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Cannabis and metabolic disorder

Brain

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Cannabis has been known to mankind for more than 4 thousand years and is used for medical and recreational purposes. The first cannabinoid – cannabidiol was discovered at the end of the XIX century, and at the end of the XX century, cannabinoid receptors (CB1 and CB2) were discovered and the concept of the endocannabinoid system was formed. Today, two synthetic drugs - dronabinol and nabinol are authorised for medical use. Marijuana is gradually being legalized in different parts of the world. Researchers learn that endocannabinoids and their receptors are involved in almost every physiological and pathological process. It was this ubiquity of endocannabinoid system, the system that almost put an end to the use of its antagonists in patients with obesity. Endocannabinoid system (ECS) of the human body in a simplified form consists of endocannabinoids, enzymes for their synthesis and decomposition, CB1 and CB2 receptors.

Endocannabinoids are derivatives of polyunsaturated fatty acids that are formed in the cell "on demand" from the cell membrane phospholipids and act autocrine or paracrine on endocannabinoid receptors. The most researched cannabinoids are anandamide (arachidonic acid N-ethanolamide, AEA), arachidonic acid glycerin ether or 2-arachidonoglycerol (2-AG). Anandamide is formed from N-acylphosphatidylethanolamine (NAPE) by N-acetyltransferase and NAPE-PLD. These enzymes are found in the gastrointestinal tract and central nervous system. 2-AG is produced during hydrolysis of diacylglycerol by DAG-lipases alpha and beta. There are other ways of producing anandamide and 2-AG.

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The main receptors of the endocannabinoid system are CB1 and CB2, which are binded not only by endocannabinoids but also by phytocannabinoids (Δ9-tetrahydrocannabiol - the main component of Marijuana and cannabidiol) and synthetic cannabinoids(nabilon). However, cannabinoids act on other receptors:

1.CB1R: they are located in the brain, are responsible for antinociceptive action, cognitive function and memory disorders. These are mainly presynaptic receptors in the following structures of the central nervous system: olfactory bulb, cerebral cortex, hypothalamus, hippocampus, striatum, cerebellum. They are also found in postsynaptic membranes, astrocytes. In much smaller quantities, they are found in the heart muscle, blood vessels, gastrointestinal tract, reproductive organs, muscles, bones, and skin. CB1Rs are associated with Gi and, through the PKA cascade, reduce the release of neurotransmitters and reduce the activity of the MAPK pathway. Some CB1Rs are associated with Ca2+channels and Kir channels, or stimulate NOS.
2.CB2: they are mainly found in cells of the immune system and hematopoietic cells, as well as in peripheral tissues cells: liver, endocrine part of the pancreas, bones, neurons and microglia. One of their functions is suppression of the cytokines release.
3. Capsaicin receptor TRPV1: it is carried by primary afferents and perivascular neurons. Effects: local vasodilation, pro-inflammatory effect, cardioprotective and antihypertensive effect. Regulates the release of substance P and gene-calcitonin peptide (CGRP).
4. PPARs, G-protein-coupled receptor 55 (GPR55), nicotine receptors, 5-HT3 and A2A adenosine receptors.

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Endocannabinoids act allosterically on 5-HT2 receptors, 5-HT3 receptors, α1-adrenergic receptors, M1 and M4 muscarinic receptors and AMPA GLUA1 and GLUA3 glutamate receptors. Binding to the receptors above mediates the effects of endocannabinoids: analgesic; antispasmodic; immunosuppressive; anti-inflammatory; anti-allergic; sedative; normotimic; orexigenic; antiemetic; reduction of intraocular pressure; bronchodilation; neuroprotective; antitumor; antioxidant; tachycardia and dry mouth. The degradation of anandamide and 2-AG occurs through the reuptake of endocannabinoids by the cell and their hydrolysis by enzymes: anandamide - hydrolase of fatty acid amides, 2-AG-monoacylglycerol-lipase. 2-AG can also be oxidized by cyclooxygenase-2 to form biologically active glycerol esters of prostaglandins.

Hyperactivation of the endocannabinoid system may be a link between obesity and related diseases. Hyperactivation of the ECS is found both in the hypothalamus and in peripheral tissues, including the liver and adipose tissue. In the central nervous system, endocannabinoids perform the function of retrograde neuromodulators, which involves inhibition of the excitatory and inhibitory neurotransmitters release of through presynaptic CB1 receptors. Thus, they modulate neuronal activity, including in the parts of the brain responsible for regulating the energy balance: the hypothalamus, the brain stem, the cortico-limbic system — the nucleus accumbens (NAc) and the ventral tegmental area (VTA).

It has been shown that the orexigenic or anorexigenic effect of endocannabinoids depends on the properties of the neuron on which the presynaptic CB1 receptors are located. However, the orexigenic effect of CB1 receptor agonists on the body as a whole indicates a predominant inhibition of glutamatergic synapses. Endocannabinoids inform about instantaneous changes in the energy balance, since they are synthesized "on demand". Their concentration in the brain structures increases during fasting and decreases when the need for food is satisfied. Direct injection of AEA and 2-AG in hypothalamus or NAc of rats increases the consumption of food and sucrose solution through the CB1R-dependent mechanism. Also, the cannabinoid system regulates appetite along the lectin pathway in the hypothalamus. Leptin reduces food intake by increasing the release of appetite-reducing neuropeptides and suppressing the release of factors that stimulate hunger. A decrease in leptin levels coincides with an increase in endocannabinoid levels in the hypothalamus. Leptin suppresses the synthesis of endocannabinoids, reducing intracellular calcium, and suppresses the CB 1-dependent activation of neurons expressing melanin-concentrating hormone in the lateral hypothalamus. However, the effect of leptin is manifested only when the ECS is activated, otherwise (when the CB1 receptor gene is knocked out) leptin doesn`t decrease appetite in mice.

There is an antagonism between leptin and glucocorticoids in regard to regulation of endocannabinoid synthesis in the paraventricular nucleus (PVN). Glucocorticoids trigger endocannabinoid-mediated rapid inhibition of synaptic excitation in the PVN through the membrane receptor, which allows for the secretion of hypothalamic hormones to reduce quickly. Leptin blocks the synthesis of endocannabinoids, which is triggered by glucocorticoids. ECS and ghrelin regulate the energy balance together. The action of ghrelin requires the appearance of AMPK in the PVN, which is caused by the activation of CB1 receptors. AEA stimulates ghrelin synthesis and secretion in rat’s stomach. In people with normal weight eating for pleasure is associated with increased levels of ghrelin and 2-AG.
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Cannabinoids enhance the feeling of pleasure from eating by increasing the release of dopamine in the NAc. It is likely, that activation of dopaminergic neurons of the VTA is mediated by the action of endocannabinoids on CB1 receptors at glutamatergic terminals, which inhibit GABAergic neurons expressed from the NAc to the VTA, and thereby disinhibit dopaminergic neurons in the VTA. Taste sensations are processed in the parabrachial nucleus (PBN) and the nucleus tractus solitarii (NTS), where they are integrated with signals from the gastrointestinal tract. The processed information determines the amount of food consumed and the intervals between meals. By stimulating CB1 receptors in PBN, endocannabinoids increase the consumption of palatable food.

An increase in food consumption is achieved by increase in concentration of endocannabinoids, activation of CB1 receptors at the axon terminals of the olfactory cortex and inhibition of granular cells in the olfactory bulb, which increases sensitivity to pleasant smells. Endocannabinoid receptors colocalize with sweet receptors on the tongue papillae and enhance the pleasure of sweet food. There is no evidence that the effect of endocannabinoids on taste and smell plays a role in the pathogenesis of obesity. Up-regulation of CB1 receptors is also observed in obesity pathogenesis. Interestingly, CB1 receptor knockout mice are resistant to alimentary obesity. They have increased activity of the sympathetic nervous system, increased lipid oxidation and thermogenesis; increased levels of endocannabinoids in plasma and saliva. Plasma levels of endocannabinoids have been shown to be elevated in patients with obesity and type 2 diabetes and correlate with the degree of insulin resistance, body mass index, waist circumference and visceral fat mass. It is proposed to use these values as markers of the fat white distribution and insulin resistance for prediction of response to treatment. However, clinical application is still far away: methods for isolating and measuring the endocannabinoids' concentration have not been standardized; reference levels and the correlation of age, gender and present diseases with their values have not been established.

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Hyperactivation of cannabinoid system is reflected in change of energy metabolism in various organs:
1. Activation of CB1 receptors in isolated mouse adipocytes leads to stimulation of fatty acid synthase and lipoprotein lipase and inhibition of AMPK. The expression of adipocyte differentiation (PPAR) genes increases, mitochondrial biogenesis is disrupted;

2. Activation of CB1 receptors in hepatocytes leads to a decrease in AMPK phosphorylation and its activity. The expression of acetyl-CoA-carboxylase-1 (ACC1) and fatty acid synthase (FAS) increases, de novo fatty acid synthesis increases and liver steatosis develops. Up-regulation of inhibitory phosphorylation of the insulin receptor substrate (IRS) and inhibitory dephosphorylation of insulin-activated protein kinase B (PKB) occurs, followed by triggering endoplasmic reticulum stress. It has been shown that the CB2 receptor is involved in the pathogenesis of liver steatosis;

3. Activation of CB1 receptors in skeletal muscle suppresses glucose and fatty acid oxidation and mitochondrial biogenesis, reduces basal and insulin-dependent glucose transport, reduces tissue sensitivity to insulin by PI3-kinase/PKB and Raf-MEK1/2-ERK1/2 pathways, which can lead to insulin resistance;

4. Activation of CB1R on beta cells of the pancreas recruits focal adhesion kinases (FAK). Its action causes the rebuilding of cytoskeleton; exocytosis of vesicles with insulin occurs, triggers apoptosis of beta cells and promotes infiltration of islets by macrophages and inflammation, leading to type 2 diabetes.​

Treatment of metabolic disorder and obesity by decreasing the tone of cannabinoid system.
To reduce the activity of ECS in obese patients, the antagonists of ECS and lifestyle modifications are proposed:
1. Nonselective CB1 receptor blockers;
2. Selective blockers of peripheral CB1 receptors («Compound 2p», «Compound 10q»);
3. Allosteric antagonists of CB1 receptors (hemopressin, pregnenolone, ORG27569 and PSNCBAM-1)
4. Neutral agonists (AM4113, AM6545, JD5037, TM38837, NESS06SM);
5. CB2 receptor agonists (JWH-133, JWH-015);
6. Nonselective agonists of CB1 and CB2 receptors (URB447);
7. Modulators of other receptors (TRPV1, GPR55);
8. Inhibitors of enzymes involved in synthesis of endocannabinoids;
9. A diet with a high level of omega-3 and omega-6 fatty acids.

The first CB1R blocker approved by clinical trials used for obesity treatment was rimonabant (SR141716A). In Europe, it had been sold since 2006 under the name Acomplia. It is often called a CB1R antagonist, but in fact it is an inverse agonist. Data from multinational clinical trials of rimonabant in obese patients (Rimonabant in Obesity, RIO), namely RIO-Lipids, RIO-Europe, RIONorth America and RIO-Diabetes indicate the effectiveness of rimonabant in weight loss and reduction of cardiovascular risk factors. The latter is due to the normalization of adiponectin, HDL, triglycerides and HbA1c levels in diabetic patients.

Long-term rimonabant treatment restored of the sensitivity of cells to insulin, normalized the size of fat cells and their distribution in the body, prevented the deposition of visceral fat and reduced the amount of subcutaneous fat, reduced body weight regardless of decrease in food intake. The mechanisms of the observed effects are not yet clear. One of them may be an increase in the adiponectin gene expression in visceral fat and the concentration of adiponectin in plasma during treatment with rimonabant. There is an increase in the activity of adiponectin 1 and 2 receptors in the liver. The hepatoprotective effect of rimonabant is also manifested as an increase in fat oxidation in the liver and a decrease in inflammation, which reduces the accumulation of fat in the liver.

Blockade of CB1 receptors expressed in beta cells of pancreatic islets stimulated their proliferation and increased cell size, reduced the inflammatory response and led to normalization of glucose levels and restoration of insulin sensitivity. Pharmacological blockade of CB1 is effective only with hyperactivity of the ECS and hypersecretion of insulin. Blockade of CB1 receptors in white adipocytes in vitro stimulates mitochondrial biogenesis through increased expression of endothelial NOS, reduces fatty acid synthesis and triglyceride accumulation, and induces white-to-brown fat transdifferentiation, characterized by increased expression of uncoupling protein-1 (UCP-1), alpha-coactivator PPAR-gamma (PGC-1) and AMPK activity.

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Blockade of CB1 receptors in brown adipocytes intensifies disruption in tissue respiration. However, in vivo it has been shown that ECS regulates lipogenesis and lipolysis in white adipose tissue at the level of the sympathetic nervous system, not at the tissue level. The hypophagic effect of rimonabant, achieved within an hour, depends on the activity of the sympathetic nervous system and disappears when beta-blockers are administered. At the same time, neurological and psychiatric side effects also disappear —fear, anxiety. Acomplia was withdrawn from European markets in 2008 because it was associated with suicidal behavior, depression, seizures and caused five deaths in the UK. Clinical studies of other CB1 receptor antagonists (taranabant, surinabant, ibipinabant) were discontinued at phase 2-3 in 2008-2012.

The focus of research has shifted towards peripheral CB1R blockers, allosteric inhibitors, neutral agonists, inhibitors of endocannabinoid synthesis, stimulators of their degradation, modulators of other receptors and dietary restrictions. None of the possible drugs have yet been tested on humans, although all of them have shown some effectiveness in animal models of obesity. A high-fat diet increases the content of anandamide in the liver of mice, whereas a similar diet with a high content of omega-3 fatty acids (contained in fish oil) reduces the content of 2-AG in the brain of piglets. In rats consuming large amounts of linoleic acid (the "western diet"), the content of 2-AG and anandamide in the small intestine is increased. However, in clinical studies, the same calorie amount of low- and high-fat diets did not lead to a change in plasma concentrations of endocannabinoids. A diet enriched with polyunsaturated fatty acids did not lead to weight loss in obese patients, but improved the lipid profile in patients with hypercholesterolemia.​
 
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