Evidence for inverse agonism of SR141716A at human recombinant cannabinoid CB1 and CB2 receptors

The cannabinoid receptor antagonist SR141716A has been suggested to be an inverse agonist at CB₁ receptors in some isolated intact tissues. We found that the basal incorporation of [³⁵S]-GTPγS in Chinese hamster ovary cells expressing human recombinant CB₁ and CB₂ receptors was inhibited by SR141716A (mean pEC₅₀s 8.26 and 6.00, respectively), whereas cannabinol (10 μM) had no significant effect at hCB₁ receptors but inhibited the binding at hCB₂ receptors. As cannabinol had no effect on basal [³⁵S]-GTPγS binding at hCB₁ at a concentration 100 fold higher than its binding affinity (K_i=0.1 μM), we conclude that endogenous cannabinoid receptor agonists are not a confounding factor and suggest the actions of SR141716A at the hCB₁ receptor, and the actions of SR141716A and cannabinol at the hCB₂ receptor, are due to inverse agonism.     

The cannabinoid CB1 receptor antagonists rimonabant (SR141716) and AM251 directly potentiate GABA(A) receptors

BACKGROUND AND PURPOSE

Rimonabant (SR141716) and the structurally related AM251 are widely used in pharmacological experiments as selective cannabinoid receptor CB₁ antagonists / inverse agonists. Concentrations of 0.5–10 µM are usually applied in in vitro experiments. We intended to show that these drugs did not act at GABA_A receptors but found a significant positive allosteric modulation instead.

EXPERIMENTAL APPROACH

Recombinant GABA_A receptors were expressed in Xenopus oocytes. Receptors were exposed to AM251 or rimonabant in the absence and presence of GABA. Standard electrophysiological techniques were used to monitor the elicited ionic currents.

KEY RESULTS

AM251 dose-dependently potentiated responses to 0.5 µM GABA at the recombinant α₁β₂γ₂ GABA_A receptor with an EC₅₀ below 1 µM and a maximal potentiation of about eightfold. The Hill coefficient indicated that more than one binding site for AM251 was located in this receptor. Rimonabant had a lower affinity, but a fourfold higher efficacy. AM251 potentiated also currents mediated by α₁β₂, α_xβ₂γ₂ (x = 2,3,5,6), α₁β₃γ₂ and α₄β₂δ GABA_A receptors, but not those mediated by α₁β₁γ₂. Interestingly, the CB₁ receptor antagonists {"type":"entrez-nucleotide","attrs":{"text":"LY320135","term_id":"1257555575","term_text":"LY320135"}}LY320135 and O-2050 did not significantly affect α₁β₂γ₂ GABA_A receptor-mediated currents at concentrations of 1 µM.

CONCLUSIONS AND IMPLICATIONS

This study identified rimonabant and AM251 as positive allosteric modulators of GABA_A receptors. Thus, potential GABAergic effects of commonly used concentrations of these compounds should be considered in in vitro experiments, especially at extrasynaptic sites where GABA concentrations are low.

LINKED ARTICLES

This article is part of a themed section on Cannabinoids in Biology and Medicine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue-8. To view Part I of Cannabinoids in Biology and Medicine visit http://dx.doi.org/10.1111/bph.2011.163.issue-7     

SR 141716, a CB1 cannabinoid receptor antagonist, selectively reduces sweet food intake in marmoset

SR 141716 (1 and 3 mg/kg p.o.), a selective central (CB1) cannabinoid receptor antagonist, selectively reduced feeding of a very highly palatable cane-sugar mixture in marmosets. In contrast, standard primate pellet intake was not modified at the lower dose, but was slightly increased (+29%;p < 0.01) by the higher dose of SR 141716. These results are in agreement with the hypothesis that endogenous cannabinoid systems are involved in the modulation of the appetitive value of food.     

Rapid tolerance to the intestinal prokinetic effect of cannabinoid CB1 receptor antagonist, SR 141716 (Rimonabant)

The cannabinoid CB(1) receptor antagonist, SR 141716 (Rimonabant), has been reported to stimulate, when acutely administered, intestinal motility in mice. The present study was aimed at determining whether tolerance develops to its repeated administration. Mice were treated twice a day for up to 8 consecutive days with 0, 3 and 5.6 mg/kg SR 141716 (i.p.). On days 1, 3, 5 and 8, separate groups of mice were treated intragastrically with a non-absorbable colored marker (carmine). The distance traveled by the head of the marker in the small intestine was recorded. On day 1, SR 141716 markedly activated intestinal peristalsis, but complete tolerance to this effect developed within the third day of treatment. The results may have some relevance to the proposed future clinical use of SR 141716.     

Blockade by the cannabinoid CB(1) receptor antagonist,SR 141716, of alcohol deprivation effect in alcohol-preferring rats

The present study investigated the effect of the cannabinoid CB(1) receptor antagonist, SR 141716 (N-piperidino-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-3-pyrazole-carboxamide), on alcohol deprivation effect (i.e. the temporary increase in alcohol intake after a period of alcohol withdrawal) in Sardinian alcohol-preferring (sP) rats. As expected, alcohol-deprived rats virtually doubled voluntary alcohol intake during the first hour of re-access. Acute administration of SR 141716 (0, 0.3, 1 and 3 mg/kg, i.p.) completely abolished the alcohol deprivation effect. These results suggest that the cannabinoid CB(1) receptor is part of the neural substrate mediating the alcohol deprivation effect and that SR 141716 may possess anti-relapse properties.     

The cannabinoid CB(1) receptor antagonist SR141716A increases norepinephrine outflow in the rat anterior hypothalamus

The effects of the selective cannabinoid CB(1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide-hydrochloride (SR141716A) on extracellular concentrations of norepinephrine and 5-hydroxytryptamine (5-HT) were assessed by in vivo microdialysis in the anterior hypothalamus of freely moving rats. SR14716A (0.3, 1, 3 mg/kg, i.p.) dose-dependently increased norepinephrine efflux to about 300% of baseline, without affecting 5-HT levels. This increase in norepinephrine outflow could play an important role in the pharmacological and potentially therapeutic actions of SR141716A.     

Effects of the cannabinoid CB(1) receptor antagonist, SR141716A, after Delta(9)-tetrahydrocannabinol withdrawal

Rats were trained to lever press according to variable interval 10 s schedules during daily experimental sessions composed of six 3 min food reinforcement periods and were treated twice daily for 6 days with either vehicle or escalating regimens of Delta(9)-tetrahydrocannabinol. On days 7 and 8, the rats were challenged with vehicle and cumulative doses of SR141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4, -dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxyamide hydrochloride), a cannabinoid CB(1) receptor antagonist, up to 3 and 9 mg/kg, respectively. Response rates increased during Delta(9)-tetrahydrocannabinol withdrawal and towards those of the vehicle treatment group suggesting a waning of the direct effects of Delta(9)-tetrahydrocannabinol. SR141716A reduced response rates but only in rats pre-treated with Delta(9)-tetrahydrocannabinol. These data suggest that dependence upon Delta(9)-tetrahydrocannabinol was induced and SR141716A precipitated withdrawal.     

Cannabinoid receptor antagonism and inverse agonism in response to SR141716A on cAMP production in human and rat brain

The effects of cannabinoid drugs on cAMP production were examined in mammalian brain. The cannabinoid receptor agonist (R)-(+)-[2,3-dihydro-5-methyl-3-[(4-morpholinyl)methyl]pyrrolo[1,2,3,-d,e-1,4-benzoxazin-6-yl]-(1-naphthalenyl) methanone (WIN55,212-2) decreased forskolin-induced cAMP accumulation in a concentration-dependent manner (10(-8)-10(-5) M) in membranes from several rat and human brain regions, this effect being antagonized by 10(-5) M N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (SR141716A). Furthermore, high micromolar concentrations of SR141716A evoked a dose-dependent increase in basal cAMP in rat cerebellum and cortex, as well as in human frontal cortex. This effect was antagonized by WIN55,212-2 and abolished by N-ethylmaleimide, consistent with the involvement of cannabinoid CB(1) receptors through the activation of G(i/o) proteins. These results suggest a ligand-independent activity for cannabinoid CB(1) receptor signaling cascade in mammalian brain.     

The cannabinoid CB1 receptor antagonist SR141716A reduces appetitive and consummatory responses for food

Rationale The CB1 receptor antagonist SR141716A reduces food intake in rats. This effect is likely to depend on modulation of reward related processes.Objective To investigate the effects of SR141716A on responding for food under a second order instrumental task in which responding and consumption of food can be separated, and on Pavlovian responding for a stimulus predictive of food reward.Methods Instrumental responding and pellet consumption following administration of SR141716A (0–3 mg/kg) were recorded under an FI5 min FR5(5:S) operant schedule that incorporates both a 5 min initial appetitive phase and a 25 min consummatory phase. We compared the drug-induced change in responding to that recorded following a reduction in motivational state induced by pre-feeding. In a second experiment we assessed the effects of SR141716A (0–3 mg/kg) on Pavlovian approach behaviour for a stimulus (lever) associated with food reward (CS+) and a neutral stimulus (lever) not associated with reward (CS–).Results SR141716A reduced pellet consumption and instrumental responding during both the appetitive and consummatory phases of the second order schedule. Pre-feeding had a similar effect on responding during the appetitive phase, suggesting an effect on incentive motivation. SR141716A also blocked an enhancement of responding that occurred during the consummatory phase in pre-fed animals. SR141716A and pre-feeding had no effect on responding in the Pavlovian autoshaping paradigm.Conclusions SR141716A impacts on motivational processes in both the appetitive and consummatory phases of feeding behaviour.     

Suppressing effect of the cannabinoid CB1 receptor antagonist, SR 141716, on alcohol's motivational properties in alcohol-preferring rats

Administration of the cannabinoid CB(1) receptor antagonist, SR 141716 [N-piperidino-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-3-pyrazole-carboxamide], has been reported to reduce alcohol intake and alcohol self-administration in different models of excessive alcohol consumption, including the selectively bred Sardinian alcohol-preferring (sP) rats. The present study investigated whether SR 141716 was also capable of decreasing, in this rat line, alcohol's motivational properties. Extinction responding for alcohol, defined as the maximal number of lever responses reached in the absence of alcohol in rats trained to lever-press for alcohol, was used as index of alcohol's motivational properties. Rats were initially trained to lever-press for oral alcohol (15%, v/v) under a fixed ratio (FR) schedule of FR4. Once self-administration behavior was established, extinction sessions were conducted. SR 141716 (0, 0.3, 1 and 3 mg/kg; i.p.) was acutely administered before extinction sessions. In order to assess the specificity of SR 141716 action on extinction responding for alcohol, a separate group of sP rats was trained to lever-press for a 3% (w/v) sucrose solution under an FR4 schedule. SR 141716 administration produced a dose-dependent, virtually complete suppression of extinction responding for alcohol. In contrast, extinction responding for sucrose was not significantly altered by treatment with SR 141716. Further to the consummatory aspects, these results also extend the suppressing effect of SR 141716 to the appetitive aspects of alcohol drinking behavior in sP rats. The results also implicate the cannabinoid CB1 receptor in the neural substrate mediating alcohol's motivational properties in this rat line.     

The cannabinoid CB1 receptor antagonist SR141716 blocks the orexigenic effects of intrahypothalamic ghrelin

The paraventricular nucleus (PVN) of the hypothalamus plays a key role in the control of appetite and energy balance. Both ghrelin and cannabinoid receptor agonists increase food intake when administered into this nucleus: this study investigated possible interactions between the two systems in relation to eating. The orexigenic effect of ghrelin (100 pmol) when infused in to the PVN was reversed by a small, systemic dose of the CB(1) cannabinoid receptor antagonist SR141716 (1 mg kg(-1)). This is the first demonstration of a functional relationship between brain ghrelin and endocannabinoid systems, and, although it needs to be further investigated, the effect of ghrelin on food intake when injected into the PVN seems to be mediated by stimulation of cannabinoid release.     

Improvement of memory in rodents by the selective CB1 cannabinoid receptor antagonist,SR 141716

Social short-term memory in rodents is based on the recognition of a juvenile by an adult conspecific when the juvenile is presented on two successive occasions. Cannabimimetics are claimed to induce memory deficits in both humans and animals. In the brain, they mainly bind to CB1 receptors for which anandamide is a purported endogenous ligand. SR 141716, a specific antagonist of CB1 receptors, dose-dependently reverses biochemical and pharmacological effects of cannabimimetics. More particularly, it antagonizes the inhibition of hippocampal long-term potentiation induced by WIN 55,212-2 and anandamide, and it increases arousal when given alone. The present experiments study the ability of SR 141716 (from 0.03 to 3 mg/kg SC) to facilitate short-term olfactory memory in the social recognition test in rodents. SR 141716 improved social recognition in a long intertrial paradigm with a threshold dose of 0.1 mg/kg SC. At 1 mg/kg, it antagonized the memory disturbance elicited by retroactive inhibition. Scopolamine (0.06 mg/kg IP) partially reversed its memory-enhancing effect. Moreover, SR 141716 reduced memory deficit in aged rats (0.03–0.1 mg/kg) and mice (0.3–1 mg/kg). As SR 141716 is not known to exhibit any pharmacological activity which is not mediated by CB1 receptors, the results strongly support the concept that blockade of CB1 receptors plays an important role in consolidation of short-term memory in rodents and suggest there may be a role for an endogenous cannabinoid agonist tone (anandaminergic) in forgetting.     

Selective inhibition of sucrose and ethanol intake by SR 141716, an antagonist of central cannabinoid (CB1) receptors

SR 141716, a selective central CB1 cannabinoid receptor antagonist, markedly and selectively reduces sucrose feeding and drinking as well as neuropeptide Y-induced sucrose drinking in rats. SR 141716 also decreases ethanol consumption in C57BL/6 mice. In contrast, blockade of CB1 receptors only marginally affects regular chow intake or water drinking. The active doses of SR 141716 (0.3–3 mg/kg) are in the range known to antagonize the characteristic effects induced by cannabinoid receptor agonists. These results suggest for the first time that endogenous cannabinoid systems may modulate the appetitive value of sucrose and ethanol, perhaps by affecting the activity of brain reward systems. Received: 31 January 1997/Final version: 16 March 1997     

SR141716A, a cannabinoid CB1 receptor antagonist, improves memory in a delayed radial maze task

An endogenous cannabinoid system may play an important role in controlling memory processes. SR141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamidehydrochloride), a selective cannabinoid CB(1) receptor antagonist, was studied in an eight-arm radial maze task in which either deficits or improvements in memory could be detected. This task required well-trained rats to recall after either a relatively short (3 h) or long (7 h) delay period where they had received rewards during an information phase in order to obtain the remaining rewards during a retention phase. SR141716A was administered intraperitoneally immediately after the information phase in order to determine the drug's effects on memory consolidation. Although SR141716A had no effect on the number of errors committed after a short interval, SR141716A significantly reduced the number of errors that occurred after 7 h. These results suggest that a cannabinoid CB(1) receptor antagonist can improve consolidation processes and thus may be useful in treating memory disorders.     

Cannabinoid CB(1) antagonist SR 141716A attenuates reinstatement of heroin self-administration in heroin-abstinent rats

Rats with a previous history of heroin self-administration were studied to assess interactions occurring between cannabinoids and opioids in an animal model of reinstatement of heroin-seeking behaviour. Rats were trained to self-administer heroin and after a long-term extinction were primed with one of the following non-contingent non-reinforced drug administrations: saline (or vehicle), heroin, synthetic cannabinoid CB₁ receptor agonists (WIN 55,212-2 or CP 55,940), opioid antagonist (naloxone) or CB₁ antagonist (SR 141716A), alone or in combination. After primings, lever-pressing activity was recorded and compared to those observed during previous phases of training and extinction. Results of this study showed that (i) priming injections of heroin (0.1 mg/kg) as well as CB₁ agonists WIN 55,212-2 (0.15 or 0.30 mg/kg) and CP 55,940 (0.05 or 0.1 mg/kg) completely restore heroin-seeking behaviour; (ii) primings of naloxone (1 mg/kg) and SR 141716A (0.3 mg/kg) had no effect when administered alone; (iii) heroin-induced reinstatement was fully prevented by pre-treatment with either naloxone or SR 141716A; (iv) pre-treatment with SR 141716A significantly reduced WIN 55,212-2 and CP 55,940 priming effects. These results suggest that cannabinoid CB₁ receptors play an important role in the mechanisms underlying relapse to heroin-seeking and depict CB₁ antagonists as possible therapeutic agents for use in the prevention of relapse to heroin abuse.     

Blockade of cannabinoid (CB1) receptors by SR 141716 selectively antagonizes drug-induced reinstatement of exploratory behaviour in gerbils

  Rationale: A cannabinoid hypothesis of schi- zophrenia has been proposed according to which cognitive dysfunction could be associated with dysregulation of an endogenous cannabinoid system. Objective: The present study investigated whether SR 141716, a selective CB1 receptor antagonist, was able to reduce the hyperactivity induced in gerbils by various stimulant drugs known to produce or exacerbate schizophrenic symptoms. Methods: Cocaine, d-amphetamine, morphine, and Win 55212-2 were administered intraperitoneally (IP) either immediately before placing the animals in the test apparatus (non-habituated gerbils) or after a 2- to 3-h habituation period in the actimeter (habituated gerbils). SR 141716 was given IP 30 min before the injection of stimulant drugs. Horizontal activity was recorded every 10 min for 1 h in Digiscan activity monitor. Results: SR 141716 (0.3–3 mg/kg) dose-dependently suppressed the enhanced locomotor activity induced by each stimulant drug in habituated gerbils, but not in non-habituated animals. Clozapine, an atypical antipsychotic compound, but not haloperidol, shared with SR 141716, the ability to differentially affect drug-induced hyperactivity in habituated versus non-habituated gerbils. Conclusion: The activation of cannabinoid systems is a required, permissive element in the ability of cocaine, d-amphetamine, morphine, and Win 55212-2 to reinstate behaviour, i.e., to override stimulus satiation. Received: 1 August 1998 / Final version: 15 December 1998     

SR141716A诱导大鼠产生与CB1受体激动剂相反的行为模式(英文)

AIM: To examine the acute actions of the CB1cannabinoid receptor antagonist SR141716A [N-piperidino-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methylpyrazole-3-carbowamide] on typical behavioralpattern of psychoactive cannabinoids in rats.METHODS:At different time after injection the tail-flick response latency,the rectal temperature,thelocomotor activity,and the immobility on a ring as wellas the numbers of rears,self-grooming episodes(lasting 5 s),and fecal pellets were measured.RESULTS:Achte administration of SR141716A(3 mg/kg ip) induced a significant increase inhorizontal locomotor activity assayed by an activitymeter,in stereotypic activity(such as rearing and self-     

Effect of cannabinoid CB1 receptor antagonist SR141716A and CB1 receptor knockout on cue-induced reinstatement of Ensure and corn-oil seeking in mice.

The cannabinoid CB1 receptor antagonist SR141716A decreases cue-induced reinstatement of sucrose and drug seeking in rats. Reinstatement behavior is not well characterized in C57Bl/6 mice, including CB1 receptor knockout mice generated on a C57Bl/6 background. In the present study, male C57Bl/6, CB1 knockout (CB1 KO), and wild-type littermate (WT) mice were trained to respond for the sweet reinforcer Ensure or corn oil. Responding was maintained on a fixed ratio 1 (FR1) schedule of reinforcement for 10 days, and then extinguished by the removal of the reinforcer and associated cues. Subsequently, the effect of either pretreatment with SR141716A or CB1 receptor knockout on cue-induced reinstatement of Ensure or corn-oil seeking was assessed. Both 1.0 and 3.0 mg/kg SR141716A decreased reinstatement of Ensure seeking in C57Bl/6 mice. A tenfold higher dose of SR141716A (10.0 mg/kg) was required to attenuate reinstatement behavior in C57Bl/6 mice responding for corn oil, suggesting that CB1 receptors may be selectively involved in the neurobiology underlying reinstatement of responding for some food reinforcers but not others. Whereas CB1 receptor antagonism selectively attenuated reinstatement of responding for Ensure, genetic deletion of the CB1 receptor produced only a trend in decreasing reinstatement of Ensure seeking, and did not attenuate reinstatement of corn-oil seeking. Baseline differences in levels of operant responding were also observed in WT vs CB1 KO mice maintained by Ensure and corn oil. This and other possible reasons for the observed discrepancy between pharmacological blockade vs genetic invalidation of the CB1 receptor on reinstatement of Ensure seeking are discussed.     

The cannabinoid CB1 receptor antagonist SR141716A (Rimonabant) enhances the metabolic benefits of long-term treatment with oleoylethanolamide in Zucker rats

Anandamide and oleoylethanolamide (OEA) are lipid mediators that regulate feeding and lipid metabolism. While anandamide, a cannabinoid CB1 receptor agonist, promotes feeding and lipogenesis, oleoylethanolamide, an endogenous agonist of peroxisome proliferator activated receptor alpha (PPAR-alpha), decreases food intake and activates lipid mobilization and oxidation. The treatment with a cannabinoid CB1 receptor antagonist results in reduction of body weight gain and cholesterol in obese humans and rodents. In the present study, we show the benefits of the treatment of obese Zucker rats with a combination of a cannabinoid CB1 receptor antagonist (Rimonabant) and oleoylethanolamide. This combinational therapy improved the separate effects of Rimonabant and OEA, and resulted in marked decreases on feeding, body weight gain, and plasma cholesterol levels. Additionally, the treatment with both drugs reduced the hepatic steatosis observed in Zucker rats, decreasing liver fat deposits and damage, as revealed by the levels of alanine aminotransferase activity in serum. The combined treatment inhibits the expression of stearoyl coenzyme-A desaturase-1 (SCD-1), a pivotal enzyme in lipid biosynthesis and triglyceride mobilization that is linked to obesity phenotypes. These results support the use of combined therapies with cannabinoid CB1 receptor antagonists and PPAR-alpha agonists for the treatment of obesity associated with dyslipemia.     

SR 141716A, a CB1 cannabinoid receptor antagonist, potentiates the locomotor stimulant effects of amphetamine and apomorphine

The intraperitoneal (i.p.) injection of apomorphine or d-amphetamine significantly increased locomotor activity in Sprague-Dawley rats. Prior administration of the cannabinoid receptor antagonist, SR 141716A, significantly enhanced the stimulant effect of both d-amphetamine and apomorphine in a dose-dependent manner. Administration of SR 141716A alone had no effect on locomotor activity. These data indicate that endogenous cannabinoids exert an inhibitory action on the increase in locomotor activity produced by amphetamine and apomorphine.