Cardiovascular risk management by blocking the endocannabinoid system.

The specific blockade of endocannabinoids at the level of the cannabinoid receptor 1 (CB (1) receptor) is a new therapeutic option to reduce body weight and manage cardiovascular risk. Although clinical trials are underway to document the safety and efficacy of this approach, much is still unknown about this endogenous system. Endocannabinoids and their receptors are expressed in the central nervous system as well as in the periphery and regulate the central neural circuits for food uptake and peripheral metabolic circuits. Within the context of food uptake, the stimulation of the CB (1) receptor with Delta (9)-tetrahydrocannabinol (Delta (9)-THC) enhances food consumption, while its blockade with receptor antagonists is an emerging relevant therapeutic means to reduce body weight. Rimonabant is the first of a new class of drugs that interferes with the endocannabinoid system by blocking the CB (1) receptor. In recent clinical studies, a substantial reduction in body weight and waist circumference was associated with an improvement of the cardiovascular risk profile. In particular, increased HDL cholesterol, decreased serum triglycerides and improved insulin sensitivity were observed. Further research will serve to establish the role of these compounds in cardiovascular risk management.     

Endocannabinoids and the control of energy balance.

Two receptors have been cloned to date for the psychotropic compound Delta(9)-tetrahydrocannabinol, and termed cannabinoid CB(1) and CB(2) receptors. Their endogenous ligands, the endocannabinoids, have also been identified. CB(1) receptors and endocannabinoids are present in brain structures controlling energy intake and in peripheral cells (hepatocytes, adipocytes, pancreatic islet cells) regulating energy homeostasis. CB(2) receptors are more abundant in lymphocytes and macrophages, and participate in immune and inflammatory reactions. Metabolic hormones and peptides regulate the levels of the endocannabinoids and, hence, the activity of cannabinoid receptors in several tissues in a seemingly coordinated way. The endocannabinoids, particularly after stress and brief food deprivation, act in turn as local modulators of the expression and action of neurotransmitters, hormones and adipokines involved in metabolic control. Endocannabinoid overactivity seems to accompany metabolic and eating disorders and to contribute to the development of abdominal obesity, dyslipidemia and hyperglycemia. Accordingly, clinical trials have shown that CB(1) receptor antagonists are efficacious at reducing not only food intake, but also abdominal adiposity and its metabolic sequelae.     

Blocking the endocannabinoid system -- weight reduction and cardiovascular risk management

Blocking the endocannabinoid system is an option that substantially reduces cardiovascular risk beyond reducing body weight. Endocannabinoids and their receptors are expressed in the central nervous system as well as in the peripheral organs and regulate the central circuits for food uptake and peripheral metabolic circuits. Within the context of food uptake the cannabinoid receptors 1 (CB (1)-receptor) is of crucial importance. Its stimulation with Delta (9)-tetrahydrocannabiol (Delta (9)-THC) or its blockade with rimonabant are clinically relevant therapeutic means to maintain body weight. Rimonabant is the first of a new class of drugs, that interferes with the endocannabinoid system by blocking the CB (1)-Receptor. In recent clinical studies a substantial reduction of body weight and waist circumference was associated with an improvement of the cardiovascular risk profile, which was marked by increased HDL-cholesterol, serum triglycerides and improved insulin sensivity.     

Impact of endocannabinoid system in modulation of cardiometabolic risk factors

Endocannabinoid system, the complex of specific cannabinoid receptors (CB1 and CB2 subtypes) and their endogenous agonistic ligands (endocannabinoids) plays, besides others, an important role in the central and peripheral regulation of food intake, fat accumulation, and lipid and glucose metabolism. Alterations of these functions are associated with endocannabinoid system hyperactivity. The cannabinoid receptor CB1 antagonist rimonabant normalizes the over activated endocannabinoid system which contributes to the regulation of energy homeostasis, and improves lipid and glucose metabolism--decreases body weight, waist circumference, intra-abdominal obesity and triglycerides, increases HDL-C, improves insulin sensitivity according to HOMA index. Results of the international multicentric clinical trials confirm that rimonabant is well tolerated and show antiatherogenic effects (increased adiponectin, decreased marker of inflammation CRP and improvement of LDL profile) as well as decreased percentage of subjects with NCEP/ATPIII (National Cholesterol Education Program Adult Treatment Panel III) defined metabolic syndrome. Thus, the CB1 cannabinoid receptor antagonist rimonabant is suggested to be a prospective drug decreasing cardiometabolic risk factors.     

GPR55 is a cannabinoid receptor that increases intracellular calcium and inhibits M current

The CB(1) cannabinoid receptor mediates many of the psychoactive effects of Delta(9)THC, the principal active component of cannabis. However, ample evidence suggests that additional non-CB(1)/CB(2) receptors may contribute to the behavioral, vascular, and immunological actions of Delta(9)THC and endogenous cannabinoids. Here, we provide further evidence that GPR55, a G protein-coupled receptor, is a cannabinoid receptor. GPR55 is highly expressed in large dorsal root ganglion neurons and, upon activation by various cannabinoids (Delta(9)THC, the anandamide analog methanandamide, and JWH015) increases intracellular calcium in these neurons. Examination of its signaling pathway in HEK293 cells transiently expressing GPR55 found the calcium increase to involve G(q), G(12), RhoA, actin, phospholipase C, and calcium release from IP(3)R-gated stores. GPR55 activation also inhibits M current. These results establish GPR55 as a cannabinoid receptor with signaling distinct from CB(1) and CB(2).     

The Role of Endocannabinoids System in Fatty Liver Disease and Therapeutic Potentials

Non-alcoholic fatty liver disease (NAFLD) is a major cause of liver morbidity and mortality with no proven effective therapy as of yet. Its prevalence is increasing globally in parallel with obesity and metabolic syndrome pandemic. The endocannabinoid (EC) system has been implicated in the pathogenesis of several diseases, including fatty liver diseases. This system refers to the cannabinoid receptors type 1 (CB1) and type 2 (CB2), with both their endogenous ligands and machinery dedicated to EC synthesis and degradation. There is accumulating evidence on the role CB1 as a key mediator of insulin resistance and liver lipogenesis in both animals and humans. On the other hand, CB2 receptors have been shown to promote inflammation with anti-fibrogenic properties. The pharmacological modulation of the EC system activity for the treatment of metabolic syndrome and NAFLD are promising yet premature. The initial limited success due to deleterious central nervous system side-effects are likely to be bypassed with the use of peripherally restricted drugs.     

Endocannabinoid overactivity and intestinal inflammation

Cannabinoid receptors of type 1 and 2 (CB(1) and CB(2)), endogenous ligands that activate them (endocannabinoids), and mechanisms for endocannabinoid biosynthesis and inactivation have been identified in the gastrointestinal system. Activation of CB(1 )receptors by endocannabinoids produces relaxation of the lower oesophageal sphincter and inhibition of gastric acid secretion, intestinal motility, and fluid stimulated secretion. However, stimulation of cannabinoid receptors impacts on gastrointestinal functions in several other ways. Recent data indicate that the endocannabinoid system in the small intestine and colon becomes over stimulated during inflammation in both animal models and human inflammatory disorders. The pathological significance of this "endocannabinoid overactivity" and its possible exploitation for therapeutic purposes are discussed here.     

Endocannabinoids regulate interneuron migration and morphogenesis by transactivating the TrkB receptor

In utero exposure to Delta(9)-tetrahydrocannabinol (Delta(9)-THC), the active component from marijuana, induces cognitive deficits enduring into adulthood. Although changes in synaptic structure and plasticity may underlie Delta(9)-THC-induced cognitive impairments, the neuronal basis of Delta(9)-THC-related developmental deficits remains unknown. Using a Boyden chamber assay, we show that agonist stimulation of the CB(1) cannabinoid receptor (CB(1)R) on cholecystokinin-expressing interneurons induces chemotaxis that is additive with brain-derived neurotrophic factor (BDNF)-induced interneuron migration. We find that Src kinase-dependent TrkB receptor transactivation mediates endocannabinoid (eCB)-induced chemotaxis in the absence of BDNF. Simultaneously, eCBs suppress the BDNF-dependent morphogenesis of interneurons, and this suppression is abolished by Src kinase inhibition in vitro. Because sustained prenatal Delta(9)-THC stimulation of CB(1)Rs selectively increases the density of cholecystokinin-expressing interneurons in the hippocampus in vivo, we conclude that prenatal CB(1)R activity governs proper interneuron placement and integration during corticogenesis. Moreover, eCBs use TrkB receptor-dependent signaling pathways to regulate subtype-selective interneuron migration and specification.     

Peripheral cannabinoid receptor, CB2, regulates bone mass

The endogenous cannabinoids bind to and activate two G protein-coupled receptors, the predominantly central cannabinoid receptor type 1 (CB1) and peripheral cannabinoid receptor type 2 (CB2). Whereas CB1 mediates the cannabinoid psychotropic, analgesic, and orectic effects, CB2 has been implicated recently in the regulation of liver fibrosis and atherosclerosis. Here we show that CB2-deficient mice have a markedly accelerated age-related trabecular bone loss and cortical expansion, although cortical thickness remains unaltered. These changes are reminiscent of human osteoporosis and may result from differential regulation of trabecular and cortical bone remodeling. The CB2(-/-) phenotype is also characterized by increased activity of trabecular osteoblasts (bone-forming cells), increased osteoclast (the bone-resorbing cell) number, and a markedly decreased number of diaphyseal osteoblast precursors. CB2 is expressed in osteoblasts, osteocytes, and osteoclasts. A CB2-specific agonist that does not have any psychotropic effects enhances endocortical osteoblast number and activity and restrains trabecular osteoclastogenesis, apparently by inhibiting proliferation of osteoclast precursors and receptor activator of NF-kappaB ligand expression in bone marrow-derived osteoblasts/stromal cells. The same agonist attenuates ovariectomy-induced bone loss and markedly stimulates cortical thickness through the respective suppression of osteoclast number and stimulation of endocortical bone formation. These results demonstrate that the endocannabinoid system is essential for the maintenance of normal bone mass by osteoblastic and osteoclastic CB2 signaling. Hence, CB2 offers a molecular target for the diagnosis and treatment of osteoporosis, the most prevalent degenerative disease in developed countries.     

Differential expression of cannabinoid receptors in the human colon: cannabinoids promote epithelial wound healing

BACKGROUND & AIMS: Two G-protein-coupled cannabinoid receptors, termed CB1 and CB2, have been identified and several mammalian enteric nervous systems express CB1 receptors and produce endocannabinoids. An immunomodulatory role for the endocannabinoid system in gastrointestinal inflammatory disorders has been proposed and this study sought to determine the location of both cannabinoid receptors in human colon and to investigate epithelial receptor function. METHODS: The location of CB1 and CB2 receptors in human colonic tissue was determined by immunohistochemistry. Primary colonic epithelial cells were treated with both synthetic and endogenous cannabinoids in vitro, and biochemical coupling of the receptors to known signaling events was determined by immunoblotting. Human colonic epithelial cell lines were used in cannabinoid-binding studies and as a model for in vitro wound-healing experiments. RESULTS: CB1-receptor immunoreactivity was evident in normal colonic epithelium, smooth muscle, and the submucosal myenteric plexus. CB1- and CB2-receptor expression was present on plasma cells in the lamina propria, whereas only CB2 was present on macrophages. CB2 immunoreactivity was seen in the epithelium of colonic tissue characteristic of inflammatory bowel disease. Cannabinoids enhanced epithelial wound closure either alone or in combination with lysophosphatidic acid through a CB1-lysophosphatidic acid 1 heteromeric receptor complex. CONCLUSIONS: CB1 receptors are expressed in normal human colon and colonic epithelium is responsive biochemically and functionally to cannabinoids. Increased epithelial CB2-receptor expression in human inflammatory bowel disease tissue implies an immunomodulatory role that may impact on mucosal immunity.     

内源性大麻素系统对肝硬化的影响

内源性大麻素系统包括内源性大麻素物质、特异性大麻素受体1(cannabinoid receptor1,CB1)和大麻素受体2(cannabinoid receptor2,CB2).许多研究已经证实在慢性肝病发展过程中,肝脏肌成纤维细胞和血管内皮细胞的CB1和CB2表达升高,内源性大麻素系统与肝纤维化及肝硬化关系密切.本文就内源性大麻素系统、内源性大麻素系统和脂肪性肝病、内源性大麻素系统和肝纤维化、内源性大麻素系统和肝硬化并发症等作一综述.     

Expression and function of cannabinoid receptors in mouse islets

The endocannabinoid system plays a key role in energy homeostasis, with agonists and antagonists of CB1 receptors acting centrally to stimulate and inhibit food intake, respectively. In addition to their established effects on the central nervous system, cannabinoid receptor agonists also exert peripheral effects by modulating cellular cyclic AMP and calcium levels and there have been reports that they regulate β-cell function. However, the few reports to date on islet expression of cannabinoid receptors and effects of agonists on insulin secretion have failed to reach a consensus. We have therefore investigated cannabinoid receptor expression by mouse islet β-and α-cells and the effects of selective receptor agonists on cyclic AMP and calcium levels, and on dynamic insulin secretory responses. CB1 and CB2 mRNA and protein expression by islets was detected by RT-PCR and western blotting respectively, and cellular location of the receptors was identified by immunohistochemistry with insulin and glucagon antibody co-staining. Cyclic AMP generation was quantified by enzyme immunoassay and changes in calcium levels were measured by microfluorimetry of Fura-2-loaded mouse islet cells. Dynamic insulin secretion was quantified by radioimmunoassay after perifusion of isolated islets. We found that mouse islets expressed both CB1 and CB2 receptors, and they were localised to β-cells. Activation of mouse β-cell CB1 and CB2 receptors resulted in decreased cyclic AMP, increased calcium and potentiation of glucose-stimulated insulin secretion. Thus, activation of islet cannabinoid receptors by locally produced endocannabinoids such as 2-aminoglycerol may be another regulatory pathway by which islets stimulate insulin secretion to maintain glucose homeostasis.     

Evidence for an interaction between CB1 cannabinoid and melanocortin MCR-4 receptors in regulating food intake

Melanocortin receptor 4 (MCR4) and CB(1) cannabinoid receptors independently modulate food intake. Although an interaction between the cannabinoid and melanocortin systems has been found in recovery from hemorrhagic shock, the interaction between these systems in modulating food intake has not yet been examined. The present study had two primary purposes: 1) to examine whether the cannabinoid and melanocortin systems act independently or synergistically in suppressing food intake; and 2) to determine the relative position of the CB(1) receptors in the chain of control of food intake in relation to the melanocortin system. Rats were habituated to the test environment and injection procedure and then received intracerebroventicular injections of various combinations of the MCR4 receptor antagonist JKC-363, the CB(1) receptor agonist Delta(9)-tetrahydrocannabinol, the MCR4 receptor agonist alpha-MSH, or the cannabinoid CB(1) receptor antagonist SR 141716. Food intake and locomotor activity were then recorded for 120 min. When administrated alone, SR 141716 and alpha-MSH dose-dependently attenuated baseline feeding, whereas sub-anorectic doses of SR 141716 and alpha-MSH synergistically attenuated baseline feeding when combined. Delta(9)-Tetrahydrocannabinol-induced feeding was not blocked by alpha-MSH, whereas SR 141716 dose-dependently attenuated JKC-363-induced feeding. Locomotor activity was not significantly affected by any drug treatment, suggesting that the observed effects on feeding were not due to a nonspecific reduction in motivated behavior. These findings revealed a synergistic interaction between the cannabinoid and melanocortin systems in feeding behavior. These results further suggested that CB(1) receptors are located downstream from melanocortin receptors and CB(1) receptor signaling is necessary to prevent the melanocortin system from altering food intake.     

Expression and function of cannabinoid receptors in mouse islets

The endocannabinoid system plays a key role in energy homeostasis, with agonists and antagonists of CB1 receptors acting centrally to stimulate and inhibit food intake, respectively. In addition to their established effects on the central nervous system, cannabinoid receptor agonists also exert peripheral effects by modulating cellular cyclic AMP and calcium levels and there have been reports that they regulate β-cell function. However, the few reports to date on islet expression of cannabinoid receptors and effects of agonists on insulin secretion have failed to reach a consensus. We have therefore investigated cannabinoid receptor expression by mouse islet β-and α-cells and the effects of selective receptor agonists on cyclic AMP and calcium levels, and on dynamic insulin secretory responses. CB1 and CB2 mRNA and protein expression by islets was detected by RT-PCR and western blotting respectively, and cellular location of the receptors was identified by immunohistochemistry with insulin and glucagon antibody co-staining. Cyclic AMP generation was quantified by enzyme immunoassay and changes in calcium levels were measured by microfluorimetry of Fura-2-loaded mouse islet cells. Dynamic insulin secretion was quantified by radioimmunoassay after perifusion of isolated islets. We found that mouse islets expressed both CB1 and CB2 receptors, and they were localised to β-cells. Activation of mouse β-cell CB1 and CB2 receptors resulted in decreased cyclic AMP, increased calcium and potentiation of glucose-stimulated insulin secretion. Thus, activation of islet cannabinoid receptors by locally produced endocannabinoids such as 2-aminoglycerol may be another regulatory pathway by which islets stimulate insulin secretion to maintain glucose homeostasis.     

Evidence for functional CB1 cannabinoid receptor expressed in the rat thyroid

OBJECTIVE: Previous reports have shown that the Delta(9)-tetrahydrocannabinol (Delta(9)TCH), the major psychoactive cannabinoid components of marijuana, is able [corrected] to inhibit thyroid hormonal activity. The aim of this study was to characterize the CB1 functional expression in the rat thyroid by a multi-methods approach. METHODS AND RESULTS: RT-PCR was used to detect the mRNA expression of the CB1 cannabinoid receptor (17.8+/-4.0% of the normalizing reference gene beta(2) microglobulin), as well as the expression of the endocannabinoid hydrolyzing enzyme, fatty acid amide hydrolase (46.9+/-4.3% of beta(2) microglobulin), in the rat thyroid gland. The CB1-encoded protein was detected in its glycosylated form (63 kDa) by Western blot, employing a polyclonal antibody, while CB1 immunohistochemical localization showed an intracellular positive staining in both follicular and parafollicular cells. In addition, a 30% decrease in serum levels of both 3,5,3' tri-iodothyronine (T(3)) and thyroxine (T(4)) was detected 4 h after the administration of the synthetic cannabinoid receptor agonist, WIN 55,212-2 (10 mg/kg i.p.). These effects were antagonized by pretreatment with the CB1 antagonist SR 141716A (3 mg/kg i.p.); thyrotrophin levels were unaffected by both treatments. CONCLUSION: These data indicate that functional CB1 receptors which are able to modulate the release of T(3) and T(4) are expressed in the rat thyroid, and suggest a possible role of cannabinoids in the regulation of rat thyroid hormonal activity.     

The emerging role of the endocannabinoid system in endocrine regulation and energy balance

During the last few years, the endocannabinoid system has emerged as a highly relevant topic in the scientific community. Many different regulatory actions have been attributed to endocannabinoids, and their involvement in several pathophysiological conditions is under intense scrutiny. Cannabinoid receptors, named CB1 receptor and CB2 receptor, first discovered as the molecular targets of the psychotropic component of the plant Cannabis sativa, participate in the physiological modulation of many central and peripheral functions. CB2 receptor is mainly expressed in immune cells, whereas CB1 receptor is the most abundant G protein-coupled receptor expressed in the brain. CB1 receptor is expressed in the hypothalamus and the pituitary gland, and its activation is known to modulate all the endocrine hypothalamic-peripheral endocrine axes. An increasing amount of data highlights the role of the system in the stress response by influencing the hypothalamic-pituitary-adrenal axis and in the control of reproduction by modifying gonadotropin release, fertility, and sexual behavior. The ability of the endocannabinoid system to control appetite, food intake, and energy balance has recently received great attention, particularly in the light of the different modes of action underlying these functions. The endocannabinoid system modulates rewarding properties of food by acting at specific mesolimbic areas in the brain. In the hypothalamus, CB1 receptor and endocannabinoids are integrated components of the networks controlling appetite and food intake. Interestingly, the endocannabinoid system was recently shown to control metabolic functions by acting on peripheral tissues, such as adipocytes, hepatocytes, the gastrointestinal tract, and, possibly, skeletal muscle. The relevance of the system is further strenghtened by the notion that drugs interfering with the activity of the endocannabinoid system are considered as promising candidates for the treatment of various diseases, including obesity.     

The unfolding cannabinoid story on energy homeostasis: central or peripheral site of action?

Presentations in this symposium addressed effects and modes of action of endocannabinoids in various tissues in relation to metabolic disorders. Endocannabinoids are produced and exert their effect in various brain sites, including the mesolimbic reward circuitry and the hypothalamus. Both of these regions have direct ties to energy metabolism regulation, particularly food intake and energy expenditure. These data clearly suggest that the observed beneficial effects of CB1 (cannabinoid receptor 1) receptor antagonists on obesity may be related to the central endocannabinoid system. On the other hand, data presented on cannabinoid action in the liver and white adipose tissues clearly indicate that CB1-mediated events in affecting metabolic phenotype may occur in peripheral tissues as well. This together with the reported results from human trials on CB1 antagonists showing that the initial anorectic effect of rimonabant is diminished after the first weeks while longer lasting weight loss is achieved do indicate that peripheral action of cannabinoids are very important in body weight regulation. Should this hold true in the long run, antagonizing CB1 receptors with compound not crossing the blood-brain barrier could revolutionize pharmaceutical approaches to obesity by offering a tool that short cuts the central nervous system.     

Presence of functional cannabinoid receptors in human endocrine pancreas

Aims/hypothesis We examined the presence of functional cannabinoid receptors 1 and 2 (CB1, CB2) in isolated human islets, phenotyped the cells producing cannabinoid receptors and analysed the actions of selective cannabinoid receptor agonists on insulin, glucagon and somatostatin secretion in vitro. We also described the localisation on islet cells of: (1) the endocannabinoid-producing enzymes N-acyl-phosphatidyl ethanolamine-hydrolysing phospholipase D and diacylglycerol lipase; and (2) the endocannabinoid-degrading enzymes fatty acid amidohydrolase and monoacyl glycerol lipase. Methods Real-time PCR, western blotting and immunocytochemistry were used to analyse the presence of endocannabinoid-related proteins and genes. Static secretion experiments were used to examine the effects of activating CB1 or CB2 on insulin, glucagon and somatostatin secretion and to measure changes in 2-arachidonoylglycerol (2-AG) levels within islets. Analyses were performed in isolated human islets and in paraffin-embedded sections of human pancreas. Results Human islets of Langerhans expressed CB1 and CB2 (also known as CNR1 and CNR2) mRNA and CB1 and CB2 proteins, and also the machinery involved in synthesis and degradation of 2-AG (the most abundant endocannabinoid, levels of which were modulated by glucose). Immunofluorescence revealed that CB1 was densely located in glucagon-secreting alpha cells and less so in insulin-secreting beta cells. CB2 was densely present in somatostatin-secreting delta cells, but absent in alpha and beta cells. In vitro experiments revealed that CB1 stimulation enhanced insulin and glucagon secretion, while CB2 agonism lowered glucose-dependent insulin secretion, showing these cannabinoid receptors to be functional. Conclusions/interpretation Together, these results suggest a role for endogenous endocannabinoid signalling in regulation of endocrine secretion in the human pancreas. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorised users.