The cannabinoid CB(1) receptor antagonist CE prolongs spatial memory duration in a rat delayed radial arm maze memory task

The cannabinoid receptor system plays an integral role in learning and memory. Moreover, the cannabinoid CB(1) receptor antagonist rimonabant has been found to improve performance in a variety of animal memory models. The present study tested whether a novel and potent cannabinoid CB(1) receptor antagonist, CE, would prolong the duration of spatial memory. Rats were trained in a two-phase radial arm maze procedure, consisting of acquisition and retrieval tests, which were separated by an 18 h delay. CE was administered 30 min before the acquisition phase, immediately after the acquisition phase, or 30 min before the retrieval test to assess its effects on acquisition and retrieval processes. CE administered before and immediately after the acquisition phase significantly decreased the number of errors committed during the retrieval test. On the other hand, CE administered 30 min before the retrieval test had no effect on the number of errors committed. These findings demonstrate that CE improves memory by acting on consolidation, rather than retrieval, processes and further suggest that the endocannabinoid system has an important role in modulating memory duration.     

Additive subthreshold dose effects of cannabinoid CB(1) receptor antagonist and selective serotonin reuptake inhibitor in antidepressant behavioral tests

The main clinically used antidepressant drugs are selective monoamine reuptake inhibitors, including selective serotonin reuptake inhibitors (citalopram, sertraline), selective dopamine reuptake inhibitor (nomifensine) and selective noradrenaline reuptake inhibitor (reboxetine), but they have various side effects. Because cannabinoid CB(1) receptor antagonists (SR141716A, AM251) enhance monoamine release, they might be beneficial in the therapy of affective disorders. We hypothesized that the use of monoamine reuptake inhibitors in combination with cannabinoid CB(1) receptor antagonists would allow a lower dose of monoamine reuptake inhibitors to be used in the therapy of depression, thereby reducing or eliminating the side effects. To test this hypothesis, we examined the combination of SR141716A or AM251 with citalopram, sertraline, nomifensine or reboxetine at subthreshold doses to see whether these combinations would show an additive effect in the forced swimming test and the tail suspension test with mice. Subthreshold doses of cannabinoid CB(1) receptor antagonist and selective serotonin reuptake inhibitors, which separately had no effect on the immobility of mice in the tests, showed a clear effect when the drugs were administered at 40 and 30 min, respectively, before the tests, without any change of motor activity. Therefore, the use of subthreshold doses of these agents in combination might be useful to enhance mainly serotonergic neurotransmission, and to reduce or eliminate the side effects of citalopram and sertraline.     

Novel pharmacological targets based on receptor heteromers

Studies performed in the last 10 years have provided solid evidence indicating that G-protein-coupled receptors are expressed on the plasma membrane as homo and heterodimers. The first consequence of this fact is that homo and heterodimers are the true targets of natural (hormones, neurotransmitters) and synthetic drugs. Furthermore a given receptor in a heteromer may display a different functional and/or pharmacological profile than the same receptor characterized as monomer or as homodimer. Recent evidence indicates that receptor heteromers are sensors that lead to a fine-tuning in neurotransmission or hormone regulation; mainly this is achieved by a modification of the signaling pathways activated via a given receptor when it is forming a given heteromer. Quite often antagonists display variable affinities when a given receptor is expressed with different heteromeric partners. This fact should be taken into account in the development of new drugs. Finally it should be pointed out that radioligand binding data has to be analyzed by a model that considers receptors as dimers and not as monomers. This model provides a novel approach to characterize drugs interacting with the orthosteric center (agonists/antagonists) or with allosteric centers (allosteric regulators).     

Description of a Bivalent Cannabinoid Ligand with Hypophagic Properties

A series of bivalent cannabinoid ligands is proposed. The synthesis of double amides based on the rimonabant structure separated by an alkyl chain and the evaluation of their affinities for cannabinoid receptors are reported. The data of 4d confirmed that a bivalent structure is a suitable scaffold for CB₁ cannabinoid receptor binding. The compound 4d was selected for in vitro and in vivo pharmacological evaluations. Moreover, intraperitoneal administration of 4d to food‐deprived rats resulted in a dose‐dependent inhibition of feeding that was maintained up to 240 min.     

慢性胃食管反流大鼠背根神经节中辣椒素受体1和大麻素受体的表达

背景：食管痛觉高敏参与胃食管反流病（GERD）的发病,而辣椒素受体1（TRPV1）和大麻素系统的激活在疼痛调控中发挥双向作用。目的：检测慢性胃食管反流大鼠相应节段背根神经节（DRG）中TRPV1和大麻素受体（CB1、CB2）的表达情况,探讨两者在GERD中的作用。方法：雄性Sprague-Dawley大鼠随机分为反流组（R组）、对照组（S组）。采用胃底结扎联合幽门限制法构建慢性胃食管反流模型。采用免疫荧光法和蛋白质印迹法检测大鼠DRG中TRPV1和CB1、CB2的表达,并分析TRPV1蛋白表达与CB1、CB2蛋白表达的相关性。结果：免疫荧光法示R组DRG中TRPV1表达较S组显著上调（905.24±134.82对648.43±135.13,P=0.000）;而CB1（677.06±123.75对836.89±101.00,P=0.013）、CB2（513.99±79.80对709.63±43.25,P=0.000）表达显著下调。蛋白质印迹法亦显示,R组TRPV1蛋白表达显著高于S组（0.98±0.01对0.64±0.09,P=0.001）,CB1（0.86±0.05对0.96±0.06,P=0.013）、CB2（0.75±0.03对0.81±0.05,P=0.019）蛋白表达显著低于S组。TRPV1蛋白表达与 CB1蛋白表达呈负相关（r=－0.836,P=0.001）,而与CB2蛋白表达无关（r=－0.351,P=0.263）。结论：慢性胃食管反流大鼠中TRPV1蛋白表达上调与CB1蛋白表达下调密切相关,酸在调节受体分子中起关键作用。     

外周内源性大麻素系统在快眼动睡眠剥夺所致大鼠内脏感觉降低中的作用

背景：内脏感觉过敏是功能性胃肠病的重要病理生理机制之一．快眼动（REM）睡眠剥夺可降低大鼠内脏感觉,但具体机制尚不明。目的：研究外周内源性大麻素系统在REM睡眠剥夺所致大鼠内脏感觉降低中的作用。方法：24只雄性Sprague—Dawley大鼠随机分为实验对照组、睡眠剥夺组和利莫那班干预组。采用花瓶技术制作REM睡眠剥夺大鼠模型。睡眠剥夺48h后行结直肠扩张（CRD）,记录腹壁肌电图,以反映内脏感觉功能的变化。采用逆转录聚合酶链反应（RT—PCR）和蛋白质印迹法分别测定大鼠肠道组织中1型大麻素（CBl）受体、脂肪酰胺水解酶（FAAH）和单酰基甘油脂酶（MAGL）mRNA和蛋白表达。结果：予不同扩张压力（40、60和80mmHg）后,睡眠剥夺组大鼠腹外斜肌放电次数显著低于实验对照组（P〈0．05）;利莫那班干预组放电次数显著高于睡眠剥夺组（P〈0．05）,但与实验对照组无明显差异。与实验对照组相比,睡眠剥夺组大鼠肠道CB1受体mRNA表达上调（P〈0．05）,结肠FAAH和MAGLmRNA和蛋白表达显著下调（P〈0．05）。结论：REM睡眠剥夺降低大鼠内脏感觉的作用与外周内源性大麻素代谢降低有关。     

Detailed analysis of food-reinforced operant lever pressing distinguishes effects of a cannabinoid CB1 inverse agonist and dopamine D1 and D2 antagonists

Overt similarities exist between the effects of systemic cannabinoid CB1 inverse agonists and dopamine (DA) antagonists on appetitive behavior. The present set of studies was undertaken to apply a fine-grained analysis of food-reinforced operant lever pressing in rats in order to compare the pattern of effects produced by administration of the CB1 inverse agonist AM 251 and those induced by the DA D1 antagonist SKF 83566, and the D2 antagonist raclopride. Three groups of rats were trained on a fixed-ratio 5 (FR5) schedule and administered these compounds over a range of doses expected to suppress responding. All three drugs produced a dose-related suppression of total lever pressing. In addition to main effects of dose, regression analyses were performed to determine which of several response timing- and rate-related variables correlated most strongly with overall responding in each group. It was found that total session time spent pausing from responding was significantly better at predicting responding in the AM 251 group, while both DA antagonists produced significantly stronger regression coefficients (versus AM 251) from fast responding measures. These results suggest that, while several similarities exist, CB1, D1, and D2 antagonists are not identical in their pattern of suppression of food-maintained lever pressing.     

Acute cannabinoid administration attenuates female socio-sexual motivation

Endocannabinoids may normally inhibit the generation and expression of female estrous behaviors. Previous work in our laboratory demonstrated that acute administration of a CB₁ receptor antagonist (AM251) increased sexual incentive motivation in estrous female rats. The current experiment examined the effect of CP55,940, a synthetic cannabinoid agonist, on sexual motivation. Seventy-two ovariectomized female Long-Evans rats were tested for their socio-sexual motivation via a runway methodology. Baseline motivation to approach and maintain close proximity to an empty goalbox, a female conspecific, and a male conspecific was assessed over six trials. Subjects were then grouped into nine experimental conditions and re-tested for their socio-sexual motivation after one of three possible hormonal treatments and three drug doses. Hormone treatments were: oil (nonestrous), 10 μg estradiol benzoate (partially estrous), and 10 μg estradiol + 500 μg progesterone (fully estrous). Drug doses were: 0, 20, or 40 μg/kg CP55,940 (IP, 30 min prior to testing). As expected, hormonal priming with both estradiol and progesterone significantly increased sexual motivation in females that did not receive drug treatment. This occurred even though females were kept sexually-naïve throughout the experiment. CP55,940 dose-dependently attenuated sexual motivation for a male target in estrous females; the 40 μg/kg dose completely blocked sexual motivation. However, this same dose also significantly reduced social motivation for another female. Cannabinoid agonists reduce female sexual motivation, either directly by inhibiting estrus or indirectly by increasing social anxiety.     

Possible inhibitory roles of endogenous 2-arachidonoylglycerol during corticotropin-releasing factor-induced activation of central sympatho-adrenomedullary outflow in anesthetized rats

We previously reported that intracerebroventricularly (i.c.v.) administered corticotropin-releasing factor (CRF) (0.5-3.0 nmol/animal) dose-dependently elevates plasma noradrenaline and adrenaline through brain phospholipase C-, diacylglycerol lipase- and prostanoids-mediated mechanisms in rats. Diacylglycerol produced by phospholipase C from phospholipids can be hydrolyzed by diacylglycerol lipase into 2-arachidonoylglycerol, which may be further hydrolyzed by monoacylglycerol lipase into arachidonic acid, a precursor of prostanoids. Recently, 2-arachidonoylglycerol has been recognized as a major brain endocannabinoid, which can modulate synaptic transmission through presynaptic cannabinoid CB₁ receptors. Released 2-arachidonoylglycerol is rapidly deactivated by uptake into cells and enzymatic hydrolysis. In the present study, therefore, we examined (1) the involvement of brain 2-arachidonoylglycerol, (2) the regulatory role of 2-arachidonoylglycerol as a brain endocannabinoid, and (3) the effect of exogenous cannabinoid receptor agonist, on the CRF-induced elevation of plasma noradrenaline and adrenaline using anesthetized rats. The elevation of both catecholamines induced by a submaximal dose of CRF (1.5 nmol/animal, i.c.v.) was reduced by i.c.v. administered MAFP (monoacylglycerol lipase inhibitor) (0.7 and 1.4 µmol/animal), AM 404 (endocannabinoid uptake-inhibitor) (80 and 250 nmol/animal) and ACEA (cannabinoid CB₁ receptor agonist) (0.7 and 1.4 µmol/animal), while AM 251 (cannabinoid CB₁ receptor antagonist) (90 and 180 nmol/animal, i.c.v.) potentiated the response induced by a small dose of CRF (0.5 nmol/animal, i.c.v.). These results suggest a possibility that 2-arachidonoylglycerol is endogenously generated in the brain during CRF-induced activation of central sympatho-adrenomedullary outflow, thereby inhibiting the peptide-induced response by activation of brain cannabinoid CB₁ receptors in anesthetized rats.     

Anxiety-like effects of SR141716-precipitated delta9-tetrahydrocannabinol withdrawal in mice in the elevated plus-maze

Marijuana discontinuation has been recently reported to be anxiogenic in humans, which may predict relapse. Limited animal research has been carried out to model this withdrawal-associated negative affect. The current study sought to investigate the potential anxiety-like effects of cannabinoid withdrawal in mice. Male ICR mice were injected s.c. with delta9-tetrahydrocannabinol (THC) at 10 mg/kg or vehicle once daily for 10 days. To precipitate withdrawal, the cannabinoid CB1 antagonist SR141716 (0.3, 1.0, or 3.0 mg/kg) or vehicle was administrated i.p. 4 h following the last THC or vehicle treatment. Thirty minutes later, mice were tested on the elevated plus-maze (EPM) for 5 min. SR141716 did not significantly change EPM behaviors in vehicle-treated mice. In contrast, SR141716 precipitated a reduction in exploration of the open arms of EPM in mice repeatedly treated with THC vs vehicle. At 3.0 mg/kg, SR141716 significantly reduced % open arm entries of the total arm entries, % open arm time of total time in arms, and the absolute time spent in open arms. No significant differences in the number of closed or total arm entries were observed, indicating that the behavioral changes were not due to altered motor activity. Collectively, the present results constitute the first evidence that cannabinoid withdrawal produces anxiety-like effects in mice. This animal model may help to identify the mechanisms that contribute to adaptations in the neuronal circuitry of the brain that are expressed as emotional symptoms of cannabinoid withdrawal.