Enhancement of endocannabinoid signaling by fatty acid amide hydrolase inhibition: A neuroprotective therapeutic modality

AimsThis review posits that fatty acid amide hydrolase (FAAH) inhibition has therapeutic potential against neuropathological states including traumatic brain injury; Alzheimer's, Huntington's, and Parkinson's diseases; and stroke.Main methodsThis proposition is supported by data from numerous in vitro and in vivo experiments establishing metabolic and pharmacological contexts for the neuroprotective role of the endogenous cannabinoid (“endocannabinoid”) system and selective FAAH inhibitors.Key findingsThe systems biology of endocannabinoid signaling involves two main cannabinoid receptors, the principal endocannabinoid lipid mediators N-arachidonoylethanolamine (“anandamide”) (AEA) and 2-arachidonoyl glycerol (2-AG), related metabolites, and the proteins involved in endocannabinoid biosynthesis, biotransformation, and transit. The endocannabinoid system is capable of activating distinct signaling pathways on-demand in response to pathogenic events or stimuli, thereby enhancing cell survival and promoting tissue repair. Accumulating data suggest that endocannabinoid system modulation at discrete targets is a promising pharmacotherapeutic strategy for treating various medical conditions. In particular, neuronal injury activates cannabinoid signaling in the central nervous system as an intrinsic neuroprotective response. Indirect potentiation of this salutary response through pharmacological inhibition of FAAH, an endocannabinoid-deactivating enzyme, and consequent activation of signaling pathways downstream from cannabinoid receptors have been shown to promote neuronal maintenance and function.SignificanceThis therapeutic modality has the potential to offer site- and event-specific neuroprotection under conditions where endocannabinoids are being produced as part of a physiological protective mechanism. In contrast, direct application of cannabinoid receptor agonists to the central nervous system may activate CB receptors indiscriminately and invite unwanted psychotrophic effects.     

Endocannabinoids Are Expressed in Bone Marrow Stromal Niches and Play a Role in Interactions of Hematopoietic Stem and Progenitor Cells with the Bone Marrow Microenvironment

Endocannabinoids are lipid signaling molecules that act via G-coupled receptors, CB₁ and CB₂. The endocannabinoid system is capable of activation of distinct signaling pathways on demand in response to pathogenic events or stimuli, hereby enhancing cell survival and promoting tissue repair. However, the role of endocannabinoids in hematopoietic stem and progenitor cells (HSPCs) and their interaction with hematopoietic stem cells (HSC) niches is not known. HSPCs are maintained in the quiescent state in bone marrow (BM) niches by intrinsic and extrinsic signaling. We report that HSPCs express the CB₁ receptors and that BM stromal cells secrete endocannabinoids, anandamide (AEA) (35 pg/10⁷ cells), and 2-AG (75.2 ng/10⁷ cells). In response to the endotoxin lipopolysaccharide (LPS), elevated levels of AEA (75.6 pg/10⁷ cells) and 2-AG (98.8 ng/10⁷ cells) were secreted from BM stromal cells, resulting in migration and trafficking of HSPCs from the BM niches to the peripheral blood. Furthermore, administration of exogenous cannabinoid CB₁ agonists in vivo induced chemotaxis, migration, and mobilization of human and murine HSPCs. Cannabinoid receptor knock-out mice Cnr1^−/− showed a decrease in side population (SP) cells, whereas fatty acid amide hydrolase (FAAH)^−/− mice, which have elevated levels of AEA, yielded increased colony formation as compared with WT mice. In addition, G-CSF-induced mobilization in vivo was modulated by endocannabinoids and was inhibited by specific cannabinoid antagonists as well as impaired in cannabinoid receptor knock-out mice Cnr1^−/−, as compared with WT mice. Thus, we propose a novel function of the endocannabinoid system, as a regulator of HSPC interactions with their BM niches, where endocannabinoids are expressed in HSC niches and under stress conditions, endocannabinoid expression levels are enhanced to induce HSPC migration for proper hematopoiesis.     

Energy metabolism of cerebral mitochondria during aging,ischemia and post-ischemic recovery assessed by functional proteomics of enzymes

Stroke is a leading cause of death and disability, but most of the therapeutic approaches failed in clinical trials. The energy metabolism alterations, due to marked ATP decline, are strongly related to stroke and, at present, their physiopathological roles are not fully understood. Thus, the aim of this study was to evaluate the effects of aging on ischemia-induced changes in energy mitochondrial transduction and the consequences on overall brain energy metabolism in an in vivo experimental model of complete cerebral ischemia of 15 min duration and during post-ischemic recirculation after 1, 24, 48, 72 and 96 h, in 1 year “adult” and 2 year-old “aged” rats.     

The combination of Panax ginseng and Angelica sinensis alleviates ischemia brain injury by suppressing NLRP3 inflammasome activation and microglial pyroptosis

BackgroundThe combination of Panax ginseng and Angelica sinensis (CPA) has been used to treat stroke for one thousand years and demonstrated clinically to have satisfied effects. However, the underlying mechanism remains unknown.PurposeWe investigate whether CPA has neuroprotective effects via suppressing Nod-like receptor protein 3 (NLRP3) inflammasome and microglial pyroptosis against ischemic injury in transient middle cerebral artery occlusion (MCAO) rats.MethodsMale rats were divided randomly into sham operated, MCAO, MCC950 (NLRP3-specific inhibitor) and CPA groups. Neurological deficits, glucose uptake, infarct size, activation of NLRP3 inflammasomes, microglial pyroptosis and related signaling pathways were detected. BV-2 microglial cells subjected to oxygen-glucose deprivation/reoxygenation (OGD/R) were used in in vitro experiments.ResultsCompared with sham rats, elevated level of proinflammatory interleukin-1β (IL-1β) in plasma, neurological function deficit, reduced glucose uptake in ipsilateral hemisphere, obvious infarct size, the activation of NLRP3 inflammasomes and enhanced microglial pyroptosis were presented in MCAO rats. The administrations of MCC950 and CPA respectively reversed the results. In vitro OGD/R induced the release of lactate dehydrogenase, promoted NLRP3 inflammasomes activation and pyroptosis in BV-2 cells, which was significantly suppressed by treatment with ginsenoside Rd (Rd) and Z-ligustilide (LIG). Mechanistically, OGD/R induced high expression of dynamin-related protein 1 (Drp1) and mitochondrial fission, as well as NLRP3 inflammasomes activation and pyroptosis in BV-2 cells, which was attenuated by treatment with Rd and LIG. Moreover, the increased expression of Drp1 was validated in MCAO rats, and also abolished by MCC950 or CPA treatments.ConclusionCPA treatment attenuates cerebral injury via inhibition of NLRP3 inflammasomes activation and microglial pyroptosis after stroke, which at least partially involved in the amelioration of Drp1-mediated mitochondrial fission.     

2-arachidonoylglycerol induces the migration of HL-60 cells differentiated into macrophage-like cells and human peripheral blood monocytes through the cannabinoid CB2 receptor-dependent mechanism

2-Arachidonoylglycerol is an endogenous ligand for the cannabinoid receptors (CB1 and CB2) and has been shown to exhibit a variety of cannabimimetic activities in vitro and in vivo. Recently, we proposed that 2-arachidonoylglycerol is the true endogenous ligand for the cannabinoid receptors, and both receptors (CB1 and CB2) are primarily 2-arachidonoylglycerol receptors. The CB1 receptor is assumed to be involved in the attenuation of neurotransmission. On the other hand, the physiological roles of the CB2 receptor, which is abundantly expressed in several types of leukocytes such as macrophages, still remain unknown. In this study, we examined the effects of 2-arachidonoylglycerol on the motility of HL-60 cells differentiated into macrophage-like cells. We found that 2-arachidonoylglycerol induces the migration of differentiated HL-60 cells. The migration induced by 2-arachidonoylglycerol was blocked by treatment of the cells with either SR144528, a CB2 receptor antagonist, or pertussis toxin, suggesting that the CB2 receptor and Gi/Go are involved in the 2-arachidonoylglycerol-induced migration. Several intracellular signaling molecules such as Rho kinase and mitogen-activated protein kinases were also suggested to be involved. In contrast to 2-arachidonoylglycerol, anandamide, another endogenous cannabinoid receptor ligand, failed to induce the migration. The 2-arachidonoylglycerol-induced migration was also observed for two other types of macrophage-like cells, the U937 cells and THP-1 cells, as well as human peripheral blood monocytes. These results strongly suggest that 2-arachidonoylglycerol induces the migration of several types of leukocytes such as macrophages/monocytes through a CB2 receptor-dependent mechanism thereby stimulating inflammatory reactions and immune responses.     

Attenuated inflammatory response in aged mice brains following stroke

Background

Increased age is a major risk factor for stroke incidence, post-ischemic mortality, and severe and long-term disability. Stroke outcome is considerably influenced by post-ischemic mechanisms. We hypothesized that the inflammatory response following an ischemic injury is altered in aged organisms.

Methods and Results

To that end, we analyzed the expression pattern of pro-inflammatory cytokines (TNF, IL-1α, IL-1β, IL-6), anti-inflammatory cytokines (IL-10, TGFβ1), and chemokines (Mip-1α, MCP-1, RANTES) of adult (2 months) and aged (24 months) mice brains at different reperfusion times (6 h, 12 h, 24 h, 2 d, 7 d) following transient occlusion of the middle cerebral artery. The infarct size was assessed to monitor possible consequences of an altered inflammatory response in aged mice. Our data revealed an increased neuro-inflammation with age. Above all, we found profound age-related alterations in the reaction to stroke. The response of pro-inflammatory cytokines (TNF, and IL-1β) and the level of chemokines (Mip-1α, and MCP-1) were strongly diminished in the aged post-ischemic brain tissue. IL-6 showed the strongest age-dependent decrease in its post-ischemic expression profile. Anti-inflammatory cytokines (TGFβ1, and IL-10) revealed no significant age dependency after ischemia. Aged mice brains tend to develop smaller infarcts.

Conclusion

The attenuated inflammatory response to stroke in aged animals may contribute to their smaller infarcts. The results presented here highlight the importance of using aged animals to investigate age-associated diseases like stroke, and should be considered as a major prerequisite in the development of age-adjusted therapeutic interventions.     

TIPE2, a Novel Regulator of Immunity,Protects against Experimental Stroke

The inflammatory responses accompanying stroke are recognized to contribute to secondary ischemic injury. TIPE2 is a very recently identified negative regulator of inflammation that maintains immune homeostasis. However, it is unknown whether TIPE2 is expressed in the brain and contributes to the regulation of cerebral diseases. In this study, we explored the potential roles of TIPE2 in cerebral ischemia/reperfusion injury. TIPE2^−/− mice were used to assess whether TIPE2 provides neuroprotection following cerebral ischemia/reperfusion induced by middle cerebral artery occlusion (MCAO), and in vitro primary cerebral cell cultures were used to investigate the expression and regulation of TIPE2. Our results show that genetic ablation of the Tipe2 gene significantly increased the cerebral volume of infarction and neurological dysfunction in mice subjected to MCAO. Flow cytometric analysis revealed more infiltrating macrophages, neutrophils, and lymphocytes in the ischemic hemisphere of TIPE2^−/− mice. The responses to inflammatory cytokines and chemokines were significantly increased in TIPE2^−/− mouse brain after MCAO. We further observed that TIPE2 was highly induced in WT mice after cerebral ischemia and was expressed mainly in microglia/macrophages, but not in neurons and astrocytes. Finally, we found that regulation of TIPE2 expression was associated with NADPH oxidase activity. These findings demonstrate, for the first time, that TIPE2 is involved in the pathogenesis of stroke and suggest that TIPE2 plays an essential role in a signal transduction pathway that links the inflammatory immune response to specific conditions after cerebral ischemia. Targeting TIPE2 may be a new therapeutic strategy for stroke treatment.     

Cannabinoids reduce markers of inflammation and fibrosis in pancreatic stellate cells

Background

While cannabinoids have been shown to ameliorate liver fibrosis, their effects in chronic pancreatitis and on pancreatic stellate cells (PSC) are unknown.

Methodology/Principal Findings

The activity of the endocannabinoid system was evaluated in human chronic pancreatitis (CP) tissues. In vitro, effects of blockade and activation of cannabinoid receptors on pancreatic stellate cells were characterized. In CP, cannabinoid receptors were detected predominantly in areas with inflammatory changes, stellate cells and nerves. Levels of endocannabinoids were decreased compared with normal pancreas. Cannabinoid-receptor-1 antagonism effectuated a small PSC phenotype and a trend toward increased invasiveness. Activation of cannabinoid receptors, however, induced de-activation of PSC and dose-dependently inhibited growth and decreased IL-6 and MCP-1 secretion as well as fibronectin, collagen1 and alphaSMA levels. De-activation of PSC was partially reversible using a combination of cannabinoid-receptor-1 and -2 antagonists. Concomitantly, cannabinoid receptor activation specifically decreased invasiveness of PSC, MMP-2 secretion and led to changes in PSC phenotype accompanied by a reduction of intracellular stress fibres.

Conclusions/Significance

Augmentation of the endocannabinoid system via exogenously administered cannabinoid receptor agonists specifically induces a functionally and metabolically quiescent pancreatic stellate cell phenotype and may thus constitute an option to treat inflammation and fibrosis in chronic pancreatitis.     

Synthesis and biological evaluation of bivalent cannabinoid receptor ligands based on hCB2R selective benzimidazoles reveal unexpected intrinsic properties

The design of bivalent ligands targeting G protein-coupled receptors (GPCRs) often leads to the development of new, highly selective and potent compounds. To date, no bivalent ligands for the human cannabinoid receptor type 2 (hCB₂R) of the endocannabinoid system (ECS) are described. Therefore, two sets of homobivalent ligands containing as parent structure the hCB₂R selective agonist 13a and coupled at different attachment positions were synthesized. Changes of the parent structure at these positions have a crucial effect on the potency and efficacy of the ligands. However, we discovered that bivalency has an influence on the effect at both cannabinoid receptors. Moreover, we found out that the spacer length and the attachment position altered the efficacy of the bivalent ligands at the receptors by turning agonists into antagonists and inverse agonists.     

Cannabinoid-mediated modulation of neuropathic pain and microglial accumulation in a model of murine type I diabetic peripheral neuropathic pain

Background

Despite the frequency of diabetes mellitus and its relationship to diabetic peripheral neuropathy (DPN) and neuropathic pain (NeP), our understanding of underlying mechanisms leading to chronic pain in diabetes remains poor. Recent evidence has demonstated a prominent role of microglial cells in neuropathic pain states. One potential therapeutic option gaining clinical acceptance is the cannabinoids, for which cannabinoid receptors (CB) are expressed on neurons and microglia. We studied the accumulation and activation of spinal and thalamic microglia in streptozotocin (STZ)-diabetic CD1 mice and the impact of cannabinoid receptor agonism/antagonism during the development of a chronic NeP state. We provided either intranasal or intraperitoneal cannabinoid agonists/antagonists at multiple doses both at the initiation of diabetes as well as after establishment of diabetes and its related NeP state.

Results

Tactile allodynia and thermal hypersensitivity were observed over 8 months in diabetic mice without intervention. Microglial density increases were seen in the dorsal spinal cord and in thalamic nuclei and were accompanied by elevation of phosphorylated p38 MAPK, a marker of microglial activation. When initiated coincidentally with diabetes, moderate-high doses of intranasal cannabidiol (cannaboid receptor 2 agonist) and intraperitoneal cannabidiol attenuated the development of an NeP state, even after their discontinuation and without modification of the diabetic state. Cannabidiol was also associated with restriction in elevation of microglial density in the dorsal spinal cord and elevation in phosphorylated p38 MAPK. When initiated in an established DPN NeP state, both CB1 and CB2 agonists demonstrated an antinociceptive effect until their discontinuation. There were no pronociceptive effects demonstated for either CB1 or CB2 antagonists.

Conclusions

The prevention of microglial accumulation and activation in the dorsal spinal cord was associated with limited development of a neuropathic pain state. Cannabinoids demonstrated antinociceptive effects in this mouse model of DPN. These results suggest that such interventions may also benefit humans with DPN, and their early introduction may also modify the development of the NeP state.     

Endocannabinoids and liver disease. III. Endocannabinoid effects on immune cells: implications for inflammatory liver diseases

Recent studies have implicated dysregulation of the endocannabinoid system in various liver diseases and their complications (e.g., hepatitis, fibrosis, cirrhosis, cirrhotic cardiomyopathy, and ischemia-reperfusion), and demonstrated that its modulation by either cannabinoid 2 (CB(2)) receptor agonists or CB(1) antagonists may be of significant therapeutic benefits. This review is aimed to focus on the triggers and sources of endocannabinoids during liver inflammation and on the novel role of CB(2) receptors in the interplay between the activated endothelium and various inflammatory cells (leukocytes, lymphocytes, etc.), which play pivotal role in the early development and progression of inflammatory and other liver diseases.     

Cannabinoid receptors in atherosclerosis

PURPOSE OF REVIEW: Recent findings suggesting that cannabinoid receptors are potential targets for the treatment of atherosclerosis are reviewed. RECENT FINDINGS: Cannabinoids, such as Delta9-tetrahydrocannabinol, the major psychoactive compound of marijuana, their synthetic analogs and endogenous cannabinoid ligands, produce their biological effects by interacting with specific receptors. In the apolipoprotein E knockout mouse model of atherosclerosis, Delta9-tetrahydrocannabinol was shown to inhibit disease progression through pleiotropic effects on inflammatory cells. Blocking of cannabinoid receptor CB2, the main cannabinoid receptor expressed on immune cells, abolished the observed effects. The development of novel cannabinoid receptor ligands that selectively target CB2 receptors or pharmacological modulation of the endocannabinoid system might offer novel therapeutic strategies in the treatment of atherosclerosis. Several reports demonstrating an implication of the endocannabinoid system in different inflammatory conditions support this hypothesis. SUMMARY: The immunomodulatory capacity of cannabinoids is now well established and suggests a broad therapeutic potential of cannabinoids for a variety of conditions, including atherosclerosis. New strategies based on nonpsychotropic cannabinoid receptor ligands or compounds modulating endocannabinoid synthesis or stability might solve the problem of the unwanted side effects associated with cannabinoid administration.     

The cannabinoid WIN55,212-2 protects against oxidized LDL-induced inflammatory response in murine macrophages

The endocannabinoid system has recently been attracted interest for its anti-inflammatory and anti-oxidative properties. In this study, we investigated the role of the endocannabinoid system in regulating the oxidized low-density lipoprotein (oxLDL)-induced inflammatory response in macrophages. RAW264.7 mouse macrophages and peritoneal macrophages isolated from Sprague-Dawley (SD) rats were exposed to oxLDL with or without the synthetic cannabinoid WIN55,212-2. To assess the inflammatory response, reactive oxygen species (ROS) and tumor necrosis factor alpha (TNF- α) levels were determined, and activation of the mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-kappa B signaling pathways were assessed. We observed that: i) oxLDL strongly induced ROS generation and TNF- α secretion in murine macrophages; ii) oxLDL-induced TNF- α and ROS levels could be lowered considerably by WIN55,212-2 via inhibition of MAPK (ERK1/2) signaling and NF-kappa B activity; and iii) the effects of WIN55212-2 were attenuated by the selective CB2 receptor antagonist AM630. These results demonstrate the involvement of the endocannabinoid system in regulating the oxLDL-induced inflammatory response in macrophages, and indicate that the CB2 receptor may offer a novel pharmaceutical target for treating atherosclerosis.     

Occurrence and possible biological role of the endocannabinoid system in the sea squirt Ciona intestinalis

A cannabinoid receptor orthologue (CiCBR) has been described in the sea squirt Ciona intestinalis. Here we report that CiCBR mRNA expression is highest in cerebral ganglion, branchial pharynx, heart and testis of C. intestinalis, and that this organism also contains cannabinoid receptor ligands and some of the enzymes for ligand biosynthesis and inactivation. Using liquid chromatography-mass spectrometry, the endocannabinoid anandamide was found in all tissues analysed (0.063-5.423 pmol/mg of lipid extract), with the highest concentrations being found in brain and heart. The endocannabinoid 2-arachidonoylglycerol (2-AG) was fivefold more abundant than anandamide, and was most abundant in stomach and intestine and least abundant in heart and ovaries (2.677-50.607 pmol/mg of lipid extract). Using phylogenomic analysis, we identified orthologues of several endocannabinoid synthesizing and degrading enzymes. In particular, we identified and partly sequenced a fatty acid amide hydrolase (FAAH) orthologue, showing 44% identity with human FAAH and containing nearly all the amino acids necessary for a functional FAAH enzyme. Ciona intestinalis also contained specific binding sites for cannabinoid receptor ligands, and an amidase enzyme with pH-dependency and subcellular/tissue distribution similar to mammalian FAAHs. Finally, a typical C. intestinalis behavioural response, siphon reopening after closure induced by mechanical stimulation, was inhibited by the cannabinoid receptor agonist HU-210, and this effect was significantly attenuated by mammalian cannabinoid receptor antagonists.     

The Synergistic Combination of Everolimus and Paroxetine Exerts Post-ischemic Neuroprotection In Vitro

Ischemic stroke is a debilitating multi-factorial cerebrovascular disorder, representing an area of tremendous unmet medical need. Combination treatment has been proposed as a promising therapeutic approach towards combating ischemic stroke. The present study employs in vitro oxygen glucose deprivation (OGD) model to evaluate the post-ischemic neuroprotective efficacy of Everolimus and Paroxetine, alone and in combination. Post-OGD treatment with Everolimus and Paroxetine, alone or in combination, significantly improved the cell survival (~?80%) when compared to the cells subjected to ischemic injury alone. The individual neuroprotective doses of Everolimus and Paroxetine were found to be at 6.25 and 25 nM, respectively. Whereas, the synergistic neuroprotective dose for Everolimus:Paroxetine was 2:10 nM, calculated using the Chou-Talalay combination index and other four mathematical models. The synergistic combination dose downregulated neuroinflammatory genes (Tnf-α, Il1b, Nf-κB, and iNos) and upregulated the neuroprotective genes (Bcl-2, Bcl-xl, Hif-1, and Epo). The mitochondrial functioning and ROS neutralizing ability increased with combination treatment. Further, the active role of nitric oxide synthase and calmodulin were revealed while exploring the bio-activity of Everolimus and Paroxetine through network pharmacology. The present study for the first time demonstrates the synergistic post-ischemic neuroprotective efficacy of combination treatment with Everolimus and Paroxetine in vitro. Taken together, these findings clearly suggest that Everolimus in combination with Paroxetine may represent a promising therapeutic strategy for the treatment of ischemic stroke, further supporting the combination treatment strategy for this debilitating disorder.     

Anandamide increases swelling and reduces calcium sensitivity of mitochondria

The endocannabinoid anandamide alters mitochondria-dependent signal transduction, thus controlling key cellular events like energy homeostasis and induction of apoptosis. Here, the ability of anandamide to directly affect the integrity of mitochondria was investigated on isolated organelles. We found that anandamide dose-dependently increases mitochondrial swelling, and reduces cytochrome c release induced by calcium ions. The effects of anandamide were independent of its target receptors (e.g., cannabinoid or vanilloid receptors), and were paralleled by decreased membrane potential and increased membrane fluidity. Overall, our data suggest that anandamide can impact mitochondrial physiology, by reducing calcium sensitivity and perturbing membrane properties of these organelles.     

Formation of B and T cell subsets require the cannabinoid receptor CB2

A recent and surprising body of research has linked changes in immune function to biologic and therapeutic targeting of cannabinoid receptors, which prototypically respond to delta-9 tetrahydrocannabinol. The peripheral cannabinoid receptor CB2 is highly expressed in immune cell types (macrophages, dendritic cells, and B cells), and pharmacologically alters their cytokine production and responsiveness. Accordingly, cannabinoid agonists can powerfully alter susceptibility to certain microbial infections, atherosclerosis, and cancer immunotherapy. What is unknown is the physiologic role of natural levels of endocannabinoids and their receptors in normal immune homeostasis. Gαi2−/− mice are deficient in the formation of certain B and T cell subsets and are susceptible to immune dysregulation, notably developing inflammatory bowel disease. A key issue is the identity of the Gi-coupled receptors relevant to this Gαi2-signaling pathway. We find that mice deficient in CB2, the Gi-coupled peripheral endocannabinoid receptor, have profound deficiencies in splenic marginal zone, peritoneal B1a cells, splenic memory CD4⁺ T cells, and intestinal natural killer cells and natural killer T cells. These findings partially phenocopy and extend the lymphocyte developmental disorder associated with the Gαi2−/− genotype, and suggest that the endocannabinoid system is required for the formation of T and B cell subsets involved in immune homeostasis. This noncompensatable requirement for physiologic function of the endocannabinoid system is novel. Because levels of endocannabinoids are highly restricted microanatomically, local regulation of their production and receptor expression offers a new principle for regional immune homeostasis and disease susceptibility, and extends and refines the rationale for CB2-targeted immunotherapy in immune and inflammatory diseases.     

Role of Endocannabinoids and Cannabinoid-1 Receptors in Cerebrocortical Blood Flow Regulation

Background

Endocannabinoids are among the most intensively studied lipid mediators of cardiovascular functions. In the present study the effects of decreased and increased activity of the endocannabinoid system (achieved by cannabinoid-1 (CB1) receptor blockade and inhibition of cannabinoid reuptake, respectively) on the systemic and cerebral circulation were analyzed under steady-state physiological conditions and during hypoxia and hypercapnia (H/H).

Methodology/Principal Findings

In anesthetized spontaneously ventilating rats the CB1-receptor antagonist/inverse agonist AM-251 (10 mg/kg, i.v.) failed to influence blood pressure (BP), cerebrocortical blood flow (CoBF, measured by laser-Doppler flowmetry) or arterial blood gas levels. In contrast, the putative cannabinoid reuptake inhibitor AM-404 (10 mg/kg, i.v.) induced triphasic responses, some of which could be blocked by AM-251. Hypertension during phase I was resistant to AM-251, whereas the concomitant CoBF-increase was attenuated. In contrast, hypotension during phase III was sensitive to AM-251, whereas the concomitant CoBF-decrease was not. Therefore, CoBF autoregulation appeared to shift towards higher BP levels after CB1-blockade. During phase II H/H developed due to respiratory depression, which could be inhibited by AM-251. Interestingly, however, the concomitant rise in CoBF remained unchanged after AM-251, indicating that CB1-blockade potentially enhanced the reactivity of the CoBF to H/H. In accordance with this hypothesis, AM-251 induced a significant enhancement of the CoBF responses during controlled stepwise H/H.

Conclusion/Significance

Under resting physiological conditions CB1-receptor mediated mechanisms appear to have limited influence on systemic or cerebral circulation. Enhancement of endocannabinoid levels, however, induces transient CB1-independent hypertension and sustained CB1-mediated hypotension. Furthermore, enhanced endocannabinoid activity results in respiratory depression in a CB1-dependent manner. Finally, our data indicate for the first time the involvement of the endocannabinoid system and CB1-receptors in the regulation of the cerebral circulation during H/H and also raise the possibility of their contribution to the autoregulation of CoBF.     

Endogenous cannabinoid receptor ligand induces the migration of human natural killer cells

2-Arachidonoylglycerol is an endogenous ligand for the cannabinoid receptors (CB1 and CB2). Evidence is gradually accumulating which shows that 2-arachidonoylglycerol plays important physiological roles in several mammalian tissues and cells, yet the details remain ambiguous. In this study, we first examined the effects of 2-arachidonoylglycerol on the motility of human natural killer cells. We found that 2-arachidonoylglycerol induces the migration of KHYG-1 cells (a natural killer leukemia cell line) and human peripheral blood natural killer cells. The migration of natural killer cells induced by 2-arachidonoylglycerol was abolished by treating the cells with SR144528, a CB2 receptor antagonist, suggesting that the CB2 receptor is involved in the 2-arachidonoylglycerol-induced migration. In contrast to 2-arachidonoylglycerol, anandamide, another endogenous cannabinoid receptor ligand, did not induce the migration. Delta9-tetrahydrocannabinol, a major psychoactive constituent of marijuana, also failed to induce the migration; instead, the addition of delta9-tetrahydrocannabinol together with 2-arachidonoylglycerol abolished the migration induced by 2-arachidonoylglycerol. It is conceivable that the endogenous ligand for the cannabinoid receptor, that is, 2-arachidonoylglycerol, affects natural killer cell functions such as migration, thereby contributing to the host-defense mechanism against infectious viruses and tumor cells.