Decreased alcohol self-administration and increased alcohol sensitivity and withdrawal in CB1 receptor knockout mice

Recent advances in the understanding of the neurobiological basis of alcohol dependence suggest that the endocannabinoid system may play a key role in the reinforcing effects of ethanol. In the present study, disruption of CB1 receptors in mice generated on a CD1 background decreased both ethanol consumption and preference. This decreased ethanol self-administration was associated with increased sensitivity to the acute intoxicating effects of ethanol. Mutant mice were more sensitive to the hypothermic and sedative/hypnotic effects of acute ethanol administration (1.5-4.0 g/kg), although plasma ethanol concentrations did not differ from those of controls. Moreover, wild-type mice exhibited normal locomotor activation caused by 1.0-2.5 g/kg injection of ethanol, whereas mutant mice displayed sedation in response to the injection of the same ethanol doses. The severity of alcohol withdrawal-induced convulsions was also increased in CB1(-/-) mice. Our results suggest that CB1 receptors participate in the regulation of ethanol drinking and demonstrate that their disruption lead to increased ethanol sensitivity and withdrawal severity.     

Expression of opioid receptors and c-fos in CB1 knockout mice exposed to neuropathic pain

The development of neuropathic pain is associated with multiple changes in gene expression occurring in the dorsal root ganglia (DRG) and spinal cord. The goal of this study was to evaluate whether the disruption of CB1 cannabinoid receptor gene modulates the changes induced by neuropathic pain in the expression of mu- (MOR), delta- (DOR) and kappa-opioid receptors (KOR) mRNA levels in the DRG and spinal cord. The induction of c-fos expression in the lumbar and sacral regions of the spinal cord was also evaluated in these animals. Opioid receptors mRNA levels were determined by using real-time PCR and Fos protein levels by immunohistochemistry. Nerve injury significantly reduced the expression of MOR in the DRG and the lumbar section of the spinal cord from CB1 cannabinoid knockout (KO) mice and wild-type littermates (WT). In contrast, mRNA levels of DOR and KOR were not significantly changed in any of the different sections analysed. Furthermore, sciatic nerve injury evoked a similar increase of c-fos expression in lumbar and sacral regions of the spinal cord of both KO and WT. In all instances, no significant differences were observed between WT and KO mice. These data revealed specific changes induced by neuropathic pain in MOR expression and c-fos levels in the DRG and/or spinal cord that were not modified by the genetic disruption of CB1 cannabinoid receptors.     

Lack of CB1 cannabinoid receptors modifies nicotine behavioural responses,but not nicotine abstinence

Cannabis is the most widely consumed illicit drug and its consumption is currently associated with tobacco, which contains another psychoactive compound, namely nicotine. Interactions between cannabinoids and other drugs of abuse, such as opioids, have been previously reported. The aim of the present study was to evaluate the possible role of CB1 cannabinoid receptor in responses induced by acute and repeated nicotine administration by using knockout mice lacking the CB1 cannabinoid receptor and their wild-type littermates. Acute nicotine (0.5, 1, 3 and 6 mg/kg, sc) administration decreased locomotor activity and induced antinociceptive responses in the tail-immersion and the hot-plate test, in wild-type animals. The antinociceptive effects in the tail-immersion test were significantly enhanced in CB1 knockout mice. In wild-type mice nicotine (0.5 mg/kg, sc) produced a significant rewarding effect, as measured by a conditioned place preference paradigm. This response was absent in CB1 knockout mice. Finally, a model of mecamylamine-induced abstinence in chronic nicotine-treated mice (10 mg/kg/day, sc) was developed. Mecamylamine (1 and 2 mg/kg, sc) precipitated several somatic signs of nicotine withdrawal in wild-type dependent mice. However, no difference in the severity of nicotine withdrawal was observed in CB1 knockout mice. These results demonstrate that some acute effects and motivational responses elicited by nicotine can be modulated by the endogenous cannabinoid system and support the existence of a physiological interaction between these two systems.     

Differential signalling in human cannabinoid CB1 receptors and their splice variants in autaptic hippocampal neurones

BACKGROUND AND PURPOSE

Cannabinoids such as Δ⁹- tetrahydrocannabinol, the major psychoactive component of marijuana and hashish, primarily act via cannabinoid CB₁ and CB₂ receptors to produce characteristic behavioural effects in humans. Due to the tractability of rodent models for electrophysiological and behavioural studies, most of the studies of cannabinoid receptor action have used rodent cannabinoid receptors. While CB₁ receptors are relatively well-conserved among mammals, human CB₁ (hCB₁) differs from rCB₁ and mCB₁ receptors at 13 residues, which may result in differential signalling. In addition, two hCB₁ splice variants (hCB_1a and hCB_1b) have been reported, diverging in their amino-termini relative to hCB₁ receptors. In this study, we have examined hCB₁ signalling in neurones.

EXPERIMENTAL APPROACH

hCB₁, hCB_1a hCB_1b or rCB₁ receptors were expressed in autaptic cultured hippocampal neurones from CB₁^−/− mice. Such cells express a complete endogenous cannabinoid signalling system. Electrophysiological techniques were used to assess CB₁ receptor-mediated signalling.

KEY RESULTS

Expressed in autaptic hippocampal neurones cultured from CB₁^−/− mice, hCB₁, hCB_1a and hCB_1b signal differentially from one another and from rodent CB₁ receptors. Specifically, hCB₁ receptors inhibit synaptic transmission less effectively than rCB₁ receptors.

CONCLUSIONS AND IMPLICATIONS

Our results suggest that cannabinoid receptor signalling in humans is quantitatively very different from that in rodents. As the problems of marijuana and hashish abuse occur in humans, our results highlight the importance of studying hCB₁ receptors. They also suggest further study of the distribution and function of hCB₁ receptor splice variants, given their differential signalling and potential impact on human health.

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     

Methamphetamine-seeking behavior is due to inhibition of nicotinic cholinergic transmission by activation of cannabinoid CB1 receptors

We previously reported the involvement of cannabinoid CB1 receptors (CB1Rs) and nicotinic acetylcholine receptors (nAChRs) in the reinstatement of methamphetamine (MAP)-seeking behavior (lever-pressing response for MAP reinforcement under saline infusion). The present study examined whether the reinstatement involves interactions between these receptors. Rats were trained to self-administer MAP with a light and tone (MAP-associated cues). Then, extinction sessions under saline infusion without cues were conducted. After that, a reinstatement tests were conducted by either presenting the cues or a MAP-priming injection. Systemic and intracranial administration of HU210, a cannabinoid CB1R agonist, into the nucleus accumbens core (NAC) and prelimbic cortex (PrC) reinstated MAP-seeking behavior. The reinstatement caused by the systemic HU210 treatment was attenuated by intracranial administration of AM251, a cannabinoid CB1R antagonist, into each region mentioned above. Meanwhile, reinstatement induced by the MAP-associated cues and MAP-priming injection was also attenuated by intracranial administration of AM251 in each region. In these regions, the attenuating effects of AM251 on the reinstatement induced by each stimulus were blocked by the intracranial administration of mecamylamine, a non-selective nAChR antagonist, but not by scopolamine, a muscarinic ACh receptor (mAChR) antagonist. Furthermore, the intracranial administration of DHβE, an α4β2 nAChR antagonist, but not MLA, an α7 nAChR antagonist, into each region blocked the AM251-induced attenuation of the reinstatement. These findings suggest that relapses to MAP-seeking behavior may be due to two steps, first inhibition of ACh transmission by the activation of cannabinoid CB1Rs and then the inactivation of α4β2 nAChRs.     

The CB1 inverse agonist AM251, but not the CB1 antagonist AM4113, enhances retention of contextual fear conditioning in rats

The effects of CB1 antagonist/inverse agonists on the acquisition and consolidation of conditioned fear remain uncertain. Recent studies suggest that the CB1 antagonist/inverse agonist AM251 affects acquisition or consolidation of both contextual and discretely cued fear memories. AM251 is frequently referred to as a CB1 antagonist; however in vitro signal transduction assays indicate that this drug also elicits inverse agonist activity at CB1 receptors. The present studies were undertaken to compare the effects of AM251 on conditioned fear with those produced by AM4113, a novel CB1 antagonist with minimal inverse agonist activity. All drugs were administered prior to conditioning. In retention tests conducted two weeks after conditioning, both AM251 (4.0 mg/kg) and AM4113 (6.0 mg/kg)-treated animals exhibited reduced freezing during a conditioned tone cue played within a novel context. In contextual fear retention tests, animals previously treated with 4.0 or 8.0 mg/kg AM251 exhibited enhanced freezing. By contrast, no dose of AM4113 had any significant effect on contextual fear memory, which is consistent with the lower signal transduction activity of AM4113 at CB1 receptors compared to AM251. These results suggest that CB1 neutral antagonists may be less likely than CB1 inverse agonists to facilitate the acquisition or consolidation of contextual fear that may contribute to some clinical disorders.     

Development and expression of neuropathic pain in CB1 knockout mice

Neuropathic pain is a clinical manifestation characterized by the presence of spontaneous pain, allodynia and hyperalgesia. Here, we have evaluated the involvement of CB1 cannabinoid receptors in the development and expression of neuropathic pain. For this purpose, partial ligation of the sciatic nerve was performed in CB1 cannabinoid receptor knockout mice and their wild-type littermates. The development of mechanical and thermal allodynia, and thermal hyperalgesia was evaluated by using the von Frey filaments, cold-plate and plantar tests, respectively. Pre-surgical tactile and thermal withdrawal thresholds were similar in both genotypes. In wild-type mice, sciatic nerve injury led to a neuropathic pain syndrome characterized by a marked and long-lasting reduction of the paw withdrawal thresholds to mechanical and thermal stimuli. These manifestations developed similarly in mice lacking CB1 cannabinoid receptors. We have also investigated the consequences of gabapentin administration in these animals. Gabapentin (50 mg/kg/day, i.p.) induced a similar suppression of mechanical and thermal allodynia in both wild-type and CB1 knockout mice. Mild differences between genotypes were observed concerning the effect of gabapentin in the expression of thermal hyperalgesia. Taken together, our results indicate that CB1 cannabinoid receptors are not critically implicated in the development of neuropathic pain nor in the anti-allodynic and anti-hyperalgesic effects of gabapentin in this model.     

Lean mean fat reducing “ghrelin” machine: Hypothalamic ghrelin and ghrelin receptors as therapeutic targets in obesity

Obesity has reached epidemic proportions not only in Western societies but also in the developing world. Current pharmacological treatments for obesity are either lacking in efficacy and/or are burdened with adverse side effects. Thus, novel strategies are required. A better understanding of the intricate molecular pathways controlling energy homeostasis may lead to novel therapeutic intervention. The circulating hormone, ghrelin represents a major target in the molecular signalling regulating food intake, appetite and energy expenditure and its circulating levels often display aberrant signalling in obesity. Ghrelin exerts its central orexigenic action mainly in the hypothalamus and in particular in the arcuate nucleus via activation of specific G-protein coupled receptors (GHS-R). In this review we describe current pharmacological models of how ghrelin regulates food intake and how manipulating ghrelin signalling may give novel insight into developing better and more selective anti-obesity drugs. Accumulating data suggests multiple ghrelin variants and additional receptors exist to play a role in energy metabolism and these may well play an important role in obesity. In addition, the recent findings of hypothalamic GHS-R crosstalk and heterodimerisation may add to the understanding of the complexity of bodyweight regulation.     

The role of halogen substitution in classical cannabinoids: A CB1 pharmacophore model

The presence of halogens within the classical cannabinoid structure leads to large variations in the compounds' potencies and affinities for the CB1 receptors. To explore the structure activity relationships within this class of analogs we have used a series of halogen-substituted (-)-Delta8-tetrahydrocannabinol analogs and compared their affinities for the CB1 cannabinoid receptor. Our results indicate that halogen substitution at the end-carbon of the side chain leads to an enhancement in affinity with the bulkier halogens (Br, I) producing the largest effects. Conversely, 2-iodo substitution on the phenolic ring leads to a 2-fold reduction in affinity while iodo-substitution in the C1'-position of the side chain lowers the compound's affinity for CB1 by more than 8-fold. The pharmacophoric requirements resulting from halogen-substitution are explored using computer modeling methods.     

Peripheral, but not central effects of cannabidiol derivatives: mediation by CB(1) and unidentified receptors

Delta-9 tetrahydrocannabinol (Δ⁹-THC) and (−)-cannabidiol ((−)-CBD) are major constituents of the Cannabis sativa plant with different pharmacological profiles: (Δ⁹-THC activates cannabinoid CB₁ and CB₂ receptors and induces psychoactive and peripheral effects. (−)-CBD possesses no, or very weak affinity for these receptors. We tested a series of (+)- and (−)-CBD derivatives for central and peripheral effects in mice. None of the (−)-CBD derivatives were centrally active, yet most inhibited intestinal motility. Of the five (+)-CBD derivatives, all with CB₁ receptor affinity, only (+)-7-OH-CBD-DMH (DMH=1,1-dimethylheptyl), acted centrally, while all five arrested defecation. The effects of (+)-CBD-DMH and (+)-7-OH-CBD-DMH were inhibited by the CB₁ receptor antagonist SR141716. The CB₂ receptor antagonist SR144528, and the vanilloid TRPV1 receptor antagonist capsazepine, had no influence. Further, the (−)-CBD derivatives (−)-7-COOH-CBD and (−)-7-COOH-CBD-DMH, displayed antiinflammatory activity.

We suggest that (+)-CBD analogues have mixed agonist/antagonist activity in the brain. Second, (−)-CBD analogues which are devoid of cannabinoid receptor affinity but which inhibit intestinal motility, suggest the existence of a non-CB₁, non-CB₂ receptor. Therefore, such analogues should be further developed as antidiarrheal and/or antiinflammatory drugs. We propose to study the therapeutic potential of (−)- and (+)-CBD derivatives for complex conditions such as inflammatory bowel disease and cystic fibrosis.     

Characterization of the endocannabinoid system, CB(1) receptor signalling and desensitization in human myometrium

BACKGROUND AND PURPOSE

The endocannabinoid plays vital roles in several aspects of reproduction, including gametogenesis, fertilization and parturition. However, little is known regarding the presence or role of the endocannabinoid system in myometrial function. Here the presence of the endocannabinoid system and signalling properties of cannabinoid receptors were characterized.

EXPERIMENTAL APPROACH

Components of the endocannabinoid system were identified using qRT-PCR, immunohistochemical, immunoblotting and radioligand binding experiments. Cannabinoid receptor signalling pathways were characterized using standard MAPK and second messenger assays.

KEY RESULTS

Primary myometrium expresses the endocannabinoid synthesizing enzyme N-acyl-phosphatidyl ethanolamine-specific phospholipase D, endocannabinoid degrading enzyme fatty acid amide hydrolase and cannabinoid CB₁, but not CB₂ receptors or transient receptor potential vanilloid-type-1 channels. The CB₁ receptor ligand anandamide caused a Gα_i/o-dependent inhibition of adenylate cyclase reducing intracellular cAMP levels, and Gα_i/o, phosphoinositide-3-kinase, Src-kinase-dependent ERK activation. CB₁ receptor-generated signals declined following continual anandamide stimulation, possibly due to ligand metabolism since free anandamide concentrations declined during the experiment from 2.5 µM initially, to 500 nM after >30 min. However, identical loss of CB₁ receptor responsiveness occurred in the presence of the metabolically stable derivative methanandamide. Moreover, RNAi-mediated depletion of arrestin3 (a negative regulator of receptor signalling) prevented loss of CB₁ receptor activity, enhancing and prolonging ERK signals.

CONCLUSIONS AND IMPLICATIONS

The myometrium has the capacity to synthesize, respond to and degrade endocannabinoids. Furthermore, reduced CB₁ receptor responsiveness occurs as a consequence of receptor desensitization, not agonist depletion and we identify a key role for arrestin3 in this process.     

Attenuation of basal and cocaine-enhanced locomotion and nucleus accumbens dopamine in cannabinoid CB1-receptor-knockout mice

Rationale  Effect of cannabinoid CB1 receptor deletion on cocaine’s actions is controversial. This is partly based on findings in CB1-receptor-knockout (CB1^−/−) mice with CD1 genetic background. Objectives  In the present study, we used CB1^−/− mice with a C57BL/6J genetic background to further investigate the role of CB1 receptors in cocaine’s action. Materials and methods  Locomotor activity was assessed using AccuScan locomotor chambers. Brain extracellular dopamine (DA) levels were measured by in vivo microdialysis and by fast-scan cyclic voltammetry in the nucleus accumbens (NAc). Results  CB1^−/− mice displayed a significant reduction in basal levels of locomotion and extracellular DA, as well as in cocaine-enhanced locomotion and extracellular DA, as compared to their wild-type (CB1^+/+) littermates. The reduction in basal and cocaine-enhanced DA appears to be related to a reduction in basal DA release, not to an increase in DA clearance, as indicated by fast-scan cyclic voltammetry in brain slices. Pharmacological blockade of CB1 receptors by SR141716 inhibited locomotion and NAc DA release in CB1^+/+ mice. Conclusions  The present findings suggest an important role for CB1 receptors in mediating cocaine’s behavioral and neurochemical effects.     

Age-dependent effects of the cannabinoid CB1 antagonist SR141716A on food intake, body weight change, and pruritus in rats

Rationale  The cannabinoid CB1 selective antagonist SR141716A (Rimonabant) has been shown to decrease body weight in laboratory animals and humans. Furthermore, SR141716A can elicit scratching behavior in rodents, a behavior that has been hypothesized to contribute to SR141716A-induced decrease in food intake. Although childhood obesity is a rising health issue, it is unknown whether SR141716A is equipotent at modulating food intake and other CB1-mediated behaviors in younger subjects. Objective  To determine whether CB1 receptor blockade is equipotent at modulating food and water intake, body weight, and scratching behavior, the effect of a range of SR141716A doses on these behaviors in food-restricted postnatal day (P) 18, 28, and 60 male rats was investigated. Brain concentrations of SR141716A were determined in each age group. Results  SR141716A dose- and age-dependently suppressed food and water intake and body weight gain and elicited head scratching, with the most potent effects observed in P18 and P28 rats. Brain concentrations of SR141716A were significantly elevated in P18 rats relative to P28 and P60 rats. SR141716A-elicited head scratching was attenuated by the 5-HT_2A/2C antagonist ketanserin. Conclusions  SR141716A is more potent at modulating food intake and head scratching in very young animals; these differences can be attributed to an increase in brain penetration of SR141716A for P18 but not for P28 and P60 rats. In addition, SR141716-elicited head scratching is modulated by 5HT receptor antagonism and is not a contributing factor to SR141716A's anorectic effects.     

CB1 receptors regulate alcohol-seeking behavior and alcohol self-administration of alcohol-preferring (P) rats

Rationale

The endogenous cannabinoid (CB) system mediates a number of behaviors associated with drug-seeking and drug self-administration. In this study the effects of CB1 receptor manipulations on operant ethanol (EtOH) responding during EtOH-seeking, EtOH-relapse as well as on-going EtOH self-administration were determined.

Methods

Alcohol-preferring (P) rats were trained in 2-lever operant chambers to self-administer 15% EtOH (v/v) and water on a concurrent fixed-ratio 5-fixed-ratio 1 (FR5-FR1) schedule of reinforcement in daily 1-h sessions. After 10 weeks, rats underwent 7 extinction sessions, followed by 2 weeks in their home cages without access to EtOH or operant chambers. Rats were then returned to the operant chambers for testing of EtOH-seeking behavior (no EtOH present) for 4 sessions. After a week in their home cages following the EtOH-seeking test, rats were returned to the operant chambers with access to EtOH and water (relapse). Rats were then maintained in the operant chambers for daily 1-h sessions with access to 15% EtOH and water for several weeks.

Results

The CB1 receptor antagonist (SR141716A), at doses of 1 and 2 mg/kg, i.p. reduced EtOH-seeking and transiently reduced EtOH self-administration during relapse and maintenance. Conversely, treatment with the CB1 receptor agonist CP 55, 940, at doses of 1 and 10 μg/kg i.p., increased EtOH-seeking and EtOH self-administration during relapse.

Conclusions

The results of this study demonstrate that activation of CB1 receptors are involved in regulating EtOH-seeking as well as the reinforcing effects of EtOH under relapse and on-going self-administration conditions.     

Additive effects of cannabinoid CB1 receptors blockade and cholecystokinin on feeding inhibition

Cannabinoid CB1 receptor and cholecystokinin-1 (CCK₁) receptors are located in peripheral nerve terminals of the gut, where they mediate satiety signals. Here we describe a detailed analysis of the interaction of both receptors in the control of feeding of food-deprived rats. Male Wistar rats were deprived for food 24 h before testing. Rats were pre-treated with SR141716A (Rimonabant) or WIN 55,212-2 before CCK-8 sulphated administration and tested for food intake 60, 120 and 240 min after last drug injection. In parallel, the effect of Lorglumide - a CCK₁ receptor antagonist - pre-treatment was evaluated on feeding behaviour after SR141716A administration. Results show that SR141716A activates c-Fos expression in brainstem areas receiving vagal inputs. Blockade of CB1 receptors with SR141716A (1 mg/kg) reduces feeding and display additive satiety induction with the CCK₁ receptor agonist CCK-8 sulphated (5, 10, 25 μg/kg). The effect of SR141716A is not blocked by Lorglumide (10 mg/kg), indicating independent sites of action. Conversely, the administration of the CB1 agonist WIN 55,212-2 (2 mg/kg) reduced satiety induced by CCK-8. In conclusion, these results report additive anorectic actions for CCK1 activation and peripheral CB1 receptor blockade providing a framework for combined therapies in the treatment of eating disorders.     

Adolescent peer-rejection persistently alters pain perception and CB1 receptor expression in female rats

Peer-interactions are particularly important during adolescence and teenagers display enhanced sensitivity toward rejection by peers. Social rejection has been shown to induce alterations in pain perception in humans. However, the neurobiological consequences of adolescent social rejection have yet to be extensively characterized, and no appropriate animal model is available. Here, we propose inadequate playful interactions in adolescent rats as a novel animal model for social peer-rejection and examine potential long-term consequences into adulthood. Acute social pairing of female adolescent Wistar rats with an age-matched rat from the less playful Fischer344 strain was found to alter social play and decrease pain reactivity, indicating Fischer rats as inadequate social partners for Wistar animals. Therefore, in a second experiment, adolescent female Wistar rats were either reared with another Wistar rat (adequate social rearing; control) or with a Fischer rat (inadequate social rearing; play-deprived). Beginning on day 50, all Wistar rats were group housed with same-strain partners and tested for behavioral, neurobiological and endocrine differences in adulthood. Playful peer-interactions were decreased during adolescence in play-deprived animals, without affecting social contact behavior. Consequently, adult play-deprived rats showed decreased pain sensitivity and increased startle reactivity compared to controls, but did not differ in activity, anxiety-related behavior or social interaction. Both groups also differed in their endocrine stress-response, and expression levels of the cannabinoid CB1 receptor were increased in the thalamus, whereas FAAH levels were decreased in the amygdala. The present animal model therefore represents a novel approach to assess the long-term consequences of peer-rejection during adolescence.     

CB1 receptor activation in the basolateral amygdala produces antinociception in animal models of acute and tonic nociception

1. Recent studies have suggested that the basolateral nucleus of the amygdala (BLA) participates in the processing of pain information, especially noxious somatic information. Cannabinoid receptors or CB1 mRNA are expressed more in the BLA than in other nuclei of the amygdala. Thus, the aim of the present study was to examine whether CB1 receptors in the BLA may be involved in modulating acute and/or tonic nociceptive processing. 2. Adult rats were exposed to intra-BLA microinjection of the cannabinoid receptor agonist (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl) pyrrolo [1,2,3,-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone mesylate [WIN 55,212-2 (1, 2.5, 5 or 10 microg/side)] and subjected to the tail flick and formalin tests. 3. The rats demonstrated a dose-dependent increase in latency to withdraw from a thermal noxious stimulus in the tail flick test and a decrease in formalin-induced pain behaviours. The antinociceptive effects of the CB1 receptor agonist WIN 55,212-2 (10 microg/side) in both tests were attenuated in the presence of the selective CB1 receptor antagonist, N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3- carboxamide (AM251; 0.55 ng/side). Administration of the CB1 receptor antagonist AM251 (0.55, 5.5, or 55.5 ng/side) alone did not alter the nociceptive thresholds in either test. Bilateral microinjection of the selective CB2 receptor antagonist N-[(1S)-endo-1,3,3-trimethyl bicyclo [2.2.1] heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528; 1 microg/side) had no effect on the antinociception produced by WIN 55,212-2, suggesting that the antinociceptive actions of WIN 55,212-2 are mediated by CB1 receptors. 4. The findings suggest the existence of a CB1-mediated inhibitory system in the BLA that, when activated, can diminish responsivity to acute and tonic noxious stimuli, but that normally has no tonic effect on the response threshold of these stimuli.     

Effects of cannabinoid CB1 receptor antagonist rimonabant in consolidation and reconsolidation of methamphetamine reward memory in mice

Rationale  Previous studies have shown that cannabinoid CB1 receptors play an important role in specific aspects of learning and memory, yet there has been no systematic study focusing on the involvement of cannabinoid CB1 receptors in methamphetamine-related reward memory. Objectives  The purpose of this study was to examine whether rimonabant, a cannabinoid CB1 receptor antagonist, would disrupt the consolidation and reconsolidation of methamphetamine-related reward memory, using conditioned place preference paradigm (CPP). Materials and methods  Separate groups of male Kunming mice were trained to acquire methamphetamine CPP. Vehicle or rimonabant (1 mg/kg or 3 mg/kg, i.p.) was given at different time points: immediately after each CPP training session (consolidation), 30 min before the reactivation of CPP (retrieval), or immediately after the reactivation of CPP (reconsolidation). Methamphetamine CPP was retested 24 h and 1 and 2 weeks after rimonabant administration. Results  Rimonabant at doses of 1 and 3 mg/kg significantly inhibited the consolidation of methamphetamine CPP. Only high-dose rimonabant (3 mg/kg) disrupted the retrieval and reconsolidation of methamphetamine CPP. Rimonabant had no effect on methamphetamine CPP in the absence of methamphetamine CPP reactivation. Conclusions  Our findings suggest that cannabinoid CB1 receptors play a major role in methamphetamine reward memory, and cannabinoid CB1 receptor antagonists may be a potential pharmacotherapy to manage relapse associated with drug-reward-related memory.     

CB1 receptors in the paraventricular nucleus of the hypothalamus modulate the release of 5-HT and GABA to stimulate food intake in rats

Endocannabinoids and their receptors not only contribute to the control of natural processes of appetite regulation and energy balance but also have an important role in the pathogenesis of obesity. CB1 receptors (CB1R) are expressed in several hypothalamic nuclei, including the paraventricular nucleus (PVN), where induce potent orexigenic responses. Activation of CB1R in the PVN induces hyperphagia by modulating directly or indirectly orexigenic and anorexigenic signals; however, interaction among these mediators has not been clearly defined. CB1R mRNA is expressed in serotonergic neurons that innervate the PVN, and activation of 5-HT receptors in the PVN constitutes an important satiety signal. Some GABAergic terminals are negatively influenced by 5-HT, suggesting that the hyperphagic effect of CB1R activation could involve changes in serotonergic and GABAergic signaling in the PVN. Accordingly, the present study was aimed to characterize the neurochemical mechanisms related to the hyperphagic effects induced by activation of CB1R in the PVN, studying in vitro and in vivo changes induced by direct activation these receptors. Here, we have found that the neurochemical mechanisms activated by stimulation of CB1 receptors in the PVN involve inhibition of 5-HT release, resulting in a decrease of serotonergic activity mediated by 5-HT_1A and 5-HT_1B receptors and inducing disinhibition of GABA release to stimulate food intake. In conclusion, these neurochemical changes in the PVN are determinant to the cannabinoid-induced stimulation of food intake. Our findings provide evidence of a functional connection among CB1R and serotonergic and GABAergic systems on the control of appetite regulation mediated by endocannabinoids.     

Stimulation of brain cannabinoid CB1 receptors can ameliorate hypertension in spontaneously hypertensive rats

Excessive activation of the sympatho-adrenomedullary system plays a pathogenic role in triggering and sustaining essential hypertension. We previously reported that, in normotensive rats, intracerebroventricularly (i.c.v.) administered neuropeptides, corticotropin-releasing factor and bombesin induced activation of the sympatho-adrenomedullary system, and that brain cannabinoid CB₁ receptors negatively regulated this activation. In this study, we investigated the effects of brain CB₁ receptor stimulation on blood pressure and the sympatho-adrenomedullary outflow in spontaneously hypertensive rats (SHRs), commonly used animal models of essential hypertension, and in Wistar-Kyoto (WKY) rats, normotensive controls of SHRs. In 18-week-old SHRs and WKY rats under urethane anaesthesia (1.0 g/kg, i.p.), SHRs exhibited significantly higher systolic, mean and diastolic blood pressures and plasma noradrenaline and adrenaline, and a lower heart rate than WKY rats. Single administration of arachidonyl 2′-chloroethylamide (ACEA, CB₁ agonist, 1.4 µmol/animal, i.c.v.) significantly but partially reduced mean and diastolic blood pressures and the plasma level of noradrenaline in SHRs compared to vehicle (N,N-dimethylformamide)-treated SHRs. These ACEA-induced reductions were abolished by central pretreatment with rimonabant (CB₁ antagonist, 300 nmol/animal, i.c.v.), which alone showed no significant effect on blood pressures or plasma noradrenaline and adrenaline levels of SHRs. On the other hand, ACEA had no significant effect on blood pressure or plasma noradrenaline and adrenaline levels in WKY rats. These results suggest that stimulation of brain CB₁ receptors can ameliorate hypertension accompanied by enhanced sympathetic outflow without affecting blood pressure under normotensive conditions.