Antitussive effect of WIN 55212-2, a cannabinoid receptor agonist

Several lines of evidence indicate that the opioid and cannabinoid systems produce synergistic interactions. The present study examined the opioid receptors involved in the antitussive effect of WIN 55212-2 ((R)-(+)-[2,3-dihydro-5-methyl-3-[4-morpholinylmethyl]-pyrrolo-[1,2,3-de]-1,4-benzoxazin-6-yl](1-naphthyl) methanone mesylate), a high-affinity cannabinoid receptor agonist, in mice. WIN 55212-2, at doses of 0.3-3 mg/kg ip, produced a dose-dependent antitussive effect. This antitussive effect of WIN 55212-2 was antagonized by pretreatment with either methysergide (3 mg/kg ip), a 5-HT receptor antagonist, or naloxone (1 mg/kg ip), an opioid receptor antagonist. Furthermore, pretreatment with N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride (SR141716A, 3 mg/kg ip), a cannabinoid CB(1) receptor antagonist, also significantly reduced the antitussive effect of WIN 55212-2. Blockade of mu-opioid receptors by pretreatment with beta-funaltrexamine (40 mg/kg sc) significantly reduced the antitussive effect of WIN 55212-2. However, pretreatment with nor-binaltorphimine (20 mg/kg sc), a kappa-opioid receptor antagonist, did not affect the antitussive effect of WIN 55212-2. Pretreatment with naloxonazine (35 mg/kg sc), a mu(1)-opioid receptor antagonist, also did not affect the antitussive effect of WIN 55212-2. These results indicate that the antitussive effect of WIN 55212-2 is mediated by the activation of cannabinoid CB(1) receptors and mu(2) (naloxonazine-insensitive)-opioid receptors, but not mu(1) (naloxonazine-sensitive)- or kappa-opioid receptors.     

Effect of the cannabinoid receptor agonist WIN55212-2 on sympathetic cardiovascular regulation

1. The aim of the present study was to analyse the cardiovascular actions of the synthetic CB1/CB2 cannabinoid receptor agonist WIN55212-2, and specifically to determine its sites of action on sympathetic cardiovascular regulation. 2. Pithed rabbits in which the sympathetic outflow was continuously stimulated electrically or which received a pressor infusion of noradrenaline were used to study peripheral prejunctional and direct vascular effects, respectively. For studying effects on brain stem cardiovascular regulatory centres, drugs were administered into the cisterna cerebellomedullaris in conscious rabbits. Overall cardiovascular effects of the cannabinoid were studied in conscious rabbits with intravenous drug administration. 3. In pithed rabbits in which the sympathetic outflow was continuously electrically stimulated, intravenous injection of WIN55212-2 (5, 50 and 500 microg kg(-1)) markedly reduced blood pressure, the spillover of noradrenaline into plasma and the plasma noradrenaline concentration, and these effects were antagonized by the CB1 cannabinoid receptor-selective antagonist SR141716A. The hypotensive and the sympathoinhibitory effect of WIN55212-2 was shared by CP55940, another mixed CB1/CB2 cannabinoid receptor agonist, but not by WIN55212-3, the enantiomer of WIN55212-2, which lacks affinity for cannabinoid binding sites. WIN55212-2 had no effect on vascular tone established by infusion of noradrenaline in pithed rabbits. 4. Intracisternal application of WIN55212-2 (0.1, 1 and 10 microg kg(-1)) in conscious rabbits increased blood pressure and the plasma noradrenaline concentration and elicited bradycardia; this latter effect was antagonized by atropine. 5. In conscious animals, intravenous injection of WIN55212-2 (5 and 50 microg kg(-1)) caused bradycardia, slight hypotension, no change in the plasma noradrenaline concentration, and an increase in renal sympathetic nerve firing. The highest dose of WIN55212-2 (500 microg kg(-1)) elicited hypotension and tachycardia, and sympathetic nerve activity and the plasma noradrenaline concentration declined. 6. The results obtained in pithed rabbits indicate that activation of CB1 cannabinoid receptors leads to marked peripheral prejunctional inhibition of noradrenaline release from postganglionic sympathetic axons. Intracisternal application of WIN55212-2 uncovered two effects on brain stem cardiovascular centres: sympathoexcitation and activation of cardiac vagal fibres. The highest dose of systemically administered WIN55212-2 produced central sympathoinhibition; the primary site of this action is not known.     

The cannabinoid receptor agonist WIN 55212-2 inhibits neurogenic inflammations in airway tissues

We examined the effects of 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), on various respiratory reactions induced by the activation of capsaicin-sensitive afferent sensory nerves (C-fibers). WIN 55212-2 significantly inhibited capsaicin-induced guinea pig bronchoconstriction, but not the neurokinin A-induced reaction. Intravenous injection of WIN 55212-2 also blocked cigarette smoke-induced rat tracheal plasma extravasation. However, substance P-induced rat tracheal plasma extravasation was not affected by the administration of WIN 55212-2. A cannabinoid CB(2) 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) reduced the inhibitory effects of WIN 55212-2, but not a cannabinoid CB(1) antagonist, [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamidehydrochloride] (SR 141716A). A Maxi-K(+) channel opener, 1-(2'-hydroxy-5'-trifluoromethylphenyl)-5-trifluoromethyl-2(3H)benzimidazolone (NS 1619), specifically inhibited capsaicin-induced guinea pig bronchoconstriction and cigarette smoke-induced rat tracheal plasma extravasation. These findings suggest that WIN 55212-2 inhibits the activation of C-fibers via cannabinoid CB(2) receptors and Maxi-K(+) channels and reduces airway neurogenic inflammatory reactions in vivo.     

Spontaneous and precipitated withdrawal with a synthetic cannabinoid, WIN 55212-2

Physical dependence on the synthetic cannabinoid-receptor agonist R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl) methanone mesylate (WIN 55212-2) was demonstrated in rats by the use of a chronic continuous infusion. Spontaneous withdrawal, of moderate intensity, was shown for the first time with this class of drugs of abuse. Behavioral withdrawal signs were also elicited after challenge with (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide.HCl (SR141716A), a specific CB(1) cannabinoid-receptor antagonist. In both instances, the high-dose regimen (4, 8, 16 and 16 mg/kg/day, i.p. on days 1-4, respectively) was sufficient to evoke a typical withdrawal syndrome quantified by the signs wet-dog shakes and facial rubs. These results are discussed relative to those obtained with Delta(9)-tetrahydrocannabinol and anandamide. With Delta(9)-tetrahydrocannabinol, precipitated but not spontaneous or abrupt withdrawal was observed, and this was ascribed to pharmacokinetic properties. Anandamide, which showed little, if any, physical dependence potential, behaved atypically. Possible implications regarding pharmacotherapeutic and human abuse issues are discussed.     

Central effects of the cannabinoid receptor agonist WIN55212-2 on respiratory and cardiovascular regulation in anaesthetised rats

1 The primary aim was to study the central respiratory effects of cannabinoids (CB). To this end, the cannabinoid receptor agonist WIN55212-2 was injected into the cisterna magna of urethane-anaesthetised rats and changes in respiratory parameters were observed. The secondary aim was to observe the centrally elicited cardiovascular actions of WIN55212-2. Involvement of opioid mechanisms in the central effects of WIN55212-2 was also studied. 2 Intracisternal (i.c.) application of WIN55212-2 (1, 3, 10 and 30 microg kg(-1)) dose-dependently decreased the respiratory rate and minute volume. Tidal volume was slightly increased, whereas peak inspiratory flow remained unchanged. In addition, WIN55212-2 increased mean arterial pressure and the plasma noradrenaline concentration and decreased heart rate. 3 I.c. injection of WIN55212-3 (1, 3, 10 and 30 microg kg(-1)), an enantiomer of WIN55212-2 lacking affinity for cannabinoid receptors, elicited no effects. All effects of WIN55212-2 were prevented by the CB1 receptor antagonist SR141716 (2 mg kg(-1) i.v.). I.c. administration of the opioid receptor agonist DAMGO (0.1, 0.3, 1 and 3 microg kg(-1)) markedly lowered the respiratory rate, tidal volume, minute volume and peak inspiratory flow. These effects were attenuated by the opioid receptor antagonist naloxone (0.2 mg kg(-1) i.v.). In contrast, naloxone did not affect the respiratory and cardiovascular effects of i.c. administered WIN55212-2. 4 Our results show that activation of CB1 cannabinoid receptors in the brain stem depresses respiration and enhances sympathetic tone and cardiac vagal tone. Opioid mechanisms are not involved in these central cannabinoid effects.     

Regional haemodynamic responses to the cannabinoid agonist, WIN 55212-2, in conscious, normotensive rats, and in hypertensive, transgenic rats

Regional haemodynamic responses to the cannabinoid agonist, WIN 55212-2 (5 - 250 microg kg(-1) i.v.) were assessed in conscious, normotensive, Hannover, Sprague-Dawley (HSD) rats, and in hypertensive, transgenic ((mRen-2)27) (abbreviated to TG) rats. In HSD rats, WIN 55212-2 caused pressor, and renal and mesenteric vasoconstrictor effects, with a hindquarters vasodilator effect occurring only at the highest dose. In TG rats, the effects of the cannabinoid agonist were qualitatively similar to those seen in HSD rats, except there was no hindquarters vasodilatation. In both strains of rat, in the presence of losartan, pentolinium and a vasopressin (V1-receptor) antagonist, the pressor and vasoconstrictor effects of WIN 55212-2 were abolished, but the hindquarters vasodilator response was enhanced (HSD rats) or was seen only in that circumstance (TG rats). Under these conditions, both strains of rat showed a modest fall in blood pressure, together with mesenteric vasodilatation. In additional experiments in normotensive SD rats from Charles River (CRSD), it was shown that, in the presence of the V1-receptor antagonist alone, or losartan alone, or the two antagonists together, the cardiovascular effects of WIN 55212-2 (50 or 150 microg kg(-1)) were not attenuated. Hence, the effects described above were likely due to pentolinium. There were no consistent differences between HSD and TG rats in their haemodynamic responses to methoxamine or noradrenaline, indicating the two strains were not likely to differ markedly in their responsiveness to any putative sympathetic activation induced by WIN 55212-2. Collectively, the results indicate that the predominant cardiovascular effects of WIN 55212-2 in conscious HSD and TG rats (i.e., pressor and vasoconstrictor actions) can be attributed largely to indirect, pentolinium-sensitive mechanisms, which appear to differ little in the normotensive and hypertensive state, at least in conscious animals. Under the conditions of our experiments, signs of cannabinoid-induced vasodilatation were modest.     

An NMDA antagonist (LY 235959) attenuates abstinence-induced withdrawal of planarians following acute exposure to a cannabinoid agonist (WIN 55212-2)

The mechanisms that facilitate the development and expression of cannabinoid physical dependence in humans and other mammals are poorly understood. The present experiments used a planarian model to provide evidence that pharmacological antagonism of NMDA receptors significantly attenuates the development of cannabinoid physical dependence. Abstinence-induced withdrawal from the cannabinoid agonist WIN 55212-2 (10 microM) was manifested as a significant (P<0.05) decrease in the rate of planarian spontaneous locomotor velocity (pLMV) when WIN 55212-2 (10 microM)-exposed planarians were placed into drug-free water. No change in pLMV occurred when WIN 55212-2 (10 microM)-exposed planarians were placed into water containing WIN 55212-2 (10 microM). WIN 55212-2 (10 microM)-exposed planarians placed into water containing LY 235959 (1 or 10 microM) did not display withdrawal (no significant difference, P>0.05, in pLMV). In addition, withdrawal was not observed (no significant difference, P>0.05, in pLMV) in planarians that were co-exposed to a solution containing WIN 55212-2 (10 microM) and LY 235959 (10 microM). The present results reveal that NMDA receptor activation mediates the development of cannabinoid physical dependence and the expression of cannabinoid withdrawal in planarians.     

Antagonism between the anti-inflammatory activity of the cannabinoid WIN 55212-2 and SR 141716A

The CB1/CB2 receptor agonist WIN 55212-2 (0.75 mg/kg, i.v.) caused a significant reduction in neurogenic plasma extravasation induced by electrical stimulation of the saphenous nerve in anesthetized rats; WIN 55212-2 at 2.5-10 mg/kg, s.c., also produced a significant reduction in the carrageenan-induced paw edema in conscious rats. The selective CB1 antagonist SR 141716A (0.075-0.75 mg/kg i.v.) antagonized the WIN 55212-2 effects in the plasma extravasation model and antagonized the WIN 55212-2 (2.5 mg/kg, s.c.)-induced decreases in rectal temperature and increases in tail-flick latencies. However, SR 141716A (10 mg/kg, p.o.) failed to antagonize the effects of Win 55212-2 (2.5 mg/kg, s.c.) in the carrageenan model, suggesting that cannabinoid receptors found in the periphery may be able to modulate inflammatory processes in rats.     

Suppression by WIN55212-2, a cannabinoid receptor agonist, of inflammatory reactions in mouse ear: Interference with the actions of an endogenous ligand, 2-arachidonoylglycerol

The effect of WIN55212-2, a cannabinoid receptor agonist, on acute inflammation of mouse ear was investigated. We found that topical application of WIN55212-2 suppressed ear swelling induced by 12-O-tetradecanoylphorbol 13-acetate or 2-arachidonoylglycerol. Similar inhibition was observed with CP55940, another cannabinoid receptor agonist, and HU-308, a cannabinoid CB(2) receptor-selective agonist. WIN55212-2 also suppressed the infiltration of leukocytes induced by 12-O-tetradecanoylphorbol 13-acetate. On the other hand, WIN55212-3, an inactive enantiomer of WIN55212-2, exerted only small effects on inflammation. Notably, SR144528, a cannabinoid CB(2) receptor antagonist, also suppressed inflammatory reactions in mouse ear. Thus, both the cannabinoid CB(2) receptor agonist and antagonist are capable of reducing inflammatory reactions. We then investigated the mechanism underlying WIN55212-2-induced suppression of inflammation using cultured cells. We found that the addition of WIN55212-2 together with 2-arachidonoylglycerol blocked 2-arachidonoylglycerol-induced migration of human promyelocytic leukemia HL-60 cells that had been differentiated into macrophage-like cells. The restoration of 2-arachidonoylglycerol-desensitized cells and WIN55212-2-desensitized cells from an anergic condition was examined next. We found that 2-arachidonoylglycerol-treated cells rapidly recovered the capacity to respond to 2-arachidonoylglycerol. On the other hand, the anergic condition toward 2-arachidonoylglycerol continued for a longer period after pretreatment with WIN55212-2. These results suggest that the anti-inflammatory activity of WIN55212-2 is attributable, at least in part, to interference with the actions of the endogenous ligand, 2-arachidonoylglycerol.     

Identification of WIN55212-3 as a competitive neutral antagonist of the human cannabinoid CB2 receptor

1. Several G protein-coupled receptors (GPCRs), including cannabinoid CB(1) and CB(2) receptors, show constitutive activity under heterologous expression. Such a tonic response is generated in the absence of an activating ligand, and can be inhibited by inverse agonists. Neutral antagonists, however, are silent at such receptors, but can reverse both agonist and inverse agonist responses. To date, no neutral antagonist for the CB(2) receptor has been reported. 2. Here, by monitoring receptor-dependent G protein activation, we demonstrate that WIN55212-3 acts as a neutral antagonist at the human CB(2) (hCB(2)) receptor. WIN55212-3 alone, at concentrations 相似文献   

Modulation of paraoxon toxicity by the cannabinoid receptor agonist WIN 55,212-2

Cannabinoids can reduce the pre-synaptic release of acetylcholine and other neurotransmitters in the mammalian brain through a retrograde signaling pathway. Organophosphorus insecticides elicit toxicity by inhibiting acetylcholinesterase and thereby increasing synaptic acetylcholine levels. Several studies suggest that some organophosphorus toxicants can potentially modify cannabinergic signaling by direct binding to cannabinoid receptors and inhibition of enzymes responsible for cannabinoid degradation (i.e., fatty acid amide hydrolase and monoacylglycerol lipase). We hypothesized that exposure to the cannabinoid receptor agonist WIN 55,212-2 (WIN) could alter the acute toxicity of the prototype anticholinesterase, paraoxon. In vitro, paraoxon inhibited hippocampal cholinesterase and fatty acid amide hydrolase activities, and displaced specific binding to the cannabinoid receptor ligand ([(3)H]CP 55,940) in a concentration-dependent manner. WIN (0.5, 1.5 or 5mg/kg/day) had a complex dose-related effect on locomotor activity when evaluated for 2h after either the first or last of seven daily exposures, and significantly decreased hippocampal CB1 binding following repeated dosing. Four hours after dosing, paraoxon (0.4 mg/kg, sc) elicited classical signs of cholinergic toxicity and significantly reduced hippocampal cholinesterase and fatty acid amide hydrolase activities as well as [(3)H]CP 55,940 binding. A single exposure to WIN (1.5 mg/kg) significantly reduced involuntary movements and SLUD signs following acute paraoxon exposure (0.4 and 0.6 mg/kg, sc). In contrast, when rats were challenged with paraoxon (0.4 mg/kg) after the seventh daily exposure to WIN (1.5mg/kg/day), involuntary movements were significantly increased at later timepoints, while SLUD signs were unaffected. These results suggest that acute and repeated exposure to cannabinoid agonists may differentially modify acute cholinergic toxicity, possibly through modulation of acetylcholine release and adaptation in cannabinergic signaling associated with repeated cannabinoid exposures.     

The cannabinoid agonist WIN 55,212-2 reduces sensorimotor gating and recognition memory in rats

Cannabinoids can disrupt short-term memory in humans and animals and induce learning deficits and other cognitive impairments. In the present study we examined the role of a full cannabinoid agonist in short-term memory, sensorimotor gating, and the acquisition and expression of an operant learning paradigm in rats. We tested the effects of the synthetic cannabinoid WIN 55,212-2 (0.6 and 1.2 mg/kg) on short-term memory in social and object recognition tests, on prepulse inhibition (PPI) of startle, as well as on lever pressing for palatable food. Injections of 0.6 and 1.2 mg/kg WIN 55,212-2 impaired recognition memory and PPI in a dose-dependent manner, but had no effect on lever-pressing acquisition or expression, or on food preference. The PPI deficit was reversed by the administration of 0.1 mg/kg haloperidol. These data suggest that the synthetic cannabinoid WIN 55,212-2 does not lead to a general impairment of learning in an appetitive instrumental task, but significantly affects short-term memory and sensorimotor integration. The impairment in recognition and PPI might be due to deficits in attention-based short-term information processing.     

Activation of cannabinoid receptors in rat brain by WIN 55212-2 produces coupling to multiple G protein alpha-subunits with different potencies

Previous studies had shown that the amplification factors for cannabinoid receptors, defined as the number of total G proteins activated per occupied receptor, differs between several rat brain regions. In this study, we sought to determine which specific Gi/Go(alpha) subunits were activated by CB1 receptors in several rat brain regions and if this coupling might explain the regional differences in receptor/G protein amplification factors. Furthermore, we examined whether cannabinoid agonists might activate different subtypes of G(alpha) subunits with varying degrees of efficacy and/or potency. Activation of specific G proteins by cannabinoid receptors was evaluated by the ability of the agonist WIN 55212-2 to stimulate incorporation of [alpha-(32)P]azidoanilido-GTP into G(alpha) subunits in membranes. Photolabeled G proteins were either directly resolved using urea/SDS-polyacrylamide gel electrophoresis or first immunoprecipitated with specific antisera for different G(alpha) subunits before electrophoresis. Individual G(alpha) subunits were separated into distinct bands on a single gel and the amount of agonist-induced increase in radioactivity was quantified by densitometry. Stimulation of CB1 receptors by WIN 55212-2 resulted in the activation of a distinct pattern of at least five different G(ialpha)/G(oalpha) subunits in several brain regions. Furthermore, although the pattern of G proteins activated by WIN 55212-2 appeared to be similar across brain regions, slight differences were observed in both the percentage of increase and the amount of the individual G(alpha) subunits activated. Most importantly, the amount of WIN 55212-2 required to half-maximally activate individual G proteins in the cerebellum varied over a 30-fold range for different G(alpha) subunits. These results suggest that cannabinoid receptors activate multiple G proteins simultaneously in several brain regions and both the efficacy and potency of cannabinoid agonists to activate individual G(alpha) subunits may vary considerably.     

Intravenous self-administration of the cannabinoid CB1 receptor agonist WIN 55,212-2 in rats

RATIONALE: Delta9-tetrahydrocannabinol (Delta9-THC), the main psychoactive ingredient of marijuana, as well as synthetic cannabinoid (CB1) receptor agonists, has led to negative or equivocal results when tested with the intravenous self-administration procedure, the best validated behavioural model for evaluating abuse liability of drugs in experimental animals. We recently reported, however, that the synthetic CB1 receptor agonist WIN 55,212-2 is intravenously self-administered by drug-naive mice and that its self-administration is blocked by the cannabinoid CB1 receptor antagonist SR 141716A. OBJECTIVE: To assess a reliable model of cannabinoid intravenous self-administration in rats. Long Evans male rats were allowed the opportunity to self-administer WIN 55,212-2 at doses ranging from 6.25 to 50 microg/kg per injection, under a fixed-ratio 1 (FR1) schedule of reinforcement and nose-pokes as the operant responses. The effect of either a change in the unit drug dose available or a pretreatment with the specific CB1 receptor antagonist SR 141716A were then investigated (maintenance phase). Finally, the extinction of the self-administration behaviour was evaluated. RESULTS: Response rate depended on the drug dose available, with maximum rates occurring at 12.5 microg/kg per injection. Response rate increased following pretreatment with the specific CB1 receptor antagonist, SR 141716A. Moreover, operant behaviour rapidly extinguished following both the substitution of saline or vehicle for cannabinoid and the disconnection of the drug delivery pumps. CONCLUSION: Rats will intravenously self-administer the synthetic CB1 receptor agonist WIN 55,212-2 under specific experimental conditions, thus allowing further investigation of the neurobiological mechanisms underlying cannabinoid-taking behaviour.     

The cannabinoid agonist WIN 55,212-2 inhibits TNF-alpha-induced neutrophil transmigration across ECV304 cells

Cannabinoids are known to possess both anti-inflammatory and neuroprotective effects. In the present study, we have investigated the ability of cannabinoids to inhibit the transmigration of neutrophils in response to chemotaxic stimuli. The cannabinoid receptor agonist WIN 55,212-2 ((R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)-pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone mesylate) significantly decreased the number of migrating neutrophils across a monolayer of tumour necrosis factor alpha (TNF-alpha) activated ECV304 cells at concentrations >or=1 microM. In contrast, the agonists HU210 and CP 55,940 (0.01-1 microM) and the endocannabinoid anandamide (0.1-10 microM) were without significant effect on the response to TNF-alpha. The ability of WIN 55,212-2 to reduce the neutrophil transmigration was still seen in the presence of the cannabinoid CB(1) receptor antagonist/inverse agonist AM251 (0.1-1 microM) and the cannabinoid CB(2) receptor antagonist/inverse agonist AM630 (0.1-1 microM). TNF-alpha treatment of ECV304 cells caused release of interleukin-8 (IL-8), but WIN 55,212-2 did not affect either the ability of neutrophils to migrate across chemotaxis plates in response to an IL-8 stimulus, or to change the percentage of CXC 1 and CXC 2 receptors expressed by the neutrophils. WIN 55,212-2 at a concentration of 1 microM, but not at lower concentrations, produced a significant inhibition of IL-8 release from ECV304 cells in response to TNF-alpha-stimulation. Thus WIN 55,212-2 reduces the transmigration of neutrophils across a monolayer of TNF-alpha-activated ECV304 cells by an indirect action upon the release of IL-8 and/or other chemokine release from the ECV304 cells, and that this effect is brought about mainly by a cannabinoid CB receptor-independent mechanism.     

Synthesis and characterization of the potent cannabinoid agonist [naphthyl‐3H] WIN 55212‐2 at high specific activity

[Naphthyl‐³H] WIN 55212‐2 was prepared at high specific activity by the catalytic tritiation of dibromo precursor 3 . Product 4 was characterized by chromatography as well as tritium NMR and proven to be a useful radioligand. Copyright © 2002 John Wiley & Sons, Ltd.     

Adolescent Exposure to WIN 55212-2 Render the Nigrostriatal Dopaminergic Pathway Activated During Adulthood

BackgroundDuring adolescence, neuronal circuits exhibit plasticity in response to physiological changes and to adapt to environmental events. Nigrostriatal dopaminergic pathways are in constant flux during development. Evidence suggests a relationship between early use of cannabinoids and psychiatric disorders characterized by altered dopaminergic systems, such as schizophrenia and addiction. However, the impact of adolescent exposure to cannabinoids on nigrostriatal dopaminergic pathways in adulthood remains unclear. The aim of this research was to determine the effects of repeated activation of cannabinoid receptors during adolescence on dopaminergic activity of nigrostriatal pathways and the mechanisms underlying this impact during adulthood.MethodsMale Sprague-Dawley rats were treated with 1.2 mg/kg WIN 55212-2 daily from postnatal day 40 to 65. Then no-net flux microdialysis of dopamine in the dorsolateral striatum, electrophysiological recording of dopaminergic neuronal activity, and microdialysis measures of gamma-aminobutyric acid (GABA) and glutamate in substantia nigra par compacta were carried out during adulthood (postnatal days 72–78).ResultsRepeated activation of cannabinoid receptors during adolescence increased the release of dopamine in dorsolateral striatum accompanied by increased population activity of dopamine neurons and decreased extracellular GABA levels in substantia nigra par compacta in adulthood. Furthermore, perfusion of bicuculline, a GABAa antagonist, into the ventral pallidum reversed the increased dopamine neuron population activity in substantia nigra par compacta induced by adolescent cannabinoid exposure.ConclusionsThese results suggest that adolescent exposure to cannabinoid agonists produces disinhibition of nigrostriatal dopamine transmission during adulthood mediated by decreased GABAergic input from the ventral pallidum.     

Effects of the cannabinoid receptor agonist WIN 55,212-2 on operant behavior and locomotor activity in rats

Cannabinoids influence the motivational state of a subject and affect motor behavior. In the present study, we examined the acute effects of the cannabinoid (CB) receptor agonist WIN 55,212-2 (WIN) in three different doses (0.6, 1.2 and 1.8 mg/kg) on the performance of rats in a progressive ratio operant behavior task and on locomotor activity. WIN dose-dependently reduced the break point and the total number of lever-presses under a progressive ratio schedule. A food preference test revealed a preference for freely available casein pellets over lab chow in all treatment groups, indicating no WIN-effects on primary motivation. There was a significant reduction in the amount of casein pellets consumed by animals treated with 1.8 mg/kg WIN. Locomotor activity in the open field was increased by 0.6 mg/kg, but not by higher doses of WIN. These data show that administration of the synthetic cannabinoid receptor agonist WIN leads to dose-dependent alterations of the performance in an operant behavior task and of motor behavior. We confirm previous findings of dose-dependent motor stimulating and inhibiting effects of cannabinoids, and show an impairment of a complex operant behavior at higher doses of WIN.     

Functional responses to the cannabinoid agonist WIN 55,212-2 in neonatal rats of both genders: influence of weaning

We have studied behavioural, biochemical and endocrine responses to the cannabinoid agonist WIN 55,212-2 (WIN) in neonatal rats, as well as the effects of weaning on such responses. We used preweanling rats (20 days of age), 25-day-old weaned rats (weaning at Day 22) and 25-day-old nonweaned rats of both sexes. The behavioural effects of WIN were assessed in the nociceptive tail immersion test and in the open field. We also analysed the effect of weaning on corticosterone responses to WIN (radioimmunoassay) as well as on WIN-stimulated [35S] GTPgammaS binding in periaqueductal grey (PAG) and striatum. The cannabinoid agonist induced a modest increase in pain thresholds, whereas the effect of the drug on open-field activity, particularly on vertical activity, was much more marked. The weaning process appeared to reduce the baseline nociceptive latencies of the female rats. No significant effect of weaning on the behavioural responses to WIN was found. However, the group of weaned females (but not males) showed a significantly reduced WIN-stimulated [35S] GTPgammaS binding in the striatum. The cannabinoid agonist significantly increased the corticosterone levels of 25-day-old rats with the effect being more marked in weaned than in nonweaned animals. The results suggest that the weaning process might produce some sexually dimorphic developmental changes in CB1 receptor function.     

Permeation of WIN 55,212-2, a potent cannabinoid receptor agonist, across human tracheo-bronchial tissue in vitro and rat nasal epithelium in vivo

The aim of this study was to investigate the intranasal absorption of R-(+)-WIN 55,212-2 mesylate in vivo and in vitro. Permeation experiments of R-(+)-WIN 55,212-2 formulations with 2% dimethyl-beta-cyclodextrin (DMbetaCD), 2% trimethyl-beta-cyclodextrin (TMbetaCD) or 2% randomly methylated-beta-cyclodextrin (RAMbetaCD) in 1:1 propylene glycol/saline and 1.5% propylene glycol +3% Tween 80 in saline were conducted using EpiAirway tissue and an anesthetized rat nasal absorption model, respectively. Samples were analysed by liquid chromatography-mass spectrometry. Mucosal tolerance was screened using paracellular marker permeation and tissue viability as indices. Nasal absorption of WIN 55,212-2 was rapid, with a t(max) (time of peak concentration) of 0.17 to 0.35 h in vivo. Relative to 1.5% propylene glycol +3% Tween 80 (control), 1:1 propylene glycol/saline, RAMbetaCD, DMbetaCD and TMbetaCD resulted in 24-, 20-, 17- and 10-fold WIN 55,212-2 permeation increases in vitro, respectively. The in vivo absolute bioavailabilities were also increased with 1:1 propylene glycol/saline, RAMbetaCD, DMbetaCD and TMbetaCD compared to 1.5% propylene glycol +3% Tween 80 (0.15 vs. 0.66-0.77). The viability of the EpiAirway tissues was significantly reduced by DMbetaCD and TMbetaCD formulations. This study showed that WIN 55,212-2 mesylate can be delivered via the nasal route. Absorption of R-(+)-WIN 55,212-2 was rapid and bioavailability was significantly improved using methylated cyclodextrins and propylene glycol-based cosolvent.