piperidines and 4-(3-3-4-p-menthadien-(1-8)-yl)olivetol

Cannabinoids have been reported to mediate changes in vascular resistance through endothelial receptor targets. We examined involvement of the endothelium in cannabinoid-mediated vasoactive responses in resistance arterioles of the retina.. Vascular responses to both intraluminal (IL) and extraluminal (EL) administration of the atypical cannabinoid, abnormal cannabidiol (abn-CBD), a prototypical agonist at the non-CB1/CB2 endothelial cannabinoid receptor (CBeR), were studied in endothelial intact and endothelial denuded, isolated perfused porcine retinal arterioles with and without endothelin-1 (ET-1) precontraction. The effects of AM251, a CB1 receptor antagonist, and O-1918, an analog of CBD reported to antagonize CBeR, were also studied.. Dose-dependent vasocontractile responses were induced by both IL and EL administration of abn-CBD in the absence of precontraction. Significantly greater vasoconstriction was induced by IL administration of abn-CBD than with EL administration. In contrast, only vasodilation to abn-CBD was observed in ET-1 precontracted retinal arterioles. Endothelium removal significantly reduced abn-CBD-induced vasoactivity when abn-CBD was used IL but not when applied EL. IL abn-CBD-induced vasoactivity was antagonized by O-1918 and AM251.. Cannabinoids show complex vasoactive actions in isolated perfused retinal arterioles. The fact that abn-CBD-mediated vasorelaxation was seen only in precontracted retinal vessels indicates that the abn-CBD-induced vasoactive response is highly dependent on vascular tone. Furthermore, IL and EL administration produced differential responses, and removal of endothelium blunted abn-CBD vasoactivity, highlighting the critical role of endothelium in abn-CBD vasoactivity. AM251 and O-1918 inhibition of abn-CBD-induced vasoactivity suggests the possibility of modulating abn-CBD-induced vasoactivity.

Topics: Animals; Arterioles; Cannabidiol; Dose-Response Relationship, Drug; Endothelin-1; Endothelium, Vascular; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Resorcinols; Retinal Vessels; Swine; Vasodilation

G-protein coupled receptor (GPR)55 is a novel lipid sensing receptor activated by both cannabinoid endogenous ligands (endocannabinoids) and other non-cannabinoid lipid transmitters. This study assessed the effects of various GPR55 agonists on glucose homeostasis.. Insulin secretion and changes in intracellular Ca(2) (+) and cAMP in response to glucose and a range of GPR55 agonists [endogenous ligands (OEA, PEA), chemically synthetic cannabidiol (CBD) analogues (Abn-CBD, 0-1602), an analogue of rimonabant (AM-251) and antagonist (CBD)] were investigated in clonal BRIN-BD11 cells and mouse pancreatic islets. Cytotoxicity was assessed by LDH release, cellular localization by double-staining immunohistochemistry and in vivo effects assessed in mice.. The most potent and selective GPR55 agonist was the synthetic CBD analogue, Abn-CBD (pEC50 10.33), maximum stimulation of 67% at 10(-4)  mol·L(-1) (P < 0.001) in BRIN-BD11 cells. AM-251 (pEC50 7.0), OEA (pEC50 7.0), 0-1602 (pEC50 7.3) and PEA (pEC50 6.0) stimulated insulin secretion. Results were corroborated by islet studies, with no cytotoxic effects. Concentration-dependent insulin secretion by GPR55 agonists was glucose-sensitive and accompanied by elevations of [Ca(2) (+) ]i (P < 0.01-P < 0.001) and cAMP (P < 0.05-P < 0.01). GPR55 agonists exhibited insulinotropic and glucose lowering activity in vivo. GPR55 was expressed on BRIN-BD11 cells and confined to islet beta cells with no distribution on alpha cells.. These results demonstrate GPR55 is distributed in pancreatic beta cells and is a strong activator of insulin secretion, with glucose-lowering effects in vivo. Development of agents agonizing the GPR55 receptor may have therapeutic potential in the treatment of type 2 diabetes.

Topics: Animals; Blood Glucose; Calcium; Cannabidiol; Cell Line; Clone Cells; Cyclic AMP; Diabetes Mellitus, Type 2; Dose-Response Relationship, Drug; Ethanolamines; Hypoglycemic Agents; Insulin; Insulin Secretion; Insulin-Secreting Cells; Islets of Langerhans; Mice; Oleic Acids; Palmitic Acids; Piperidines; Pyrazoles; Rats; Receptors, Cannabinoid; Receptors, G-Protein-Coupled; Resorcinols; Time Factors

The purpose of this study was to investigate the effects of abnormal-cannabidiol (abn-cbd), a non-psychoactive cannabinoid agonist, on aqueous humor outflow via the trabecular meshwork (TM) of porcine eye, and to examine the involvement of a non-CB1/CB2 cannabinoid receptor and the p42/44 mitogen-activated protein kinase (p42/44 MAPK) pathway. The effects of abn-cbd on aqueous humor outflow were measured using a porcine anterior segment perfused organ culture model. The activation of p42/44 MAPK by abn-cbd was determined in cultured TM cells with western blot analysis using an anti-phospho-p42/44 MAPK antibody. Administration of abn-cbd caused a concentration-dependent enhancement of aqueous humor outflow facility with a maximum effect (155.0 ± 11.7% of basal outflow facility) after administration of 30 nM abn-cbd. Pretreatment with 1 μM of O-1918, a cannabidiol analog that acts as a selective antagonist at the non-CB1/CB2 receptor, produced a full antagonism of 30 nM abn-cbd induced increase of aqueous humor outflow facility. Pretreatment with 1 μM of CB1 antagonist SR141716A partially blocked, whereas pretreatment with either 1 μM of CB1 antagonist AM251 or 1 μM of CB2 antagonist SR144528 had no effect on abn-cbd induced enhancement of outflow facility. Treatment of TM cells with 30 nM of abn-cbd activated p42/44 MAPK, which was blocked completely by pretreatment with O-1918, and partially by pretreatment with SR141716A, but not by either AM251 or SR144528. In addition, PD98059, an inhibitor of p42/44 MAPK pathway, blocked completely the abn-cbd induced p42/44 MAPK activation and blocked partially the abn-cbd induced enhancement of outflow facility. In conclusion, the results from this study demonstrate that abn-cbd increases aqueous humor outflow through the TM pathway of the eye, and this effect is mediated by a non-CB1/CB2 cannabinoid receptor, with an involvement of p42/44 MAPK signaling pathway.

Topics: Animals; Anisoles; Aqueous Humor; Blotting, Western; Cannabinoids; Cells, Cultured; Cyclohexanes; Dose-Response Relationship, Drug; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Organ Culture Techniques; Piperidines; Pyrazoles; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Resorcinols; Rimonabant; Swine; Trabecular Meshwork

The receptor(s) used by cannabinoids to relax vascular smooth muscle is unknown. Here, we investigated the effects of 2-arachidonylglyceryl ether (2-AG ether), a metabolically stable endocannabinoid, and abnormal cannabidiol (abn-CBD) on relaxation of permeabilized pulmonary arterial strips monitored with force, and on extracellular signal-regulated mitogen-activated protein kinases (ERK1/2) phosphorylation in permeabilized vascular smooth muscle cells using immunoblotting. We found that 2-AG ether and abn-CBD caused relaxation and increased phosphorylation of ERK1/2. 2-AG ether effects were completely abolished by N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251), and N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (SR141716A), and partially blocked by (-)-1.3-dimethoxy-2-(3-3,4-trans-p-menthadien-(1,8)-yl)-orcinol (O-1918). In contrast, abn-CBD effects were completely abolished by O-1918, and only partially blocked by AM251, and SR141716A. Both 2-AG ether and abn-CBD effects were partially blocked by pertussis toxin, an inhibitor of Gi/o proteins. PD98059, an inhibitor of mitogen activated protein kinase kinase (MEK), completely abolished the relaxation, but only partially blocked the increased phosphorylation of ERK1/2 by 2-AG ether. In contrast, abn-CBD-induced relaxation was partially blocked and the increased phosphorylation of ERK1/2 was abolished by PD98059. These findings suggest that 2-AG ether and abn-CBD-induced vascular smooth muscle relaxation are mediated by the cannabinoid CB1 receptor, and the abn-CBD receptor, respectively, and are modulated by cross-talk between the receptors. These responses occur mainly by coupling to pertussis toxin sensitive G proteins, but also, in part independent of these G proteins, which have been classically thought to initiate MEK/ERK1/2 signaling to relax vascular smooth muscle.

Topics: Animals; Animals, Newborn; Cells, Cultured; Dose-Response Relationship, Drug; Flavonoids; Glycerides; In Vitro Techniques; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Pertussis Toxin; Phosphorylation; Piperidines; Protein Kinase Inhibitors; Pulmonary Artery; Pyrazoles; Rabbits; Receptor Cross-Talk; Receptor, Cannabinoid, CB1; Receptors, G-Protein-Coupled; Resorcinols; Rimonabant; Vasodilation; Vasodilator Agents

We have previously shown that the endocannabinoid anandamide and its metabolically stable analog (R)-methanandamide produce vasorelaxation in rabbit aortic ring preparations in an endothelium-dependent manner that could not be mimicked by other CB(1) cannabinoid receptor agonists (Am J Physiol 282: H2046-H2054, 2002). Here, we show that (R)-methanandamide and abnormal cannabidiol stimulated nitric oxide (NO) production in rabbit aortic endothelial cells (RAEC) in a dose-dependent manner but that other CB(1) and CB(2) receptor agonists, such as cis-3R-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4R-3(3-hydroxypropyl)-1R-cyclohexanol (CP55940) and (R)-(+)-[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl) pyrrolo-[1,2,3-d,e]-1,4-benzoxazin-6-yl]-1-naphthalenyl-methanone (WIN55212-2), failed to do so. CB(1) antagonists rimonabant [also known as SR141716; N-piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] and 6-methoxy-2-(4-methoxyphenyl)benzo[b]-thien-3-yl][4-cyanophenyl]methanone (LY320135) and CB(2) antagonist N-[(1S)-endo-1,3,3,-trimethylbicyclo[2.2.1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528) failed to block (R)-methanandamide-mediated NO production in RAEC. However, anandamide receptor antagonist (-)-4-(3-3,4-trans-p-menthadien-(1,8)-yl)-orcinol (O-1918) blocked (R)-methanandamide-mediated NO production in RAEC. Reverse transcriptase-polymerase chain reaction and Western blot analyses failed to detect the CB(1) receptor in RAEC, making this a good model to study non-CB(1) responses to anandamide. (R)-Methanandamide produced endothelial nitric-oxide synthase (eNOS) phosphorylation via the activation of phosphoinositide 3-kinase-Akt signaling. Inhibition of G(i) signaling with pertussis toxin, or phosphatidylinositol 3-kinase activity with 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), resulted in a decrease in (R)-methanandamide-induced Akt phosphorylation and NO production. Results from this study suggest that in RAEC, (R)-methanandamide acts on a novel non-CB(1) and non-CB(2) anandamide receptor and signals through G(i) and phosphatidylinositol 3-kinase, leading to Akt activation, eNOS phosphorylation, and NO production.

Topics: Animals; Arachidonic Acids; Benzofurans; Benzoxazines; Camphanes; Cannabinoid Receptor Modulators; Cells, Cultured; Chromones; Cyclohexanols; Dose-Response Relationship, Drug; Endocannabinoids; Endothelial Cells; Enzyme Inhibitors; GTP-Binding Protein alpha Subunits, Gi-Go; Morpholines; Naphthalenes; Nitric Oxide; Pertussis Toxin; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Piperidines; Polyunsaturated Alkamides; Proto-Oncogene Proteins c-akt; Pyrazoles; Rabbits; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Resorcinols; Rimonabant; Signal Transduction