UMLS:C0027819 - FACTA Search

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Query: UMLS:C0027819 (neuroblastoma)
27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously shown that the endogenous putative cannabinoid ligand arachidonylethanolamide (anandamide, 20:4, n - 6) induces in vivo and in vitro effects typical of a cannabinoid agonist. We now report that two other endogenous anandamides, docosatetraenylethanolamide (anandamide, 22:4, n - 6) and homo-gamma-linolenylethanolamide (anandamide, 20:3, n - 6), have similar activities. The new anandamides bind to SV40-transformed African green monkey kidney cells transfected with the rat brain cannabinoid receptor cDNA and display K1 values of 253.4 +/- 41.1 and 244.8 +/- 38.7, respectively. The value found for arachidonylethanolamide was 155.1 +/- 13.8 nM. In addition, the new anandamides inhibit prostaglandin E1-stimulated adenylate cyclase activity in Chinese hamster ovary-K1 cells transfected with the cannabinoid receptor, as well as in N18TG2 mouse neuroblastoma cells that express the cannabinoid receptor naturally. The IC50 values for the inhibition of adenylate cyclase in transfected Chinese hamster ovary-K1 cells were 116.8 +/- 8.7 and 109.3 +/- 8.6 nM for docosatetraenylethanolamide and homo-gamma-linolenylethanolamide, respectively. These values were similar to that obtained with arachidonylethanolamide (100.5 +/- 7.7 nM), but were significantly higher than the IC50 value observed with the plant cannabinoid delta9-tetrahydrocannabinol (9.2 +/- 8.6 nM). The inhibitory effects of the anandamides on adenylate cyclase activity were blocked by pertussis toxin, indicating the involvement of pertussis toxin-sensitive GTP-binding protein(s). In a tetrad of behavioral assays for cannabinoid-like effects, the two new anandamides exerted similar behavioral effects to those observed with delta9-tetrahydrocannabinol and arachidonylethanolamide: inhibition of motor activity in an open field, hypothermia, catalepsy on a ring, and analgesia on a hot plate.
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PMID:Cannabinomimetic behavioral effects of and adenylate cyclase inhibition by two new endogenous anandamides. 874 28

The cannabinoid receptors expressed in the mouse neuroblastoma X rat glioma NG108-15 cell and the rat pituitary tumor GH4C1 cell were determined by polymerase chain reaction, dideoxysequencing and pharmacologically. The CB1 but not the CB2 or CB1A cannabinoid receptor was found in both cell lines. The cDNA identified in GH4C1 cells corresponds to the rat CB1 receptor. Interestingly, NG108-15 cells express two distinct cDNAs, one corresponds to the rat and the other to the mouse CB1 receptor. The newly developed CB1 receptor selective antagonist SR141716A was found to reverse cannabinoid agonist (WIN55212-2 or CP55940)-induced adenylyl cyclase inhibition. These results provide more direct evidence that the CB1 receptor is mediating the pharmacological actions of cannabinoids in NG108-15 and GH4C1 cells.
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PMID:Determination of the cannabinoid receptors in mouse x rat hybridoma NG108-15 cells and rat GH4C1 cells. 883 54

Anandamide (arachidonoyl-ethanolamide, AnNH) and 2-arachidonoyl-glycerol (2-AG) have been suggested to act as endogenous agonists at the brain cannabinoid receptor, and their biosynthetic and degradative mechanisms in nervous tissues and cells have also been partially elucidated. Here we present evidence for the presence, in mouse N18TG2 neuroblastoma cells, of enzymatic activities potentially responsible for the biosynthesis of AnNH and 2-AG from a common phospholipid precursor. Cell homogenates were shown to catalyze: (a) the transfer of an arachidonoyl moiety from the sn-1 position of sn-1,2-di-arachidonoyl-phosphatidylcholine (AAPC) to phosphatidyl-ethanolamine (PE) to form N-arachidonoyl-PE (N-ArPE) and sn-1-lyso-2-arachidonoyl-PC (lyso-APC), (b) the hydrolysis of N-AtPE to AnNH, (c) the hydrolysis of lyso-APC to 2-AG, (d) the hydrolysis of AAPC to sn-1,2-di-arachidonoyl-glycerol (AAG), and (e) the hydrolysis of AAG to 2-AG. From these findings it is possible to suggest that AAPC may serve as precursor for both AnNH and 2-AG biosynthesis through three different pathways.
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PMID:Potential biosynthetic connections between the two cannabimimetic eicosanoids, anandamide and 2-arachidonoyl-glycerol, in mouse neuroblastoma cells. 885 37

The predominant animal model in which the pharmacology of cannabinoids is studied is the mouse. Nonetheless, the structure and functional expression of the mouse cannabinoid receptor (CB1) gene have not been reported. We have cloned and expressed the gene for the mouse CB1 receptor and compared its properties with those of native mouse CB1 receptors in brain and N18TG2 neuroblastoma cells. The mouse CB1 gene was isolated from a mouse 129 strain genomic library. Sequence analysis of a 6-kb BamHI fragment of the mouse CB1 genomic clone indicates 95% nucleic acid identity between mouse and rat (99.5% amino acid identity) and 90% nucleic acid identity (97% amino acid identity) between mouse and human. Examination of the 5' untranslated sequence of the mouse CB1 genomic clone revealed a splice junction site approximately 60 bp upstream from the translation start site, indicating the possibility of splice variants of the CB1 receptors. The coding region of the mouse CB1 receptor was stably expressed in 293 cells, and binding by [3H]SR 141716A and [3H]CP-55,940 was determined. The Bmax and Kd values obtained with [3H]SR 141716A (921 +/- 58 fmol/mg and 0.73 +/- 0.13 nM, respectively) were similar to those of native mouse CB1 receptors in brain (Bmax of 1.81 +/- 0.44 pmol/mg, Kd of 0.16 +/- 0.01 nM) and N18TG2 cells (Bmax of 197 +/- 29 fmol/mg, Kd of 0.182 +/- 0.08 nM). The mouse CB1 receptor genomic clone will be a useful tool for studying the function and regulation of the CB1 receptor in mice.
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PMID:Isolation and expression of a mouse CB1 cannabinoid receptor gene. Comparison of binding properties with those of native CB1 receptors in mouse brain and N18TG2 neuroblastoma cells. 903 53

Anandamide amidase (EC 3.5.1.4) is responsible for the hydrolysis of arachidonoyl ethanolamide (anandamide). Relatively selective and potent enzyme reversible inhibitors effective in the low micromolar range, such as arachidonyl trifluoromethyl ketone (Arach-CF3), have been described (Koutek et al., J Biol Chem 269: 22937-22940, 1994). In the current study, methyl arachidonyl fluorophosphonate (MAFP), an arachidonyl binding site directed phosphonylation reagent, was tested as an inhibitor of anandamide amidase and as a ligand for the CB1 cannabinoid receptor. MAFP was 800 times more potent than Arach-CF3 and phenylmethylsulfonyl fluoride (PMSF) as an amidase inhibitor in rat brain homogenates. In intact neuroblastoma cells, MAFP was also approximately 1000-fold more potent than Arach-CF3. MAFP demonstrated selectivity towards anandamide amidase for which it was approximately 3000 and 30,000-fold more potent than it was towards chymotrypsin and trypsin, respectively. MAFP displaced [3H]CP-55940 binding to the CB1 cannabinoid receptor with an IC50 of 20 nM vs 40 nM for anandamide. It bound irreversibly and prevented subsequent binding of the cannabinoid radioligand [3H]CP-55940 at that locus. These studies suggest that MAFP is a potent and specific inhibitor of anandamide amidase and, in addition, can interact with the cannabinoid receptors at the cannabinoid binding site. This is the first report of a potent and relatively selective irreversible inhibitor of arachidonoyl ethanolamide amidase.
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PMID:Methyl arachidonyl fluorophosphonate: a potent irreversible inhibitor of anandamide amidase. 906 28

The monoacylglycerol 2-arachidonoylglycerol (2-AG) has been recently suggested as a possible endogenous agonist at cannabinoid receptors both in brain and peripheral tissues. Here we report that a widely used model for neuronal cells, mouse N18TG2 neuroblastoma cells, which contain the CB1 cannabinoid receptor, also biosynthesize, release and degrade 2-AG. Stimulation with ionomycin (1-5 microM) of intact cells prelabelled with [3H]arachidonic acid ([3H]AA) led to the formation of high levels of a radioactive component with the same chromatographic behaviour as synthetic standards of 2-AG in TLC and HPLC analyses. The amounts of this metabolite were negligible in unstimulated cells, and greatly decreased in cells stimulated in the presence of the Ca2+-chelating agent EGTA. The purified component was further characterized as 2-AG by: (1) digestion with Rhizopus arrhizus lipase, which yielded radiolabelled AA; (2) gas chromatographic-MS analyses; and (3) TLC analyses on borate-impregnated plates. Approx. 20% of the 2-AG produced by stimulated cells was found to be released into the incubation medium when this contained 0.1% BSA. Subcellular fractions of N18TG2 cells were shown to contain enzymic activity or activities catalysing the hydrolysis of synthetic [3H]2-AG to [3H]AA. Cell homogenates were also found to convert synthetic [3H]sn-1-acyl-2-arachidonoylglycerols (AcAGs) into [3H]2-AG, suggesting that 2-AG might be derived from AcAG hydrolysis. When compared with ionomycin stimulation, treatment of cells with exogenous phospholipase C, but not with phospholipase D or A2, led to a much higher formation of 2-AG and AcAGs. However, treatment of cells with phospholipase A2 10 min before ionomycin stimulation caused a 2.5-3-fold potentiation of 2-AG and AcAG levels with respect to ionomycin alone, whereas preincubation with the phospholipase C inhibitor neomycin sulphate did not inhibit the effect of ionomycin on 2-AG and AcAG levels. These results suggest that the Ca2+-induced formation of 2-AG proceeds through the intermediacy of AcAGs but not necessarily through phospholipase C activation. By showing for the first time the existence of molecular mechanisms for the inactivation and the Ca2+-dependent biosynthesis and release of 2-AG in neuronal cells, the present paper supports the hypothesis that this cannabimimetic monoacylglycerol might be a physiological neuromodulator.
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PMID:Biosynthesis, release and degradation of the novel endogenous cannabimimetic metabolite 2-arachidonoylglycerol in mouse neuroblastoma cells. 906 92

1. Aminoalkylindoles, typified by WIN 55212-2, bind to G protein-coupled cannabinoid receptors in brain. Although cannabinoids inhibit adenylyl cyclase in NG108-15 neuroblastoma x glioma hybrid cells, cannabinoid receptor binding in these cells has not been described previously. This study compares pharmacological characteristics of [3H]WIN 55212-2 binding sites in rat cerebellar membranes and in NG108-15 membranes. 2. Although the KD of specified [3H]WIN 55212-2 binding was similar in brain and NG108-15 membranes, the Bmax was 10 times lower in NG108-15 than in cerebellar membranes. In both brain and NG108-15 membranes, aminoalkylindole analogues were relatively potent in displacing [3H]WIN 55212-2 binding. However, IC50 values for more traditional cannabinoids were significantly higher in NG108-15 membranes than in brain, e.g., the Ki values for CP55,940 were 1.2 nM in brain and > 5000nM in NG108-15 membranes. Moreover, sodium and GTP-gamma-S decreased [3H]WIN 55212-2 binding in brain but not in NG108-15 membranes. 3. These data suggest that WIN 55212-2 does not label traditional cannabinoid receptors in NG108-15 cells and that these novel aminoalkylindole binding sites are not coupled to G proteins.
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PMID:Binding of aminoalkylindoles to noncannabinoid binding sites in NG108-15 cells. 935 90

Fatty acid amide hydrolase (FAAH) catalyzes the hydrolysis of bioactive fatty acid amides and esters such as the endogenous cannabinoid receptor ligands, anandamide (N-arachidonoyl-ethanolamine) and 2-arachidonoylglycerol, and the putative sleep inducing factor cis-9-octadecenoamide (oleamide). Most FAAH blockers developed to date also inhibit cytosolic phospholipase A2 (cPLA2) and/or bind to the CB1 cannabinoid receptor subtype. Here we report the finding of four novel FAAH inhibitors, two of which, malhamensilipin A and grenadadiene, were screened out of a series of thirty-two different algal natural products, and two others, arachidonoylethylene glycol (AEG) and arachidonoyl-serotonin (AA-5-HT) were selected out of five artificially functionalized polyunsaturated fatty acids. When using FAAH preparations from mouse neuroblastoma N18TG2 cells and [14C]anandamide as a substrate, the IC50s for these compounds ranged from 12.0 to 26 microM, the most active compound being AA-5-HT. This substance was also active on FAAH from rat basophilic leukaemia (RBL-2H3) cells (IC50 = 5.6 microM), and inhibited [14C]anandamide hydrolysis by both N18TG2 and RBL-2H3 intact cells without affecting [14C]anandamide uptake. While AEG behaved as a competitive inhibitor and was hydrolyzed to arachidonic acid (AA) by FAAH preparations, AA-5-HT was resistant to FAAH-catalyzed hydrolysis and behaved as a tight-binding, albeit non-covalent, mixed inhibitor. AA-5-HT did not interfere with cPLA2-mediated, ionomycin or antigen-induced release of [3H]AA from RBL-2H3 cells, nor with cPLA2 activity in cell-free experiments. Finally, AA-5-HT did not activate CB1 cannabinoid receptors since it acted as a very weak ligand in in vitro binding assays, and, at 10-15 mg/kg body weight, it was not active in the 'open field', 'hot plate' and rectal hypothermia tests carried out in mice. Conversely AEG behaved as a cannabimimetic substance in these tests as well as in the 'ring' immobility test where AA-5-HT was also active. AA-5-HT is the first FAAH inhibitor reported to date which is inactive both against cPLA2 and at CB1 receptors, whereas AEG represents a new type of cannabinoid receptor agonist.
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PMID:Arachidonoylserotonin and other novel inhibitors of fatty acid amide hydrolase. 970 57

In this study we employed the neuroblastoma x glioma NG 108-15 cell line as a model for investigating the effects of long-term activation of cannabinoid receptors on delta opioid receptor desensitization, down-regulation and gene expression. Exposure of NG 108-15 cells to (-)-delta9-tetrahydrocannabinol (delta9-THC) reduced opioid receptor binding, evaluated in intact cells, by approximately 40-45% in cells exposed for 24 h to 50 and 100 nM delta9-THC and by approximately 25% in cells exposed to 10 nM delta9-THC. Lower doses of delta9-THC (0.1 and 1 nM) or a shorter exposure time to the cannabinoid (6 h) were not effective. Down-regulation of 6 opioid receptors was not observed in cells exposed for 24 h to pertussis toxin (PTX) and then treated for 24 h with 100 nM delta9-THC. In cells that were exposed for 24 h to the cannabinoid, the ability of delta9-THC and of the delta opioid receptor agonist [D-Ser2, Leu5, Thr6]enkephalin to inhibit forskolin-stimulated cAMP accumulation was significantly attenuated. Prolonged exposure of NG 108-15 cells to 100 nM delta9-THC produced a significant elevation of steady-state levels of delta opioid receptor mRNA. This effect was not observed in cells pretreated with PTX. The selective cannabinoid receptor antagonist SR 141716A blocked the effects elicited by delta9-THC on delta opioid receptor desensitization, down-regulation and gene expression; thus indicating that these are mediated via activation of cannabinoid receptors. These data demonstrate the existence, in NG 108-15 cells, of a complex cross-talk between the cannabinoid and opioid receptors on prolonged exposure to delta9-THC triggered by changes in signaling through Gi and/or G0-coupled receptors.
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PMID:Regulation of delta opioid receptors by delta9-tetrahydrocannabinol in NG108-15 hybrid cells. 977 17

A CB1 cannabinoid receptor peptide fragment from the C-terminal juxtamembrane region autonomously inhibits adenylyl cyclase activity in a neuroblastoma membrane preparation. The cannabinoid receptor antagonist, SR141716A, failed to block the response. The peptide was able to evoke the response in membranes from Chinese hamster ovary (CHO) cells that do not express the CB1 receptor. These studies are consistent with a direct activation of Gi by the peptide. To test the importance of a BXBXXB sequence, Lys403 was acetylated, resulting in a peptide having similar affinity but reduced efficacy. N-Terminal truncation of Arg401 resulted in a 6-fold loss of affinity, which was not further reduced by sequential truncation of up to the first seven amino acids, four of which are charged. N-Terminal-truncated peptides exhibited maximal activity, suggesting that Gi activation can be conferred by the remaining amino acids. Truncation of the C-terminal Glu417 or substitution of Glu417 by a Leu or of Arg401 by a Norleucine reduced activity at 100 microM. The C-terminal juxtamembrane peptide was constrained to a loop peptide by placement of Cys residues at both terminals and disulfide coupling. This modification reduced the affinity 3-fold but yielded near-maximal efficacy. Blocking the Cys termini resulted in a loss of efficacy. Circular dichroism spectropolarimetry revealed that all C-terminal juxtamembrane peptide analogues exist in a random coil conformation in an aqueous environment. A hydrophobic environment (trifluoroethanol) failed to induce alpha-helix formation in the C-terminal juxtamembrane peptide but did so in less active peptides. The anionic detergent sodium dodecyl sulfate induced alpha-helix formation in all analogues except the loop peptide, where it induces a left-handed PII conformation. It is concluded that alpha-helix formation is not required for Gi activation.
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PMID:Regulation of Gi by the CB1 cannabinoid receptor C-terminal juxtamembrane region: structural requirements determined by peptide analysis. 1007 92

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