Role of cannabinoid receptors in the regulation of cardiac contractility during ischemia/reperfusion

We studied the effect of selective ligands of cannabinoid (CB) receptors on contractility of isolated Langendorff-perfused rat heart under conditions of 45-min total ischemia and 30-min reperfusion. Perfusion with a solution containing selective CB receptor agonist HU-210 for 10 min before ischemia increased the severity of reperfusion contractile dysfunction. This drug decreased left ventricular developed pressure and maximum rates of contraction and relaxation, but had no effect on heart rate and end-diastolic pressure. The negative inotropic effect of the drug was transitory and disappeared after 5-min reperfusion. Pretreatment with selective CB1 receptor antagonist SR141716A and selective CB2 receptor antagonist SR144528 had no effect on heart rate and myocardial contractility during reperfusion. Our results indicate that stimulation of CB receptors can increase the degree of reperfusion-induced cardiac contractile dysfunction. However, endogenous cannabinoids are not involved in the development of myocardial contractile dysfunction during ischemia/reperfusion of the isolated heart. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 142, No. 11, pp. 500–504, November, 2006     

Negative chronotropic effect of cannabinoids and their water-soluble emulsion is related to activation of cardiac CB1 receptors

Intravenous injection of cannabinoids dissolved in cremophore EL:ethanol:NaCl mixture and water-soluble emulsion of the same cannabinoids caused identical negative chronotropic effects in chloralose-narcotized rats. Selective CB1 and CB2 receptor antagonist HU-210 also induced a negative chronotropic effect in rats, while pre-injection of CB1 receptor antagonist SR 141716A completely abolished this effect of HU-210. Selective CB2 receptor antagonist SR 144528 had no effect on HU-210-induced bradycardia. Preinjection of ganglioblocker hexamethonium also did not abolish the negative chronotropic effect of HU-210 and ACPA. Perfusion of isolated rat heart with Krebs-Henseleit solution containing HU-210 in a final concentration of 100 nM reduced heart rate. It was shown that the negative chronotropic effect of cannabinoids is mediated through activation of cardiac CB1 receptors. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 142, No. 10, pp. 433–436, October, 2006     

Activation of type II cannabinoid receptors improves myocardial tolerance to arrhythmogenic effects of coronary occlusion and reperfusion

Preliminary intravenous injection of cannabinoid receptor agonist HU-210 (0.05 mg/kg) reduced the incidence of ventricular arrhythmias during 10-min coronary occlusion and 10-min reperfusion in chloralose-anesthetized rats. Preliminary injection of type I cannabinoid receptor antagonist SR 141716A (3 mg/kg) had no effect on the antiarrhythmic effect of HU-210, while type II cannabinoid receptor antagonist SR 144528 (1 mg/kg) completely abolished the effect of HU-210. Preconditioning with glibenclamide (0.3 mg/kg), an inhibitor of ATP-dependent K⁺-channels, did not affect the antiarrhythmic activity of HU-210. These findings suggest that antiarrhythmic effect of HU-210 is mediated through activation of type II cannabinoid receptors rather than activation of K⁺-channels.     

A beneficial aspect of a CB1 cannabinoid receptor antagonist: SR141716A is a potent inhibitor of macrophage infection by the intracellular pathogen Brucella suis

The psychoactive component of marijuana, delta9-tetrahydrocannabinol (THC) suppresses different functions of immunocytes, including the antimicrobicidal activity of macrophages. The triggering of cannabinoid receptors of CB1 and CB2 subtypes present on leukocytes may account for these effects. We investigated the influence of specific CB1 or CB2 receptor antagonists (SR141716A and SR144528, respectively) and nonselective CB1/CB2 cannabinoid receptor agonists (CP55,940 or WIN 55212-2) on macrophage infection by Brucella suis, an intracellular gram-negative bacteria. None of the compounds tested affected bacterial phagocytosis. By contrast, the intracellular multiplication of Brucella was dose-dependently inhibited in cells treated with 10-500 nM SR141716A and 1 microM SR141716A-induced cells exerted a potent microbicidal effect against the bacteria. SR144528, CP55,940, or WIN 55212-2 did not affect (or slightly potentiated) the growth of phagocytized bacteria. However, CP55,940 or WIN 55212-2 reversed the SR141716A-mediated effect, which strongly suggested an involvement of macrophage CB1 receptors in the phenomenon. SR141716A was able to pre-activate macrophages and to trigger an activation signal that inhibited Brucella development. The participation of endogenous cannabinoid ligand(s) in Brucella infection was discussed. Finally, our data show that SR141716A up-regulates the antimicrobial properties of macrophages in vitro and might be a pharmaceutical compound useful for counteracting the development of intramacrophagic gram-negative bacteria.     

Involvement of central and peripheral cannabionoid receptors in the regulation of heart resistance to arrhythmogenic effects of epinephrine

Intravenous injection of the selective cannabinoid receptor agonist HU-210 in doses of 0.05 and 0.25 mg/kg increased heart resistance to arrhythmogenic effects of epinephrine, while intracerebroventricular infusion of this substance had no effect on the incidence of epinephrine-induced arrhythmia. The selective antagonist of type I cannabinoid receptors SR141716A in a dose of 3 mg/kg and ganglion blocker hexamethonium in a dose of 10 mg/kg did not modify the antiarrhythmic effect of HU-210. This effect of HU-210 is probably related to activation of type II peripheral cannabinoid receptors. Translated fromByulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 130, No. 11, pp. 552–554, November, 2000     

Involvement of central and peripheral cannabionoid receptors in the regulation of heart resistance to arrhythmogenic effects of epinephrine

Intravenous injection of the selective cannabinoid receptor agonist HU-210 in doses of 0.05 and 0.25 mg/kg increased heart resistance to arrhythmogenic effects of epinephrine, while intracerebroventricular infusion of this substance had no effect on the incidence of epinephrine-induced arrhythmia. The selective antagonist of type I cannabinoid receptors SR141716A in a dose of 3 mg/kg and ganglion blocker hexamethonium in a dose of 10 mg/kg did not modify the antiarrhythmic effect of HU-210. This effect of HU-210 is probably related to activation of type II peripheral cannabinoid receptors.     

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.     

Selective cannabinoid receptor agonist HU-210 decreases pump function of isolated perfused heart: Role of cAMP and cGMP

The rate and strength of heart contractions decreased after 10-min perfusion of rat myocardium with Krebs—Henseleit solution containing a selective cannabinoid receptor agonist HU-210 in a final concentration of 10 nM. HU-210 completely blocked the positive inotropic and chronotropic effect of -adrenoceptor agonist isoproterenol, decreased the basal level of cAMP, and abolished the isoproterenol-induced increase in myocardial cAMP concentration. cGMP concentration remained unchanged under these conditions. The decrease in myocardial cAMP concentration after activation of cannabinoid receptors did not correlate with changes in the strength and rate of heart contractions. Our results suggest that the negative inotropic and chronotropic effects of HU-210 are not associated with decreased cAMP concentration in the myocardium.Translated from Byulleten Eksperimentalnoi Biologii i Meditsiny, Vol. 138, No. 12, pp. 622–625, December, 2004This revised version was published online in April 2005 with a corrected cover date.     

Endogenous Cannabinoid Anandamide Increases Heart Resistance to Arrhythmogenic Effects of Epinephrine: Role of CB1 and CB2 Receptors

Intravenous injection of 10 mg/kg anandamide reduces the incidence and duration of epinephrine-induced arrhythmias in rats. SR141716A and SR144528, antagonists of cannabinoid receptor I and II did not abolish the antiarrhythmic effect of anandamide. These data suggest that the antiarrhythmic effect of anandamide is nonspecific or mediated via unknown cannabinoid receptors, but not associated with activation of cannabinoid receptors I and II.     

Blockade of cannabinoid receptors reduces inflammation, leukocyte accumulation and neovascularization in a model of sponge-induced inflammatory angiogenesis

Objective

Angiogenesis depends on a complex interaction between cellular networks and mediators. The endocannabinoid system and its receptors have been shown to play a role in models of inflammation. Here, we investigated whether blockade of cannabinoid receptors may interfere with inflammatory angiogenesis.

Materials and methods

Polyester-polyurethane sponges were implanted in C57Bl/6j mice. Animals received doses (3 and 10 mg/kg/daily, s.c.) of the cannabinoid receptor antagonists SR141716A (CB1) or SR144528 (CB2). Implants were collected at days 7 and 14 for cytokines, hemoglobin, myeloperoxidase, and N-acetylglucosaminidase measurements, as indices of inflammation, angiogenesis, neutrophil and macrophage accumulation, respectively. Histological and morphometric analysis were also performed.

Results

Cannabinoid receptors expression in implants was detected from day 4 after implantation. Treatment with CB1 or CB2 receptor antagonists reduced cellular influx into sponges at days 7 and 14 after implantation, although CB1 receptor antagonist were more effective at blocking leukocyte accumulation. There was a reduction in TNF-α, VEGF, CXCL1/KC, CCL2/JE, and CCL3/MIP-1α levels, with increase in CCL5/RANTES. Both treatments reduced neovascularization. Dual blockade of cannabinoid receptors resulted in maximum inhibition of inflammatory angiogenesis.

Conclusions

Blockade of cannabinoid receptors reduced leukocyte accumulation, inflammation and neovascularization, suggesting an important role of endocannabinoids in sponge-induced inflammatory angiogenesis both via CB1 and CB2 receptors.     

Blockade of cannabinoid receptors by SR141716 selectively increases Fos expression in rat mesocorticolimbic areas via reduced dopamine D2 function.

The present study investigated, in rats, whether blockade of cannabinoid CB1 receptors may alter Fos protein expression in a manner comparable to that observed with antipsychotic drugs. Intraperitoneal administration of the selective CB1 receptor antagonist, SR141716, dose-dependently (1.0, 3.0 and 10 mg/kg) increased Fos-like immunoreactivity in mesocorticolimbic areas (prefrontal cortex, ventrolateral septum, shell of the nucleus accumbens and dorsomedial caudate-putamen), while motor-related structures such as the core of the nucleus accumbens and the dorsolateral caudate-putamen were unaffected. In the ventrolateral septum, taken as a representative structure, the Fos-inducing effect of SR141716 (10 mg/kg) was maximal 2 h after injection and returned to near control levels by 4 h. Within the prefrontal cortex, SR141716 increased the number of Fos-positive cells predominantly in the infralimbic and prelimbic cortices, presumptive pyramidal cells being the major cell types in which Fos was induced. The D1-like receptor antagonist, SCH23390 (0.1 mg/kg), did not prevent the Fos-inducing effect of SR141716 in any brain region examined (prefrontal cortex, nucleus accumbens, ventrolateral septum and dorsomedial caudate-putamen), although SCH23390 significantly reduced Fos expression induced by cocaine (20 mg/kg) in all these regions. By contrast, the dopamine D2-like agonist, quinpirole (0.25 mg/ kg), counteracted SR141716-induced Fos-like immunoreactivity in the ventrolateral septum, the nucleus accumbens and the dorsomedial caudate-putamen, while no antagonism was observed in the prefrontal cortex. Microdialysis experiments in awake rats indicated that SR141716, at doses which increased Fos expression (3 and 10 mg/kg), did not alter dopamine release in the shell of the nucleus accumbens. Finally, SR141716 increased the levels of neurotensin-like immunoreactivity in the nucleus accumbens, but not in the caudate-putamen. Collectively, the present results show that blockade of cannabinoid receptors increases Fos- and neurotensin-like immunoreactivity with characteristics comparable to those reported for atypical neuroleptic drugs.     

Hyperlocomotion and paw tremors are two highly quantifiable signs of SR141716-precipitated withdrawal from delta9-tetrahydrocannabinol in C57BL/6 mice

Increasing evidence suggests that marijuana abstinence leads to clinically significant withdrawal symptoms in humans. In mouse models, following chronic treatment with delta9-tetrahydrocannabinol (THC), administration of the selective cannabinoid CB1 receptor antagonist SR141716 (rimonabant) elicited varying behavioral responses, depending on mouse strain and dosing regimen. In the present study, C57BL/6 mice were injected s.c. with THC (25 mg/kg) or vehicle twice daily for 4.5 days. SR141716 (15 mg/kg) was administrated i.p. 4 h following the last THC treatment. During a 2-h observation period immediately following the SR141716 challenge, the total locomotor, ambulatory and stereotypic activities of THC-treated mice were 4.1, 3.3, and 3.8 times those of vehicle-treated mice, respectively. The number of paw tremors elicited in THC-treated mice was 111 ± 11 during the 45 min immediately following SR141716, whereas only 1.1 ± 0.4 was associated with vehicle-treated animals. In contrast, the number of scratching bouts was higher in vehicle-treated (182 ± 20) vs THC-treated (17 ± 4) mice. The present study is the first to demonstrate hyperlocomotion as an explicit sign of THC abstinence in mice. Together with paw tremors, the two unambiguous withdrawal signs may permit highly quantitative investigation of THC abstinence in C57BL/6 mice and may facilitate investigation of the mechanisms involved via both pharmacological and genetic manipulations, and ultimately potential treatments for cannabis dependence.     

Cannabinoid CB1 receptors in the paraventricular nucleus and central control of penile erection: immunocytochemistry, autoradiography and behavioral studies

[N-(piperidin-1-yl)-5-(4-chlorophenyl)-4-methyl-1H-pyrazole-3-carboxyamide] (SR 141716A), a selective cannabinoid CB1 receptor antagonist, injected into the paraventricular nucleus of the hypothalamus (PVN) of male rats, induces penile erection. This effect is mediated by the release of glutamic acid, which in turn activates central oxytocinergic neurons mediating penile erection. Double immunofluorescence studies with selective antibodies against CB1 receptors, glutamic acid transporters (vesicular glutamate transporters 1 and 2 (VGlut1 and VGlut2), glutamic acid decarboxylase-67 (GAD67) and oxytocin itself, have shown that CB1 receptors in the PVN are located mainly in GABAergic terminals and fibers surrounding oxytocinergic cell bodies. As GABAergic synapses in the PVN impinge directly on oxytocinergic neurons or on excitatory glutamatergic synapses, which also impinge on oxytocinergic neurons, these results suggest that the blockade of CB1 receptors decreases GABA release in the PVN, increasing in turn glutamatergic neurotransmission to activate oxytocinergic neurons mediating penile erection. Autoradiography studies with [(3)H](-)-CP 55,940 show that chronic treatment with SR 141716A for 15 days twice daily (1 mg/kg i.p.) significantly increases the density of CB1 receptors in the PVN. This increase occurs concomitantly with an almost twofold increase in the pro-erectile effect of SR 141716A injected into the PVN as compared with control rats. The present findings confirm that PVN CB1 receptors, localized mainly in GABAergic synapses that control in an inhibitory fashion excitatory synapses, exert an inhibitory control on penile erection, demonstrating for the first time that chronic blockade of CB1 receptors by SR 141716A increases the density of these receptors in the PVN. This increase is related to an enhanced pro-erectile effect of SR 141716A, which is still present 3 days after the end of the chronic treatment.     

CB1 cannabinoid receptor inhibits synaptic release of glutamate in rat dorsolateral striatum

CB1 cannabinoid receptors in the neostriatum mediate profound motor deficits induced when cannabinoid drugs are administered to rodents. Because the CB1 receptor has been shown to inhibit neurotransmitter release in various brain areas, we investigated the effects of CB1 activation on glutamatergic synaptic transmission in the dorsolateral striatum of the rat where the CB1 receptor is highly expressed. We performed whole cell voltage-clamp experiments in striatal brain slices and applied the CB1 agonists HU-210 or WIN 55,212-2 during measurement of synaptic transmission. Excitatory postsynaptic currents (EPSCs), evoked by electrical stimulation of afferent fibers, were significantly reduced in a dose-dependent manner by CB1 agonist application. EPSC inhibition was accompanied by an increase in two separate indices of presynaptic release, the paired-pulse response ratio and the coefficient of variation, suggesting a decrease in neurotransmitter release. These effects were prevented by application of the CB1 antagonist SR141716A. When Sr(2+) was substituted for Ca(2+) in the extracellular solution, application of HU-210 (1 microM) significantly reduced the frequency, but not amplitude, of evoked, asynchronous quantal release events. Spontaneous release events were similarly decreased in frequency with no change in amplitude. These findings further support the interpretation that CB1 activation leads to a decrease of glutamate release from afferent terminals in the striatum. These results reveal a novel potential role for cannabinoids in regulating striatal function and thus basal ganglia output and may suggest CB1-targeted drugs as potential therapeutic agents in the treatment of Parkinson's disease and other basal ganglia disorders.     

Behavioral effects of cannabinoid agents in animals

Two subtypes of cannabinoid receptors have been identified to date, the CB1 receptor, essentially located in the CNS, but also in peripheral tissues, and the CB2 receptor, found only at the periphery. The identification of delta9-tetrahydrocannabinol (delta9-THC) as the major active component of marijuana (Cannabis sativa), the recent emergence of potent synthetic ligands and the identification of anandamide and sn-2 arachidonylglycerol as putative endogenous ligands for cannabinoid receptors in the brain, have contributed to advancing cannabinoid pharmacology and approaching the neurobiological mechanisms involved in physiological and behavioral effects of cannabinoids. Most of the agonists exhibit nonselective affinity for CB1/CB2 receptors, and delta9-THC and anandamide probably act as partial agonists. Some recently synthesized molecules are highly selective for CB2 receptors, whereas selective agonists for the CB1 receptors are not yet available. A small number of antagonists exist that display a high selectivity for either CB1 or CB2 receptors. Cannabinomimetics produce complex pharmacological and behavioral effects that probably involve numerous neuronal substrates. Interactions with dopamine, acetylcholine, opiate, and GABAergic systems have been demonstrated in several brain structures. In animals, cannabinoid agonists such as delta9-THC, WIN 55,212-2, and CP 55,940 produce a characteristic combination of four symptoms, hypothermia, analgesia, hypoactivity, and catalepsy. They are reversed by the selective CB1 receptor antagonist, SR 141716, providing good evidence for the involvement of CB1-related mechanisms. Anandamide exhibits several differences, compared with other agonists. In particular, hypothermia, analgesia, and catalepsy induced by this endogenous ligand are not reversed by SR 141716. Cannabinoid-related processes seem also involved in cognition, memory, anxiety, control of appetite, emesis, inflammatory, and immune responses. Agonists may induce biphasic effects, for example, hyperactivity at low doses and severe motor deficits at larger doses. Intriguingly, although cannabis is widely used as recreational drug in humans, only a few studies revealed an appetitive potential of cannabimimetics in animals, and evidence for aversive effects of delta9-THC, WIN 55,212-2, and CP 55,940 is more readily obtained in a variety of tests. The selective blockade of CB1 receptors by SR 141716 impaired the perception of the appetitive value of positive reinforcers (food, cocaine, morphine) and reduced the motivation for sucrose, beer and alcohol consumption, indicating that positive incentive and/or motivational processes could be under a permissive control of CB1-related mechanisms. There is little evidence that cannabinoid systems are activated under basal conditions. However, by using SR 141716 as a tool, a tonic involvement of a CB1-mediated cannabinoid link has been demonstrated, notably in animals suffering from chronic pain, faced with anxiogenic stimuli or highly motivational reinforcers. Some effects of SR 141716 also suggest that CB1-related mechanisms exert a tonic control on cognitive processes. Extensive basic research is still needed to elucidate the roles of cannabinoid systems, both in the brain and at the periphery, in normal physiology and in diseases. Additional compounds, such as selective CB1 receptor agonists, ligands that do not cross the blood brain barrier, drugs interfering with synthesis, degradation or uptake of endogenous ligand(s) of CB receptors, are especially needed to understand when and how cannabinoid systems are activated. In turn, new therapeutic strategies would likely to emerge.     

The cannabinoid antagonist SR141716A facilitates memory acquisition and consolidation in the mouse elevated T-maze

Δ⁹-THC and synthetic cannabinoids produce memory impairment in humans as well as in laboratory animals. The high concentration of cannabinoid CB1 receptors and the presence of endocannabinoids in the hippocampus suggest that a cannabinoid neurochemical system may play a role in learning and memory processes. Thus, the objective of the present work was to study the effect of the cannabinoid antagonist SR141716A (SR) on memory acquisition, consolidation and retrieval in a recently developed elevated T-maze (ETM) model of anxiety and memory. In addition, we investigated whether pre-training SR administration was capable of reversing scopolamine-induced memory impairment. Adult male mice were exposed to the closed arm as many times as necessary for the animals to reach the avoidance criterion of remaining in the closed arm for 300 s; they were then tested (exposed to the closed arm) 24 h and 7 days after the training. SR (0.5, 1.0 or 2.0 mg/kg) was administered i.p. 20 min before the training, immediately after training or 20 min before the test in the mice. The elevated plus-maze (EPM) was used to investigate a possible influence of SR on locomotion and on the anxiety-related behavior. SR provoked memory improvement, which was observed when the drug was administered before (effect on memory acquisition/consolidation) or immediately after the training (effect on memory consolidation), but not when the drug was administered before the test (effect on memory retrieval). Also, SR administration reversed scopolamine-induced amnesia. These effects were observed in the absence of changes in locomotion or anxiety levels. Our results demonstrate that the blockade of cannabinoid receptors may improve memory acquisition and consolidation in the ETM model.     

Activation of CB1 cannabinoid receptors impairs memory consolidation and hippocampal polysialylated neural cell adhesion molecule expression in contextual fear conditioning

We investigated the role of CB1 receptors in hippocampal-dependent memory consolidation mediated by polysialylated neural cell adhesion molecule (PSA-NCAM) during contextual fear conditioning (CFC). The CB1 receptor agonist 3-(1,1-dimethylheptyl)-(-)-11-hydroxy-Δ⁸-tetrahydrocannabinol (HU-210) (0.1 mg/kg) was given immediately after training during the memory consolidation phase, and freezing behavior was measured 24 h after conditioning. Administration of HU-210 attenuated freezing behavior measured in CFC. Western blot analysis showed that CFC induced a decrease in the expression of NCAM-180, but did not change the level of NCAM-140 and increased PSA-NCAM expression measured 24 h after training in the rat hippocampus. HU-210 (0.1 mg/kg) injection did not affect the reduction in NCAM-180 levels induced by CFC, but it blocked the increase in PSA-NCAM expression. Since the dentate gyrus (DG) of the hippocampus is known to be involved in memory consolidation and expresses a high level of PSA-NCAM protein, we measured the effects of CFC and HU-210 administration on PSA-NCAM-immunoreactive (IR) cells in the DG. CFC caused an increase in the number of PSA-NCAM-IR cells in the DG, but not K_i-67- or doublecortin (DCX)-IR cells. This increase in PSA-NCAM-IR cells was abolished by HU-210 injection. Administration of the CB1 receptor antagonist N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM-251) (3 mg/kg immediately before HU-210) inhibited the effects of HU-210 on freezing behavior and PSA-NCAM expression in the DG. These results indicate that activation of CB1 receptors disturbs consolidation of fear memory in CFC, likely by affecting PSA-NCAM expression in the DG, which plays an important role in synaptic rearrangement during the formation of memory traces.     

EEG alpha power changes reflect response inhibition deficits after traumatic brain injury (TBI) in humans

We studied the effect of cannabinoids on the activity of N-methyl-d-aspartate (NMDA) receptors in the locus coeruleus from rat brain slices by single-unit extracellular recordings. As expected, NMDA (100 microM) strongly excited (by nine fold) the cell firing activity of the locus coeruleus. Perfusion with the endocannabinoid anandamide (1 and 10 microM) or the anandamide transport inhibitor AM 404 (30 microM) enhanced the NMDA-induced excitation of locus coeruleus neurons. Similarly, the synthetic agonists R(+)-WIN 55212-2 (10 microM) and CP 55940 (30 microM) enhanced the effect of NMDA. In the presence of the CB(1) receptor antagonists SR 141716A (1 microM) or AM 251 (1 microM), the enhancement induced by anandamide (10 microM) was blocked. Our results suggest that cannabinoids modulate the activity of NMDA receptors in the locus coeruleus through CB(1) receptors.     

Concentration-dependent dual effect of anandamide on sensory neuropeptide release from isolated rat tracheae

Most actions of anandamide (AEA) are mediated by the cannabinoid 1 (CB(1)) receptor activation, but on sensory neurones it is also an agonist on the vanilloid subtype 1 receptor (VR(1)). The aim of the present study was to analyse the effect of AEA (10(-6)-10(-4) M) on inhibitory CB(1) and excitatory VR(1) receptors by measuring sensory neuropeptide release such as somatostatin, substance P and calcitonin gene-related peptide, from isolated rat tracheae. AEA (10(-6) M) vas without significant effect, 10(-5) M inhibited neuropeptide release, which was abolished by the G protein-coupled receptor blocker pertussis toxin (100 ng/ml) and the CB(1) receptor antagonist SR141716A (5x10(-7) M). High concentrations of AEA (5x10(-5) M, 10(-4) M) increased the release of the peptides and this inhibition was prevented by the competitive VR(1) antagonist capsazepine (10(-5) M). These results indicate a dual, concentration-dependent action of AEA on CB(1) receptors and VR(1) on peripheral sensory nerve terminals.