Effects Of Cannabinoid Receptor Agonists Research Articles

Regulatory Effect of Cannabinoid Receptor Agonist on Chemokine-Induced Lymphocyte Chemotaxis

Cannabinoids elicit biological responses through two types of specific receptors, CB1 and CB2. Immune cells including naïve B-lymphocytes are known to selectively express peripheral cannabinoid receptors, CB2. Although the immunosuppressive effects of cannabinoids have become widely known, the mechanisms underlying their effects are not well understood. In this study, we demonstrated that splenic lymphocytes migrated toward a synthetic cannabinoid receptor agonist, WIN55,212-2. There is an optimal concentration range for induction of lymphocyte migration and a high dose fails to induce cell migration. Furthermore, a high dose of WIN55,212-2 significantly inhibited CXCL12-induced chemotaxis of lymphocytes. The inhibitory effect was transient and reversible. The inhibition was also observed when purified B-lymphocytes were used for CXCL12-induced chemotaxis. These results provide novel information regarding the cellular mechanisms underlying the effects of cannabinoids on the immune system.

Cannabinoids attenuate cancer pain and proliferation in a mouse model

We investigated the effects of cannabinoid receptor agonists on (1) oral cancer cell viability in vitro and (2) oral cancer pain and tumor growth in a mouse cancer model. We utilized immunohistochemistry and Western blot to show that human oral cancer cells express CBr1 and CBr2. When treated with WIN55,212-2 (non-selective), ACEA (CBr1-selective) or AM1241 (CBr2-selective) agonists in vitro, oral cancer cell proliferation was significantly attenuated in a dose-dependent manner. In vivo, systemic administration (0.013 M) of WIN55,212-2, ACEA, or AM1241 significantly attenuated cancer-induced mechanical allodynia. Tumor growth was also significantly attenuated with systemic AM1241 administration. Our findings suggest a direct role for cannabinoid mechanisms in oral cancer pain and proliferation. The systemic administration of cannabinoid receptor agonists may have important therapeutic implications wherein cannabinoid receptor agonists may reduce morbidity and mortality of oral cancer.

The effects of cannabinoid drugs on abnormal involuntary movements in dyskinetic and non-dyskinetic 6-hydroxydopamine lesioned rats

The long-term use of levodopa as a pharmacotherapy for Parkinson's disease is limited by the development of levodopa-induced dyskinesias. However, recent studies have suggested that pharmacological targeting of the endocannabinoid system may provide a viable adjunct to suppress these motor side effects. Thus, this study sought to determine the effect of pharmacologically activating or blocking endocannabinoid signalling on levodopa-induced dyskinesias in a rat model. Male Sprague–Dawley rats with 6-hydroxydopamine lesions were made dyskinetic by 6 weeks of daily levodopa injections (10 mg/kg s.c.). Rats that developed stable abnormal involuntary movements (AIMs) received acute injections of the cannabinoid receptor agonist, HU210 (0.0, 0.5, 5.0, and 50.0 μg/kg i.p.), or the CB 1 receptor antagonist/inverse agonist, AM251 (0.0 and 3.0 mg/kg i.p.), whereas rats that did not develop stable AIMs received injections of the CB 1 receptor antagonist/inverse agonist, rimonabant (0.0 and 3.0 mg/kg i.p.), for 18 days. In the dyskinetic rats, the highest dose of HU210 significantly reduced certain subtypes of AIMs but it also impaired normal motor functioning, while AM251 had no effect on AIMs. In the non-dyskinetic rats, rimonabant precipitated certain subtypes of AIMs. Overall, this study demonstrates that the anti-dyskinetic effects of cannabinoid receptor agonists may not be dissociable from their motor suppressant effects thereby limiting their potential usefulness for treating established dyskinesias in parkinsonism. However, it is intriguing that blockade of endocannabinoid–CB 1 signalling can unmask levodopa-induced AIMs, and this finding suggests that endocannabinoid tone may confer protection against the development of levodopa-induced dyskinesias.

Protective effects of cannabinoid receptor agonists against cocaine and other convulsant-induced toxic behavioural symptoms.

Based on the previously reported co-localization and relationship between cannabinoid and dopamine receptors, the effects of cannabinoid receptor agonists against cocaine-induced toxic behavioural symptoms, including convulsive seizures, were examined in mice. The anticonvulsant effect of several cannabimimetics against seizures induced by other convulsants was also compared. The cannabinoid receptor agonists CP 55940 ((-)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl)-cyclohexanol) and WIN 55212-2 ((R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone), and the endogenous cannabinoid anandamide were co-administered intraperitoneally with cocaine (75 mg kg(-1)) or other convulsants such as bicuculline, methyl 6,7-dimethoxy-4-ethyl-beta-carboline-carboxylate (DMCM), L-glutamic acid and N-methyl-D-aspartate (NMDA). CP 55940 (2.5 mg kg(-1)) and anandamide (15 mg kg(-1)) significantly antagonized cocaine-induced lethality, and CP 55940 and WIN 55212-2 (2.5 mg kg(-1)) significantly attenuated the severity of cocaine-induced convulsive seizures. Furthermore, ataxic hyperactivity, which was observed only in the cocaine-treated group of mice and could be evaluated by their activity counts, was also depressed in the groups of mice co-treated with each of the three cannabinoid agonists. However, none of these agonists protected against bicuculline- or DMCM-induced lethality or convulsive seizures. In contrast, all of the cannabinoid agonists, most notably anandamide, antagonized both L-glutamic acid (2 g kg(-1))- and NMDA (200 mg kg(-1))-induced convulsive seizures. These data support the previously reported close correlation between dopamine and cannabinoid receptors, and between cannabinoid agonists, especially anandamide, and glutamate (NMDA) receptors. Furthermore, these results suggest a potential therapeutic role for cannabinoid agonists against cocaine- and other-convulsant-induced toxicities.

Synaptic organization of retrograde 2-arachidonoylglycerol signaling in the dorsolateral periaqueductal grey matter

Event Abstract Back to Event Synaptic organization of retrograde 2-arachidonoylglycerol signaling in the dorsolateral periaqueductal grey matter Rita Nyilas1*, Laura C. Gregg2, Jessica M. Spradley2, Ken Mackie3, Andrea G. Hohmann2 and István Katona1 1 Hungarian Academy of Sciences, Institute of Experimental Medicine, Hungary 2 University of Georgia, United States 3 Indiana University, United States The striking antinociceptive effect of cannabinoid receptor agonists in acute nociception and chronic pain states is well-documented. 2-arachidonoylglycerol (2-AG), an endogenous ligand for both CB1 and CB2 cannabinoid receptors and a candidate retrograde synaptic endocannabinoid in the brain, has been implicated in the physiological regulation of nociception. However, the molecular machinery of synaptic 2-AG signaling involved has not been identified in the periaqueductal grey matter (PAG), a key midbrain area in the descending control of pain. To uncover the molecular architecture of 2-AG synthesis and identify sites of 2-AG action, we examined the detailed anatomical distribution of diacylglycerol lipase-α (DGL-α), metabotropic glutamate receptor 5 (mGluR5) and the CB1 cannabinoid receptor in the rodent dorsolateral (dl) PAG. Using peroxidase-based immunocytochemistry, dense distribution of DGL-α enzyme and CB1 receptor was revealed at the light microscopic level. Immunogold labeling followed by high-resolution electron microscopy showed that DGL-α, the main biosynthetic enzyme of 2-AG, was located in a remarkable postsynaptic position and co-localized with metabotropic glutamate receptor 5 (mGluR5), an upstream activator of 2-AG biosynthesis, while CB1 was found presynaptically on axon terminals. This striking positioning of DGL-α and CB1 were found at both excitatory and inhibitory synapses of the dl PAG. These findings imply a key role of the retrograde 2-AG pathway, present in dl PAG neuronal synapses, in the activity-dependent regulation of nociception. Conference: IBRO International Workshop 2010, Pécs, Hungary, 21 Jan - 23 Jan, 2010. Presentation Type: Poster Presentation Topic: Cellular neuroscience Citation: Nyilas R, Gregg LC, Spradley JM, Mackie K, Hohmann AG and Katona I (2010). Synaptic organization of retrograde 2-arachidonoylglycerol signaling in the dorsolateral periaqueductal grey matter. Front. Neurosci. Conference Abstract: IBRO International Workshop 2010. doi: 10.3389/conf.fnins.2010.10.00112 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 23 Apr 2010; Published Online: 23 Apr 2010. * Correspondence: Rita Nyilas, Hungarian Academy of Sciences, Institute of Experimental Medicine, Budapest, Hungary, nyilasr@koki.hu Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Rita Nyilas Laura C Gregg Jessica M Spradley Ken Mackie Andrea G Hohmann István Katona Google Rita Nyilas Laura C Gregg Jessica M Spradley Ken Mackie Andrea G Hohmann István Katona Google Scholar Rita Nyilas Laura C Gregg Jessica M Spradley Ken Mackie Andrea G Hohmann István Katona PubMed Rita Nyilas Laura C Gregg Jessica M Spradley Ken Mackie Andrea G Hohmann István Katona Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Peripheral and central sites of action for the non‐selective cannabinoid agonist WIN 55,212‐2 in a rat model of post‐operative pain

Activation of cannabinoid (CB) receptors decreases nociceptive transmission in inflammatory or neuropathic pain states. However, the effects of CB receptor agonists in post-operative pain remain to be investigated. Here, we characterized the anti-allodynic effects of WIN 55,212-2 (WIN) in a rat model of post-operative pain. WIN 55,212-2 was characterized in radioligand binding and in vitro functional assays at rat and human CB(1) and CB(2) receptors. Analgesic activity and site(s) of action of WIN were assessed in the skin incision-induced post-operative pain model in rats; receptor specificity was investigated using selective CB(1) and CB(2) receptor antagonists. WIN 55,212-2 exhibited non-selective affinity and agonist efficacy at human and rat CB(1) versus CB(2) receptors. Systemic administration of WIN decreased injury-induced mechanical allodynia and these effects were reversed by pretreatment with a CB(1) receptor antagonist, but not with a CB(2) receptor antagonist, given by systemic, intrathecal and supraspinal routes. In addition, peripheral administration of both CB(1) and CB(2) antagonists blocked systemic WIN-induced analgesic activity. Both CB(1) and CB(2) receptors were involved in the peripheral anti-allodynic effect of systemic WIN in a pre-clinical model of post-operative pain. In contrast, the centrally mediated anti-allodynic activity of systemic WIN is mostly due to the activation of CB(1) but not CB(2) receptors at both the spinal cord and brain levels. However, the increased potency of WIN following i.c.v. administration suggests that its main site of action is at CB(1) receptors in the brain.

CB1Receptors Mediate the Analgesic Effects of Cannabinoids on Colorectal Distension-Induced Visceral Pain in Rodents

Activation of cannabinoid receptors (CB(1), CB(2) and GPR(55)) produces analgesic effects in several experimental pain models, including visceral pain arising from the gastrointestinal tract. We assessed the role of CB(1), CB(2), and GPR(55) receptors and the endogenous cannabinoid system on basal pain responses and acute mechanical hyperalgesia during colorectal distension (CRD) in rodents. The effects of cannabinoid receptor agonists and antagonists on pain-related responses to CRD were assessed in rats and in wild-type and CB(1) receptor knock-out mice. The dual CB(1/2) agonist, WIN55,212-2, and the peripherally acting CB(1)-selective agonist, SAB-378, inhibited pain-related responses to repetitive noxious CRD (80 mmHg) in a dose-related manner in rats. The analgesic effects of WIN55,212-2 and SAB-378 were blocked by the selective CB(1) antagonist SR141716, but were not affected by the selective CB(2) antagonist SR144528. SR141716, per se, increased the responses to repetitive noxious CRD, indicative of hyperalgesia, and induced pain-related responses during non-noxious CRD (20 mmHg), indicative of allodynia. The cannabinoid receptor agonists anandamide, virodhamine and O-1602 had no effect. At analgesic doses, WIN55,212-2 did not affect colonic compliance. In accordance to the rat data, WIN55,212-2 produced analgesia, whereas SR141716 induced hyperalgesia, during noxious CRD (55 mmHg) in wild-type but not in CB(1)-knock-out mice. These data indicate that peripheral CB(1) receptors mediate the analgesic effects of cannabinoids on visceral pain from the gastrointestinal tract. The allodynic and hyperalgesic responses induced by SR141716 suggest the existence of an endogenous cannabinoid tone and the activation of CB(1) receptors during noxious CRD.

Analgesic and antiinflammatory effects of cannabinoid receptor agonists in a rat model of neuropathic pain

Cannabinoid receptor (CB) agonists are known to attenuate allodynia in a range of pain models, but their long-term effects and their mechanisms of action are controversial. The present study compares the antiallodynic effects of long-term treatment with a mixed CB1/CB2 (WIN55,212-2) and a selective CB2 (GW405833) cannabinoid receptor agonist and correlates these effects with their influences on spinal cord (SC) glial activation. The substances were applied daily in a rat neuropathic pain model. Tactile allodynia was assessed, and the development of gliosis was illustrated with immunohistochemical methods. Both substances reduced mechanical allodynia. Their analgesic effect was accompanied by a significant reduction in reactive gliosis and cathepsins (CAT) X and S expression. A daily injection of either substance for 8 days was sufficient to induce a sustained antiallodynic effect, which persisted up to 6 days after the last injection. The re-appearance of mechanical allodynia after this period was associated with a breakout of a strong gliotic response in the lumbar SC. Our results emphasize the therapeutic efficacy of cannabinoid receptor agonists and their inhibitory effects on the formation of gliosis.

Effect of Cannabinoids upon the Uptake of Folic Acid by BeWo Cells

Folic acid (FA) is an essential micronutrient that is particularly important during pregnancy for normal placental and fetal development and growth. The placenta and the fetus become frequently exposed to drugs of abuse such as cannabinoids because of maternal use of these substances. The aim of this study was to investigate the influence of cannabinoids on the uptake of FA by BeWo cells. Acute treatment with anandamide (1–10 μmol/l) caused a 15% decrease in ³H-FA uptake at pH 7.5. Moreover, tetrahydrocannabinol (THC) (1–10 μmol/l) caused a 30% increase, and AM630 (1 μmol/l) a 15% decrease in this parameter at pH 6.5. Neither the inhibitory effect of anandamide nor the stimulatory effect of THC were changed in the presence of cannabi- noid receptor type 1 or type 2 antagonists (AM251 and AM630, respectively). Chronic treatment (48 h) with THC (100 nmol/l) and AM251 (100 nmol/l) decreased the uptake of ³H-FA by 20% at pH 7.5, and anandamide (1 μmol/l) and AM630 (10–500 nmol/l) increased it by 30%. Moreover, CP55,940 (10 nmol/l) increased the uptake of ³H-FA by 30% at pH 6.5. RT-PCR analysis showed that the mRNA levels of the reduced folate transporter 1 increased by 9% after chronic treatment with AM630 (500 nmol/l). The mRNA levels of the proton-coupled folate transporter decreased by 17% and increased by 18% after chronic treatment with THC (0.1 μmol/l) and AM251 (100 nmol/l), respectively. In conclusion, ³H-FA uptake by BeWo cells is significantly, although not very markedly, changed by several distinct CB receptor agonists and antagonists, both after acute and chronic exposure of the cells. The acute effects of cannabinoid receptor agonists do not seem to be mediated by the cannabinoid receptor, and with a few exceptions the chronic effects do not seem to be related to changes in the expression levels of FA transporters.

Gz Mediates the Long-Lasting Desensitization of Brain CB1 Receptors and is Essential for Cross-Tolerance with Morphine

BackgroundAlthough the systemic administration of cannabinoids produces antinociception, their chronic use leads to analgesic tolerance as well as cross-tolerance to morphine. These effects are mediated by cannabinoids binding to peripheral, spinal and supraspinal CB1 and CB2 receptors, making it difficult to determine the relevance of each receptor type to these phenomena. However, in the brain, the CB1 receptors (CB1Rs) are expressed at high levels in neurons, whereas the expression of CB2Rs is marginal. Thus, CB1Rs mediate the effects of smoked cannabis and are also implicated in emotional behaviors. We have analyzed the production of supraspinal analgesia and the development of tolerance at CB1Rs by the direct injection of a series of cannabinoids into the brain. The influence of the activation of CB1Rs on supraspinal analgesia evoked by morphine was also evaluated.ResultsIntracerebroventricular (icv) administration of cannabinoid receptor agonists, WIN55,212-2, ACEA or methanandamide, generated a dose-dependent analgesia. Notably, a single administration of these compounds brought about profound analgesic tolerance that lasted for more than 14 days. This decrease in the effect of cannabinoid receptor agonists was not mediated by depletion of CB1Rs or the loss of regulated G proteins, but, nevertheless, it was accompanied by reduced morphine analgesia. On the other hand, acute morphine administration produced tolerance that lasted only 3 days and did not affect the CB1R. We found that both neural mu-opioid receptors (MORs) and CB1Rs interact with the HINT1-RGSZ module, thereby regulating pertussis toxin-insensitive Gz proteins. In mice with reduced levels of these Gz proteins, the CB1R agonists produced no such desensitization or morphine cross-tolerance. On the other hand, experimental enhancement of Gz signaling enabled an acute icv administration of morphine to produce a long-lasting tolerance at MORs that persisted for more than 2 weeks, and it also impaired the analgesic effects of cannabinoids.ConclusionIn the brain, cannabinoids can produce analgesic tolerance that is not associated with the loss of surface CB1Rs or their uncoupling from regulated transduction. Neural specific Gz proteins are essential mediators of the analgesic effects of supraspinal CB1R agonists and morphine. These Gz proteins are also responsible for the long-term analgesic tolerance produced by single doses of these agonists, as well as for the cross-tolerance between CB1Rs and MORs.

Effect of Cannabinoid Receptor Agonists on Streptozotocin-Induced Hyperalgesia in Diabetic Neuropathy

The effect of CB-1 and CB-2 receptor agonists, as well as an influence of a non-selective inhibitor of nitric oxide synthase (NOS), L-NOArg, and an inhibitor acting preferentially on cyclooxygenase-1 (COX-1), indomethacin, on the action of cannabinoid receptor agonists in a streptozotocin (STZ)-induced neuropathic model was investigated. When administered alone, a non-selective cannabinoid receptor agonist, WIN 55,212-2, a potentially selective CB-1 cannabinoid receptor agonist, Met-F-AEA, and a selective CB-2 cannabinoid receptor agonist, AM1241, dose-dependently reduced STZ-induced hyperalgesia. The results of the present study also demonstrated that inhibitors of COX and NOS increase antihyperalgesic activity of low doses of CB-1 and CB-2 receptor agonists. Hypothetical consequences of this phenomenon are discussed.

Interactions between Δ9-tetrahydrocannabinol and μ opioid receptor agonists in rhesus monkeys: discrimination and antinociception

Opioid receptor agonists can enhance some effects of cannabinoid receptor agonists, and cannabinoid receptor agonists can enhance some effects of opioid receptor agonists; however, the generality of these interactions is not established. This study examined interactions between the discriminative stimulus and antinociceptive effects of mu opioid receptor agonists and Delta(9)-tetrahydrocannabinol (THC) in rhesus monkeys. Neither heroin nor morphine (intravenous (i.v.) or subcutaneous (s.c.)) altered the discriminative stimulus effects of THC in monkeys (n = 5) discriminating 0.1 mg/kg THC i.v. In contrast, THC (s.c.) markedly attenuated the discriminative stimulus effect of morphine and heroin in nondependent monkeys (n = 4) discriminating 1.78 mg/kg morphine s.c. Doses of THC that attenuated the discriminative stimulus effects of morphine in nondependent monkeys failed to modify the discriminative stimulus effects of morphine in morphine-dependent (5.6 mg/kg/12 h) monkeys (n = 4) discriminating 0.0178 mg/kg naltrexone s.c. THC also failed to modify the discriminative stimulus effects of naltrexone in morphine-dependent monkeys or the effects of midazolam in monkeys (n = 4) discriminating 0.32 mg/kg midazolam s.c. Doses of THC (s.c.) that attenuated the discriminative stimulus effects of morphine in nondependent monkeys enhanced the antinociceptive effects of morphine (s.c.) in nondependent monkeys. While mu receptor agonists did not alter the discriminative stimulus effects of THC, THC altered the effects of mu receptor agonists in a context-dependent manner. That the same doses of THC enhance, attenuate, or do not affect morphine, depending on the condition, suggests that attenuation of morphine by THC can result from perceptual masking rather than common pharmacodynamic mechanisms or pharmacokinetic interactions.

The effect of cannabinoids on the uptake of 2‐deoxy‐D‐glucose by Caco‐2 cells

The endogenous cannabinoid system plays a role in the regulation of energy homeostasis acting through central pathways, and its dysregulation may be implicated in the pathogenesis of obesity. The endogenous cannabinoid system is also present in peripheral tissues. The aim of this work was to investigate the effect of cannabinoids upon the intestinal absorption of glucose, by investigating the effect of cannabinoid receptor agonists and antagonists upon the apical uptake of 3H‐2‐deoxy‐D‐glucose (3H‐DG) by human intestinal epithelial Caco‐2 cells. Uptake of 1 μM3H‐DG was both cytochalasin B‐ and phloridzin‐sensitive (±50% inhibition observed with each of these drugs), and their effect was not cumulative. On the other hand, uptake of 20 mM 3H‐DG was partially inhibited by cytochalasin B (±20%) and phloridzin had no effect. We verified that neither the cannabinoid receptor agonists THC (1–10 μM), anandamide (0.1–10 μM) and CP 55,940 (5 nM‐1 μM), nor the specific CB‐1 and CB‐2 antagonists (AM251 (10‐500 nM) and AM630 (50 nM‐1 μM), respectively) had a significant effect upon 3H‐DG uptake by Caco‐2 cells. This was true for both uptake of the low and high concentration of 3H‐DG. So, we hypothesize that cannabinoids do not interfere with the GLUT2‐mediated intestinal apical uptake of glucose. Supported by FCT (PTDC/SAU‐FCF/67805/2006).

Differential effects of cannabinoid receptor agonists on regional brain activity using pharmacological MRI

Activation of cannabinoid CB1 and/or CB2 receptors mediates analgesic effects across a broad spectrum of preclinical pain models. Selective activation of CB2 receptors may produce analgesia without the undesirable psychotropic side effects associated with modulation of CB1 receptors. To address selectivity in vivo, we describe non-invasive, non-ionizing, functional data that distinguish CB1 from CB2 receptor neural activity using pharmacological MRI (phMRI) in awake rats. Using a high field (7 T) MRI scanner, we examined and quantified the effects of non-selective CB1/CB2 (A-834735) and selective CB2 (AM1241) agonists on neural activity in awake rats. Pharmacological specificity was determined using selective CB1 (rimonabant) or CB2 (AM630) antagonists. Behavioural studies, plasma and brain exposures were used as benchmarks for activity in vivo. The non-selective CB1/CB2 agonist produced a dose-related, region-specific activation of brain structures that agrees well with published autoradiographic CB1 receptor density binding maps. Pretreatment with a CB1 antagonist but not with a CB2 antagonist, abolished these activation patterns, suggesting an effect mediated by CB1 receptors alone. In contrast, no significant changes in brain activity were found with relevant doses of the CB2 selective agonist. These results provide the first clear evidence for quantifying in vivo functional selectivity between CB1 and CB2 receptors using phMRI. Further, as the presence of CB2 receptors in the brain remains controversial, our data suggest that if CB2 receptors are expressed, they are not functional under normal physiological conditions.

The cannabinoid antagonist SR 141716A (Rimonabant) reduces the increase of extra-cellular dopamine release in the rat nucleus accumbens induced by a novel high palatable food

The assumption of a novel high palatable food (a candied cherry) occurs concomitantly with an increase in the concentration of extra-cellular dopamine and its main metabolite 3,4-dihydroxy-phenylacetic acid (DOPAC) by about 45% in the dialysate obtained by intracerebral microdialysis from the shell of the nucleus accumbens of male rats. Such increase was reversed by SR 141716A (Rimonabant), a selective cannabinoid CB1 receptor antagonist (0.3 mg/kg i.p. and 1 mg/kg i.p.), which also reduces the assumption of the high palatable food, when given 15 min before exposure to the candied cherry. SR 141716A effects on extracellular dopamine and DOPAC were prevented by WIN 55,212-2 (0.3 mg/kg i.p.) or HU 210 (0.1 mg/kg i.p.) given 15 min before SR 141716A. The present results show for the first time that SR 141716A reduces the increase in extra-cellular dopamine induced by a novel high palatable food in the nucleus accumbens. This confirms that cannabinoid CB1 receptors play a key role in food intake and/or appetite and suggests that the mesolimbic dopaminergic system is involved at least in part, in the effects of cannabinoid receptor agonists and antagonists on food intake and/or appetite.

Cannabinoid Receptor Agonists Inhibit Ca2+ Influx to Synaptosomes from Rat Brain

We examined the effects of cannabinoid receptor agonists on ⁴⁵Ca²⁺ uptake in rat brain synaptosomes. A cannabinoid receptor agonist, (R)-(+)-[2,3-dihydro-5-methyl-3-[(4-merpholino)methyl]pyrrolo-[1,2,3-de]-1,4-benzoxazin-6-yl](1-naphthyl)methanone (WIN 55212-2) dose-dependently inhibited ⁴⁵Ca²⁺ uptake in rat synaptosomes. Only an endogenous cannabinoid receptor agonist, anandamide, dose-dependently inhibited ⁴⁵Ca²⁺ uptake in rat synaptosomes, but not an endogenous cannabinoid receptor agonist, palmitoylethanolamide. Only a cannabinoid CB1 antagonist, [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamidehydrochloride] (SR 141716A), reversed the inhibitory effect of these WIN 55212-2 and anandamide on ⁴⁵Ca²⁺ uptake in rat synaptosomes, but not a cannabinoid CB2 receptor antagonist, [N-[(1S)-endo-1,3,3-trimethylbicyclo[2.2.1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)pyrazole-3-carboxamide] (SR 144528). The inhibitory effects of WIN 55212–2 and anandamide on ⁴⁵Ca²⁺ uptake in rat synaptosomes were reversed by the pretreatment of a voltage-sensitive A-type K⁺ channel blocker, dendrotoxin, but no other type of K⁺ channel blockers, i.e. iberiotoxin, charybdotoxin or glibenclamide. These findings suggest that cannabinoid receptors inhibit Ca²⁺ influx into rat brain nerves via the activation of CB1 receptors and the opening of voltage-sensitive A-type K⁺ channels.

Effects of Cannabinoid Receptor Agonists on Rat Gastric Acid Secretion: Discrepancy Between In Vitro and In Vivo Data

The effects of the cannabinoid (CB)-receptor agonists WIN55,212-2 and HU-210 and the selective CB(1)-receptor antagonist SR141716A were tested on in vitro and in vivo acid secretion assays from the rat. In the isolated gastric fundus from immature rats, WIN55,212-2 (0.001-30 microM), HU-210 (0.001-10 microM), or SR141716A (0.1-10 microM) did not change the basal acid output or acid responses to histamine, pentagastrin, or electrical field stimulation. HU-210 (0.3 micromol/kg, intravenously) inhibited the acid response to pentagastrin in anesthetized adult, young, or immature rats with lumen-perfused stomachs; moreover, HU-210 reduced vagally induced acid secretion in adult animals, its antisecretory effect being reversed by SR141716A (0.65 micromol/kg, intravenously). In vitro and in vivo data indicate that CB(1) receptors are not located on parietal cells but, rather, on vagal pathways (possibly at preganglionic sites) supplying the gastric mucosa. The lack of effect of CB-receptor ligands in vitro cannot be ascribed to the use of immature rats, since HU-210 inhibited stimulated acid secretion in vivo, irrespective of the animal age.

P.4.b.001 Anxiolytic and reward-related properties of URB597, a novel FAAH inhibitor, in CD1 mice

The present study further investigated the protective effects of cannabinoid receptor agonist WIN55,212-2 (WIN) and fatty acid amide hydrolase (FAAH) inhibitor URB597 (URB) on chronic cerebral hypoperfusion (CCH)-induced cognitive impairment in rats. Spatial learning and memory were assessed with the Morris water maze and by measuring Long-term potentiation. The expression of microtubule-associated protein-2 (MAP)-2, growth-associated protein-43 (GAP)-43, synaptophysin, cannabinoid receptor 1 (CB1), brain-derived neurotrophic factor (BDNF), FAAH, N-acylphosphatidylethanolamine phospholipase D(NAPE-PLD) and monoacyl glycerol lipase (MGL) as well as phosphoinositide 3-kinase (PI3K)/AKT signaling pathway molecules and downstream targets including AKT, phosphorylated (p-)AKT, cyclic AMP response element- binding protein (CREB), p-CREB, Bcl-2-associated death protein (BAD), p-BAD, glycogen synthase kinase (GSK)-3β, p-GSK-3β, forkhead box protein (FOXO) 3A and p-FOXO3A was determined by western blotting. WIN and URB treatment improved learning and memory performance, effects that were abolished by co-administration of the PI3K/AKT inhibitor LY294002. Moreover, WIN and URB reversed the decreases in MAP-2 and synaptophysin expression resulting from CCH, and stimulated BDNF and CB1 expression as well as CREB, FOXO3A, GSK-3β, and BAD phosphorylation, confirming that WIN and URB mediate neuroprotection by preventing neuronal apoptosis and improving cognition via PI3K/AKT signaling. These findings suggest that WIN and URB are promising agents for therapeutic management of CCH.

Δ 9-Tetrahydrocannabinol-induced desensitization of cannabinoid-mediated inhibition of synaptic transmission between hippocampal neurons in culture

Prolonged exposure to cannabinoids results in desensitization of cannabinoid receptors. Here, we compared the desensitization produced by the partial agonist, Δ 9-tetrahydrocannabinol (THC) to that produced by the full agonist Win55,212-2 on cannabinoid-mediated inhibition of glutamatergic synaptic transmission. Synaptic activity between rat hippocampal neurons was determined from network-driven increases in the intracellular Ca 2+ concentration ([Ca 2+] i spikes). To assess the effects of prolonged treatment, cultures were incubated with cannabinoids, washed in 0.5% fatty-acid-free bovine serum albumin to ensure the removal of the lipophilic drug and then tested for inhibition of [Ca 2+] i spiking by Win55,212-2. In control experiments, 0.1 μM Win55,212-2 inhibited [Ca 2+] i spiking by 93 ± 5%. Win55,212-2 produced significantly less inhibition of [Ca 2+] i spiking following 18–24 h treatment with 1 μM THC (48 ± 5%) or treatment with 1 μM Win55,212-2 (29 ± 6%). Thus, THC produced significantly less functional desensitization than Win55,212-2. The desensitization produced by THC was maximal at 0.3 μM, remained stable between 1 and 7 days of preincubation and shifted the EC 50 of acute inhibition by Win55,212-2 from 27 to 251 nM. Differences in the long-term effects of cannabinoid receptor agonists on synaptic transmission may prove important for evaluating their therapeutic and abuse potential.

Endogenous Cannabinoid Receptor Agonists Inhibit Neurogenic Inflammations in Guinea Pig Airways

Background: Although neurogenic inflammation via the activation of C fibers in the airway must have an important role in the pathogenesis of asthma, their regulatory mechanism remains uncertain. Objective: The pharmacological profiles of endogenous cannabinoid receptor agonists on the activation of C fibers in airway tissues were investigated and the mechanisms how cannabinoids regulate airway inflammatory reactions were clarified. Methods: The effects of endogenous cannabinoid receptor agonists on electrical field stimulation-induced bronchial smooth muscle contraction, capsaicin-induced bronchoconstriction and capsaicin-induced substance P release in guinea pig airway tissues were investigated. The influences of cannabinoid receptor antagonists and K⁺ channel blockers to the effects of cannabinoid receptor agonists on these respiratory reactions were examined. Results: Both endogenous cannabinoid receptor agonists, anandamide and palmitoylethanolamide, inhibited electrical field stimulation-induced guinea pig bronchial smooth muscle contraction, but not neurokinin A-induced contraction. A cannabinoid CB2 antagonist, SR 144528, reduced the inhibitory effect of endogenous agonists, but not a cannabinoid CB1 antagonist, SR 141716A. Inhibitory effects of agonists were also reduced by the pretreatment of large conductance Ca²⁺-activated K⁺ channel (maxi-K⁺ channel) blockers, iberiotoxin and charybdotoxin, but not by other K⁺ channel blockers, dendrotoxin or glibenclamide. Anandamide and palmitoylethanolamide blocked the capsaicin-induced release of substance P-like immunoreactivity from guinea pig airway tissues. Additionally, intravenous injection of palmitoylethanolamide dose-dependently inhibited capsaicin-induced guinea pig bronchoconstriction, but not neurokinin A-induced reaction. However, anandamide did not reduce capsaicin-induced guinea pig bronchoconstriction. Conclusions: These findings suggest that endogenous cannabinoid receptor agonists inhibit the activation of C fibers via cannabinoid CB2 receptors and maxi-K⁺ channels in guinea pig airways.