosB expression to a greater extent than identical nicotine exposure in adulthood, and enhanced cocaine-induced locomotor sensitization later in adulthood. These results suggest that nicotine exposure during early adolescence increases drug-induced reward in adulthood through mechanisms that may involve the induction of ΔFosB.

Full Fatty Acid Amide Hydrolase Inhibition Combined with Partial Monoacylglycerol Lipase Inhibition: Augmented and Sustained Antinociceptive Effects with Reduced Cannabimimetic Side Effects in Mice.

Inhibition of fatty acid amide hydrolase (FAAH) or monoacylglycerol lipase (MAGL), the primary hydrolytic enzymes for the respective endocannabinoids N-arachidonoylethanolamine (AEA) and 2-arachidonylglycerol (2-AG), produces antinociception but with minimal cannabimimetic side effects. Although selective inhibitors of either enzyme often show partial efficacy in various nociceptive models, their combined blockade elicits augmented antinociceptive effects, but side effects emerge. Moreover, complete and prolonged MAGL blockade leads to cannabinoid receptor type 1 (CB1) receptor functional tolerance, which represents another challenge in this potential therapeutic strategy. Therefore, the present study tested whether full FAAH inhibition combined with partial MAGL inhibition would produce sustained antinociceptive effects with minimal cannabimimetic side effects. Accordingly, we tested a high dose of the FAAH inhibitor PF-3845 (N-​3-​pyridinyl-​4-​[[3-​[[5-​(trifluoromethyl)-​2-​pyridinyl]oxy]phenyl]methyl]-​1-​piperidinecarboxamide; 10 mg/kg) given in combination with a low dose of the MAGL inhibitor JZL184 [4-nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate] (4 mg/kg) in mouse models of inflammatory and neuropathic pain. This combination of inhibitors elicited profound increases in brain AEA levels (>10-fold) but only 2- to 3-fold increases in brain 2-AG levels. This combination produced significantly greater antinociceptive effects than single enzyme inhibition and did not elicit common cannabimimetic effects (e.g., catalepsy, hypomotility, hypothermia, and substitution for Δ(9)-tetrahydrocannabinol in the drug-discrimination assay), although these side effects emerged with high-dose JZL184 (i.e., 100 mg/kg). Finally, repeated administration of this combination did not lead to tolerance to its antiallodynic actions in the carrageenan assay or CB1 receptor functional tolerance. Thus, full FAAH inhibition combined with partial MAGL inhibition reduces neuropathic and inflammatory pain states with minimal cannabimimetic effects.

Active, phosphorylated fingolimod inhibits histone deacetylases and facilitates fear extinction memory.

FTY720 (fingolimod), an FDA-approved drug for treatment of multiple sclerosis, has beneficial effects in the CNS that are not yet well understood, independent of its effects on immune cell trafficking. We show that FTY720 enters the nucleus, where it is phosphorylated by sphingosine kinase 2 (SphK2), and that nuclear FTY720-P binds and inhibits class I histone deacetylases (HDACs), enhancing specific histone acetylations. FTY720 is also phosphorylated in mice and accumulates in the brain, including the hippocampus, inhibits HDACs and enhances histone acetylation and gene expression programs associated with memory and learning, and rescues memory deficits independently of its immunosuppressive actions. Sphk2(-/-) mice have lower levels of hippocampal sphingosine-1-phosphate, an endogenous HDAC inhibitor, and reduced histone acetylation, and display deficits in spatial memory and impaired contextual fear extinction. Thus, sphingosine-1-phosphate and SphK2 play specific roles in memory functions and FTY720 may be a useful adjuvant therapy to facilitate extinction of aversive memories.

Repeated low-dose administration of the monoacylglycerol lipase inhibitor JZL184 retains cannabinoid receptor type 1-mediated antinociceptive and gastroprotective effects.

The monoacylglycerol lipase (MAGL) inhibitor 4-nitrophenyl 4-(dibenzo[d][1,3]dioxol-5-yl(hydroxy)methyl)piperidine-1-carboxylate (JZL184) produces antinociceptive and anti-inflammatory effects. However, repeated administration of high-dose JZL184 (40 mg/kg) causes dependence, antinociceptive tolerance, cross-tolerance to the pharmacological effects of cannabinoid receptor agonists, and cannabinoid receptor type 1 (CB1) downregulation and desensitization. This functional CB1 receptor tolerance poses a hurdle in the development of MAGL inhibitors for therapeutic use. Consequently, the present study tested whether repeated administration of low-dose JZL184 maintains its antinociceptive actions in the chronic constriction injury of the sciatic nerve neuropathic pain model and protective effects in a model of nonsteroidal anti-inflammatory drug-induced gastric hemorrhages. Mice given daily injections of high-dose JZL184 (≥16 mg/kg) for 6 days displayed decreased CB1 receptor density and function in the brain, as assessed in [(3)H]SR141716A binding and CP55,940 [(-)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl) cyclohexanol]-stimulated guanosine 5'-O-(3-[(35)S]thio)triphosphate binding assays, respectively. In contrast, normal CB1 receptor expression and function were maintained following repeated administration of low-dose JZL184 (≤8 mg/kg). Likewise, the antinociceptive and gastroprotective effects of high-dose JZL184 underwent tolerance following repeated administration, but these effects were maintained following repeated low-dose JZL184 treatment. Consistent with these observations, repeated high-dose JZL184, but not repeated low-dose JZL184, elicited cross-tolerance to the common pharmacological effects of Δ(9)-tetrahydrocannabinol. This same pattern of effects was found in a rimonabant [(5-(4-chlorophenyl)-1-(2,4-dichloro-phenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide)]-precipitated withdrawal model of cannabinoid dependence. Taken together, these results indicate that prolonged, partial MAGL inhibition maintains potentially beneficial antinociceptive and anti-inflammatory effects, without producing functional CB1 receptor tachyphylaxis/tolerance or cannabinoid dependence.

The monoacylglycerol lipase inhibitor JZL184 suppresses inflammatory pain in the mouse carrageenan model.

AIM: The present study tested whether the selective monoacylglycerol lipase (MAGL) inhibitor JZL184 would reduce allodynia and paw edema in the carrageenan test. MAIN METHODS: The anti-edematous and anti-allodynic effects of JZL184 were compared to those of PF-3845, an inhibitor of fatty acid amide hydrolase (FAAH), and diclofenac, a non-selective cyclooxygenase inhibitor. Cannabinoid receptor involvement in the anti-edematous and anti-allodynic effects of JZL184 was evaluated by administration of the respective CB1 and CB2 receptor antagonists rimonabant and SR144528 as well as with CB1(-/-) and CB2(-/-) mice. JZL184 (1.6, 4, 16, or 40mg/kg) was administered for six days to assess tolerance. KEY FINDINGS: JZL184 administered before or after carrageenan significantly attenuated carrageenan-induced paw edema and mechanical allodynia. Complementary genetic and pharmacological approaches revealed that the anti-allodynic effects of JZL184 required both CB1 and CB2 receptors, but only CB2 receptors mediated its anti-edematous actions. Importantly, both the anti-edematous and anti-allodynic effects underwent tolerance following repeated injections of high dose JZL184 (16 or 40mg/kg), but repeated administration of low dose JZL184 (4mg/kg) retained efficacy. SIGNIFICANCE: These results suggest that the MAGL inhibitor JZL184 reduces inflammatory nociception through the activation of both CB1 and CB2 receptors, with no evidence of tolerance following repeated administration of low doses.

Synaptic dysfunction in the hippocampus accompanies learning and memory deficits in human immunodeficiency virus type-1 Tat transgenic mice.

BACKGROUND: Human immunodeficiency virus (HIV) associated neurocognitive disorders (HAND), including memory dysfunction, continue to be a major clinical manifestation of HIV type-1 infection. Viral proteins released by infected glia are thought to be the principal triggers of inflammation and bystander neuronal injury and death, thereby driving key symptomatology of HAND. METHODS: We used a glial fibrillary acidic protein-driven, doxycycline-inducible HIV type-1 transactivator of transcription (Tat) transgenic mouse model and examined structure-function relationships in hippocampal pyramidal cornu ammonis 1 (CA1) neurons using morphologic, electrophysiological (long-term potentiation [LTP]), and behavioral (Morris water maze, fear-conditioning) approaches. RESULTS: Tat induction caused a variety of different inclusions in astrocytes characteristic of lysosomes, autophagic vacuoles, and lamellar bodies, which were typically present within distal cytoplasmic processes. In pyramidal CA1 neurons, Tat induction reduced the number of apical dendritic spines, while disrupting the distribution of synaptic proteins (synaptotagmin 2 and gephyrin) associated with inhibitory transmission but with minimal dendritic pathology and no evidence of pyramidal neuron death. Electrophysiological assessment of excitatory postsynaptic field potential at Schaffer collateral/commissural fiber-CA1 synapses showed near total suppression of LTP in mice expressing Tat. The loss in LTP coincided with disruptions in learning and memory. CONCLUSIONS: Tat expression in the brain results in profound functional changes in synaptic physiology and in behavior that are accompanied by only modest structural changes and minimal pathology. Tat likely contributes to HAND by causing molecular changes that disrupt synaptic organization, with inhibitory presynaptic terminals containing synaptotagmin 2 appearing especially vulnerable.

l-theanine attenuates abstinence signs in morphine-dependent rhesus monkeys and elicits anxiolytic-like activity in mice.

l-theanine, 2-amino-4-(ethylcarbamoyl) butyric acid, an amino acid found in green tea (Camellia sinensis), is sold in the United States as a dietary supplement to reduce stress and improve cognition and mood. The observations that l-theanine has been shown to inhibit caffeine's stimulatory effects and that caffeine produces precipitated withdrawal signs in opioid-addicted monkeys and some opioid withdrawal signs in some normal monkeys, suggest that l-theanine may suppress opioid withdrawal signs. Additionally, l-theanine produces anxiolytic effects in humans indicating that it has anti-anxiety properties. Thus, in these studies we determined whether l-theanine attenuates opioid-withdrawal signs in morphine-dependent rhesus monkeys, a model for spontaneous opioid withdrawal in human opioid addicts. We also evaluated whether l-theanine decreases anxiety-like behavior in mice, using the elevated plus maze and marble burying assays. l-theanine significantly attenuated designated opioid withdrawal signs, including fighting, rigid abdominal muscles, vocalizing on palpation of abdomen, pacing, retching, wet-dog shakes, and masturbation. It had a relatively quick onset of action that persisted for at least 2.5h. l-theanine also produced anxiolytic-like effects in the elevated plus maze and the marble burying assay in naïve mice at doses that did not significantly affect motor behavior. The results of these studies suggest that l-theanine may be useful in the pharmacotherapy of treating opioid withdrawal as well as anxiety-associated behaviors.

Dual fatty acid amide hydrolase and monoacylglycerol lipase blockade produces THC-like Morris water maze deficits in mice.

Acute administration of Δ(9)-tetrahydrocannabinol (THC) or exposure to marijuana smoke impairs short-term spatial memory in water maze tasks through a CB(1) receptor mechanism of action. N-Arachidonoylethanolamine (anandamide; AEA) and 2-arachidonoylglycerol (2-AG) are endogenous cannabinoids that are predominantly metabolized by the respective enzymes fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). Although the MAGL inhibitor JZL184 enhances short-term synaptic plasticity, it has yet to be evaluated in the Morris water maze. Previous research demonstrated that simultaneous, complete blockade of FAAH and MAGL produces full blown THC-like effects. Thus, in the following studies we tested whether dual blockade of FAAH and MAGL would impair learning in a repeated acquisition Morris water maze task. Mice treated with the dual FAAH/MAGL inhibitor JZL195 (20 mg/kg) as well as JZL184-treated FAAH -/- mice displayed robust deficits in Morris water maze performance that were similar in magnitude to THC-treated mice. While 20 or 40 mg/kg impaired water maze performance in FAAH -/- mice, only the high dose of JZL184 disrupted performance in FAAH +/+ mice. The memory impairing effects of JZL184 were blocked by the CB(1) receptor antagonist rimonabant. Neither JZL184 nor JZL195 impaired performance in a cued version of the water maze task, arguing against the notion that sensorimotor or motivational deficits accounted for the impaired acquisition performance. JZL184 increased 2-AG levels in the hippocampus, prefrontal cortex, and cerebellum to a similar degree in FAAH -/- and +/+ mice. FAAH -/- mice, regardless of drug treatment, possessed elevated AEA levels in each brain region assessed. The results of this study reveal that concomitant increases in AEA and 2-AG disrupt short-term spatial memory performance in a manner similar to that of THC.

Inhalation exposure to smoke from synthetic "marijuana" produces potent cannabimimetic effects in mice.

BACKGROUND: Use of synthetic "marijuana" has increased in recent years, produced adverse effects and prompted the temporary DEA ban of five specific cannabinoid analogs, including JWH-018. The objectives of the current study include determining the chemical content of the herbal product, Buzz, assessing its behavioral effects upon inhalation exposure to mice, determining whether CB(1) receptors mediate its pharmacological activity, and ascertaining its biodisposition in blood and various organs. METHODS: Using a nose-only exposure system, mice were exposed to smoke produced from combustion of an herbal incense product, Buzz, which contained 5.4% JWH-018. Cannabimimetic effects following smoke exposure were evaluated using the tetrad procedure, consisting of the following indices: hypomotility, antinociception, catalepsy, and hypothermia. Additionally, blood and tissues were collected for JWH-018 quantification. RESULTS: Inhalation exposure to Buzz produced dose-related tetrad effects similar to marijuana as well as dose-related increased levels of JWH-018 in the blood, brain, heart, kidney, liver, lung, and spleen. The behavioral effects were blocked by rimonabant, a CB(1) receptor antagonist. Effects produced by Buzz were similar in magnitude and time-course to those produced by marijuana, though equipotent doses of Buzz and marijuana yielded considerably lower brain levels of JWH-018 than THC for the respective materials. CONCLUSIONS: Inhalation exposure to a product containing JWH-018 penetrates into the brain and other organs and produces CB(1) receptor-mediated behavioral pharmacological effects in mice. The increased potency of JWH-018 compared to THC, the variable amount of drug added to various herbal products, and unknown toxicity, undoubtedly contribute to public health risks of synthetic cannabinoids.

Detection and disposition of JWH-018 and JWH-073 in mice after exposure to "Magic Gold" smoke.

The disposition in mice of the cannabimimetics JWH-018 and JWH-073 in blood and brain following inhalation of the smoke from the herbal incense product (HIP) "Magic Gold" containing 3.6% JWH-018, 5.7% JWH-073 and less than 0.1% JWH-398 (w/w) is presented. Specimens were analyzed by HPLC/MS/MS. The validation of the method is also presented. Five C57BL6 mice were sacrificed 20 min after exposure to the smoke of 200 mg of "Magic Gold" and a second set of five exposed mice were sacrificed after 20 h. Twenty minutes after exposure to "Magic Gold" smoke, blood concentrations of JWH-018 ranged from 42 to 160 ng/mL (mean: 88 ng/mL ± 42) and those of JWH-073 ranged from 67 to 244 ng/mL (mean: 134 ng/mL ± 62). Brain concentrations 20 min after exposure to "Magic Gold" smoke for JWH-018 ranged from 225 to 453 ng/g (mean: 317 ng/g ± 81) and those of JWH-073 ranged from 412 to 873 ng/g (mean: 584 ng/g ± 163). Twenty hours after exposure to "Magic Gold" smoke, JWH-018 was detected and quantified in only two of the five blood samples. Blood concentrations of JWH-018 were 3.4 ng/mL and 9.4 ng/mL. JWH-073 was detected in only one blood specimen 20 h after exposure at 4.3 ng/mL. Brain concentrations 20 h post exposure for JWH-018 ranged from 7 to 32 ng/g (mean: 19 ng/g ± 9). JWH-073 was not detected in 20 h post exposure brain specimens. JWH-398 was not detected in any of the blood or brain samples. The disposition data presented with the limited data available from human experience provide reasonable expectations for forensic toxicologists in JWH-018 or JWH-073 cases. As with THC after smoking marijuana, blood and brain concentrations of JWH-018 and JWH-073 after HIP smoking can be expected to rise initially to readily detected values, and then drop dramatically over the next few hours to several ng/mL or ng/g, and finally to be at extremely low or undetectable concentrations by 24h apparently due to extensive biotransformation, and redistribution to body fat.

Determination of naphthalen-1-yl-(1-pentylindol-3-yl)methanone (JWH-018) in mouse blood and tissue after inhalation exposure to 'buzz' smoke by HPLC/MS/MS.

The disposition of the cannabimimetic naphthalen-1-yl-(1-pentylindol-3-yl)methanone (JWH-018) in mice following inhalation of the smoke of the herbal incense product (HIP) 'Buzz' is presented. A high-pressure liquid chromatography with electrospray ionization triple quadrupole mass spectrometer (HPLC/MS/MS) method was validated for the analysis of JWH-018 in the specimens using deuterated Δ(9) -tetrahydrocannabinol (d(3) -THC) as the internal standard. JWH-018 was isolated by cold acetonitrile liquid-liquid extraction. Chromatographic separation was performed on a Zorbaz eclipse XDB-C(18) column. The assay was linear from 1 to 1000 ng/mL. Six C57BL6 mice were sacrificed 20 min after exposure to the smoke of 200 mg 'Buzz' containing 5.4% JWH-018. Specimen concentrations of JWH-018 were: blood, 54-166 ng/mL (mean 82 ± 42 ng/mL); brain, 316-708 ng/g (mean 510 ± 166 ng/g); and liver, 1370-3220 ng/mL (mean 1990 ± 752 ng/mL). The mean blood to brain ratio for JWH-018 was 6.8 and ranged from 4.2 to 10.9. After exposure, the responses of the mice were consistent with cannabinoid receptor type 1 activity: body temperatures dropped 7.3 ± 1.1 °C, and catalepsy, hyperreflexia, straub tail and ptosis were observed. The brain concentrations and physiological responses are consistent with the hypothesis that the behavioral effects of 'Buzz' are attributable to JWH-018.

delta(9)-Tetrahydrocannabinol-dependent mice undergoing withdrawal display impaired spatial memory.

RATIONALE: Cannabis users display a constellation of withdrawal symptoms upon drug discontinuation, including sleep disturbances, irritability, and possibly memory deficits. In cannabinoid-dependent rodents, the CB(1) antagonist rimonabant precipitates somatic withdrawal and enhances forskolin-stimulated adenylyl cyclase activity in cerebellum, an effect opposite that of acutely administered ∆(9)-tetrahydrocannabinol (THC), the primary constituent in cannabis. OBJECTIVES: Here, we tested whether THC-dependent mice undergoing rimonabant-precipitated withdrawal display short-term spatial memory deficits, as assessed in the Morris water maze. We also evaluated whether rimonabant would precipitate adenylyl cyclase superactivation in hippocampal and cerebellar tissue from THC-dependent mice. RESULTS: Rimonabant significantly impaired spatial memory of THC-dependent mice at lower doses than those necessary to precipitate somatic withdrawal behavior. In contrast, maze performance was near perfect in the cued task, suggesting sensorimotor function and motivational factors were unperturbed by the withdrawal state. Finally, rimonabant increased adenylyl cyclase activity in cerebellar, but not in hippocampal, membranes. CONCLUSIONS: The memory disruptive effects of THC undergo tolerance following repeated dosing, while the withdrawal state leads to a rebound deficit in memory. These results establish spatial memory impairment as a particularly sensitive component of cannabinoid withdrawal, an effect that may be mediated through compensatory changes in the cerebellum.

Fatty acid amide hydrolase (FAAH) knockout mice exhibit enhanced acquisition of an aversive, but not of an appetitive, Barnes maze task.

It is well established that genetic deletion or pharmacological inhibition of the CB(1) receptor disrupts extinction learning in aversive conditioning tasks, but not in appetitive tasks. Consistent with these findings is that genetic deletion or pharmacological inhibition of fatty acid amide hydrolase (FAAH), the primary catabolic enzyme of the endogenous cannabinoid anandamide (AEA), accelerates acquisition as well as extinction in aversive conditioning tasks. However, it is unknown whether FAAH blockade will affect acquisition in an appetitive conditioning task. Therefore, in the present study, we assessed FAAH (-/-) and (+/+) mice in appetitive and aversive Barnes maze conditioning procedures. Here we report that FAAH (-/-) mice displayed accelerated acquisition rates in an aversively-motivated, but not in the appetitively-motivated, Barnes maze task. The CB(1) receptor antagonist, rimonabant attenuated enhanced acquisition in the aversive procedure, consistent with the idea that elevated AEA levels mediate this apparent nootropic effect. These findings support the hypothesis that stimulation of the endocannabinoid system enhances learned behavior in aversive, but not appetitive, conditioning paradigms.

Route of administration influences substitution patterns in rats trained to discriminate methadone vs. vehicle.

Replacement therapy with the synthetic mu-opioid agonist methadone is an efficacious treatment for opioid abuse. While much is known about methadone's pharmacology, its discriminative stimulus properties remain largely unexplored. The present study sought to establish methadone discrimination in rats. Moreover, some research suggests that route of administration alters the discriminative stimulus of methadone. Thus, the present study also compared intraperitoneal (i.p.) and subcutaneous (s.c.) routes of administration. Male Sprague-Dawley rats were trained to discriminate 3.0mg/kg methadone (i.p.) from vehicle in a two-lever discrimination procedure. Generalization tests were conducted with a variety of compounds administered i.p. and s.c. Methadone fully substituted for itself, yielding ED(50)s of 1.5mg/kg (i.p.) and 0.2mg/kg (s.c.). Naltrexone (i.p.), an opioid antagonist produced a dose-dependent reduction in methadone-appropriate responding. The methadone stereoisomers fully substituted for methadone when given s.c.; however, when administered i.p., (+) and (-) methadone produced partial and no substitution, respectively. Heroin fully generalized to methadone regardless of administration route, while morphine fully substituted when given s.c., but not i.p. The kappa-agonist U50-488 failed to generalize to methadone with either route of administration. These results demonstrated that methadone's discriminative stimulus is mediated through mu-opioid receptor activity and is similar to that of commonly abused opioids (heroin, morphine). Additionally, route of administration produced differential results for many of the drugs tested, suggesting decreased drug bioavailability following i.p. administration due to hepatic first pass metabolism. Taken together, these results suggest that methadone's shared subjective effects with abused opioids, as well as its unique metabolic properties contribute to its efficacy in opioid maintenance therapy.

Hippocampal CB(1) receptors mediate the memory impairing effects of Delta(9)-tetrahydrocannabinol.

It is firmly established that the hippocampus, a brain region implicated in spatial learning, episodic memory, and consolidation, contains a high concentration of CB(1) receptors. Moreover, systemic and intrahippocampal administration of cannabinoid agonists have been shown to impair hippocampal-dependent memory tasks. However, the degree to which CB(1) receptors in the hippocampus play a specific functional role in the memory disruptive effects of marijuana or its primary psychoactive constituent Delta(9)-tetrahydrocannabinol (Delta(9)-THC) is unknown. This study was designed to determine whether hippocampal CB(1) receptors play a functional role in the memory disruptive effects of systemically administered cannabinoids, using the radial arm maze, a well characterized rodent model of working memory. Male Sprague-Dawley rats were implanted with bilateral cannulae aimed at the CA1 region of the dorsal hippocampus. The CB(1) receptor antagonist, rimonabant, was delivered into the hippocampus before to a systemic injection of either Delta(9)-THC or the potent cannabinoid analog, CP-55,940. Strikingly, intrahippocampal administration of rimonabant completely attenuated the memory disruptive effects of both cannabinoids in the radial arm maze task, but did not affect other pharmacological properties of cannabinoids, as assessed in the tetrad assay (that is, hypomotility, analgesia, catalepsy, and hypothermia). Infusions of rimonabant just dorsal or ventral to the hippocampus did not prevent Delta(9)-THC-induced memory impairment, indicating that its effects on mnemonic function were regionally selective. These findings provide compelling evidence in support of the view that hippocampal CB(1) receptors play a necessary role in the memory disruptive effects of marijuana.

Are CB(1) Receptor Antagonists Nootropic or Cognitive Impairing Agents?

For more than a decade, a considerable amount of research has examined the effects of rimonabant (SR 141716) and other CB(1) receptor antagonists in both in vivo and in vitro models of learning and memory. In addition to its utility in determining whether the effects of drugs are mediated though a CB(1) receptor mechanism of action, these antagonists are useful in providing insight into the physiological function of the endogenous cannabinoid system. Several groups have reported that CB(1) receptor antagonists enhance memory duration in a variety of spatial and operant paradigms, but not in all paradigms. Conversely, disruption of CB(1) receptor signaling also impairs extinction learning in which the animal actively suppresses a learned response when reinforcement has been withheld. These extinction deficits occur in aversively motivated tasks, such as in fear conditioning or escape behavior in the Morris water maze task, but not in appetitively motivated tasks. Similarly, in electrophysiological models, CB(1) receptor antagonists elicit a variety of effects, including enhancement of long-term potentiation (LTP), while disrupting long-term depression (LTD) and interfering with transient forms of plasticity, including depolarization-induced suppression of inhibition (DSI) and depolarization-induced suppression of excitation (DSE). The collective results of the in vivo and in vitro studies employing CB(1) receptor antagonists, demonstrate that these receptors play integral roles in different components of cognitive processing. Functionally, pharmacological blockade of CB(1) receptors may strengthen memory duration, but interferes with extinction of learned behaviors that are associated with traumatic or aversive memories.

The cannabinoid CB(1) receptor antagonist CE prolongs spatial memory duration in a rat delayed radial arm maze memory task.

The cannabinoid receptor system plays an integral role in learning and memory. Moreover, the cannabinoid CB(1) receptor antagonist rimonabant has been found to improve performance in a variety of animal memory models. The present study tested whether a novel and potent cannabinoid CB(1) receptor antagonist, CE, would prolong the duration of spatial memory. Rats were trained in a two-phase radial arm maze procedure, consisting of acquisition and retrieval tests, which were separated by an 18 h delay. CE was administered 30 min before the acquisition phase, immediately after the acquisition phase, or 30 min before the retrieval test to assess its effects on acquisition and retrieval processes. CE administered before and immediately after the acquisition phase significantly decreased the number of errors committed during the retrieval test. On the other hand, CE administered 30 min before the retrieval test had no effect on the number of errors committed. These findings demonstrate that CE improves memory by acting on consolidation, rather than retrieval, processes and further suggest that the endocannabinoid system has an important role in modulating memory duration.