Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

N18TG2 neuroblastoma cells co-express delta-opioid and CB1-cannabinoid receptors. Both receptors are negatively coupled to adenylyl cyclase through pertussis toxin-sensitive GTP-binding proteins. In the present study, we confirmed the independent activity of opioid and cannabinoid agonists, and investigated chronic interactions between the two signal transduction pathways in these cells. Opioid and cannabinoid agonists stimulated [35S]guanosine-5'-O-(3-thiotriphosphate) binding to N18TG2 membranes. When the opioid agonist etorphine and the cannabinoid agonist desacetyllevonantradol (DALN) were applied together, the stimulation was similar to the arithmetic sum of the two separate effects. This additivity existed even after partial ablation of the G-proteins reservoir with a low concentration of pertussis toxin, indicating that opioid and cannabinoid receptors activate different pools of G-proteins in N18TG2 cells. Chronic treatment of the cells with either opioid or cannabinoid agonists induced desensitization to the respective drug. In addition, asymmetric cross-desensitization was found: while long-term exposure to DALN induced homologous desensitization, and did not reduce the effect of etorphine, long-term exposure to etorphine attenuated the cannabinoid activation of G-proteins. Chronic exposure to either DALN or etorphine not only induced desensitization, but also elevated the basal activity of G-proteins in the exposed cells. The combination of the two drugs did not yield an additive activation, suggesting that chronic exposure of N18TG2 cultures to cannabinoid and opioid agonists modified a common responding element within the cells. This work presents the N18TG2 neuroblastoma as a suitable experimental model to study the molecular mechanism(s) underlying chronic interactions between opioid and cannabinoid drugs.
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PMID:Independence of, and interactions between, cannabinoid and opioid signal transduction pathways in N18TG2 cells. 973 2

1. G protein-coupled receptor kinases (GRKs) are thought to be important in mediating the agonist-induced phosphorylation and consequent desensitization of G protein-coupled receptor (GPCR) responses. We have previously shown that stable expression of a dominant negative mutant G protein-coupled receptor kinase 2 (GRK2) construct in NG108-15 mouse neuroblastoma x rat glioma cells suppresses the agonist-induced desensitization of A2A and A2B adenosine receptor-stimulated adenylyl cyclase activity (Mundell et al., 1997). To further determine the role of GRK2 in agonist-induced desensitization of these adenosine receptors, we stably overexpressed wild type GRK2 in NG108-15 cells. 2. In homogenates prepared from cells overexpressing GRK2, the acute stimulation of adenylyl cyclase by activation of A2A and A2B adenosine receptors was markedly reduced, but could be reversed by pretreating the cells with AD (adenosine deaminase), to remove extracellular adenosine from the medium. On the other hand, acute stimulation of adenylyl cyclase by secretin, iloprost, NaF and forskolin was the same in GRK2 overexpressing cells and plasmid-transfected control cells. 3. Cells overexpressing GRK2 were more sensitive to adenosine receptor agonist-induced desensitization than plasmid-transfected control cells. This effect was selective since the agonist sensitivity of desensitization for secretin and IP-prostanoid receptor-stimulated adenylyl cyclase activity was not affected by GRK2 overexpression. 4. These results further implicate GRK2 as the likely mechanism by which A2 adenosine receptors undergo short-term desensitization in NG108-15 cells, and indicate that even when overexpressed, GRK2 retains its substrate specificity for native receptors in intact cells. Furthermore, the susceptibility of GPCRs to desensitization appears to depend on the level of GRK expression, such that in cells that express high levels of GRK2, low agonist concentrations may be sufficient to trigger GRK-mediated desensitization.
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PMID:Enhanced expression of G protein-coupled receptor kinase 2 selectively increases the sensitivity of A2A adenosine receptors to agonist-induced desensitization. 978 8

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

Chronic activation of the mu-opioid receptor (MOR1TAG) results in the loss of agonist response that has been attributed to desensitization and down-regulation of the receptor. It has been suggested that opioid receptor phosphorylation is the mechanism by which this desensitization and down-regulation occurs. When MOR1TAG was stably expressed in both neuroblastoma neuro2A and human embryonic kidney HEK293 cells, the opioid agonist [D-Ala2,MePhe4, Gly5-ol]enkephalin (DAMGO) induced a time- and concentration-dependent phosphorylation of the receptor, in both cell lines, that could be reversed by the antagonist naloxone. Protein kinase C can phosphorylate the receptor, but is not involved in DAMGO-induced MOR1TAG phosphorylation. The rapid rate of receptor phosphorylation, occurring within minutes, did not correlate with the rate of the loss of agonist-mediated inhibition of adenylyl cyclase, which occurs in hours. This lack of correlation between receptor phosphorylation and the loss of response was further demonstrated when receptor phosphorylation was increased by either calyculin A or overexpression of the G-protein receptor kinases. Calyculin A increased the magnitude of MOR1TAG phosphorylation without altering the DAMGO-induced loss of the adenylyl cyclase response. Similarly, when mu- and delta-opioid (DOR1TAG) receptors were expressed in the same system, overexpression of beta-adrenergic receptor kinase 2 elevated agonist-induced phosphorylation for both receptors. However, in the same cell lines under the same conditions, overexpression of beta-adrenergic receptor kinase 2 and beta-arrestin 2 accelerated the rate of DPDPE- but not DAMGO-induced receptor desensitization. Thus, these data show that phosphorylation of MOR1TAG is not an obligatory event for the DAMGO-induced loss in the adenylyl cyclase regulation by the receptor.
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PMID:The absence of a direct correlation between the loss of [D-Ala2, MePhe4,Gly5-ol]Enkephalin inhibition of adenylyl cyclase activity and agonist-induced mu-opioid receptor phosphorylation. 1009 93

The efficacy of different opioid agonists to induce acute desensitization of the human delta-opioid receptor-mediated inhibition of cAMP accumulation was investigated in the neuroblastoma cell line SK-N-BE, which endogenously expresses these receptors. While etorphine, a non-selective alkaloid agonist, caused 50% desensitization after a 30-min incubation, the same treatment in the presence of the selective peptide agonists, DPDPE ([D-Pen2,D-Pen5]enkephalin) and deltorphin I (Tyr-D-Ala-Phe-Asp-Val-Val-Gly), almost totally desensitized the delta-opioid receptor-mediated inhibition of adenylyl cyclase. When SK-N-BE cells were prechallenged either with alkaloid or peptide agonist, we observed a cross-desensitization that was less marked when cells were pretreated with peptide agonists and then challenged with etorphine. Taken together, these results demonstrate that human delta-opioid receptors are differentially desensitized by alkaloid and peptide agonists.
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PMID:Differential desensitization of human delta-opioid receptors by peptide and alkaloid agonists. 1035 61

Huntingtin is a cytoplasmic protein of unknown function that associates with vesicle membranes and microtubules. Its protein interactions suggest that huntingtin has a role in endocytosis and organelle transport. In this study we sought to identify factors that regulate the transport of huntingtin in striatal neurons, which are the cells most affected in Huntington's disease. In clonal striatal cells derived from fusions of neuroblastoma and embryonic striatal neurons, huntingtin localization is diffuse and slightly punctate in the cytoplasm. When these neurons were differentiated by treatment with forskolin, huntingtin redistributed to perinuclear regions, discrete puncta along plasma membranes, and branch points and terminal growth cones in neurites. Huntingtin staining overlapped with clathrin, a coat protein involved in endocytosis. Immunoblot analysis of subcellular membrane fractions separated by differential centrifugation confirmed that huntingtin immunoreactivity in differentiated neurons markedly increased in membrane fractions enriched with clathrin and with huntingtin-interacting protein 1. Dopamine treatment altered the subcellular localization of huntingtin and increased its expression in clathrin-enriched membrane fractions. The dopamine-induced changes were blocked by the D1 antagonist SCH 23390 and were absent in a clonal cell line lacking D1 receptors. Results suggest that the transport of huntingtin and its co-expression in clathrin and huntingtin-interacting protein 1-enriched membranes is influenced by activation of adenylyl cyclase and stimulation of dopamine D1 receptors.
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PMID:Forskolin and dopamine D1 receptor activation increase huntingtin's association with endosomes in immortalized neuronal cells of striatal origin. 1036 4

Mood stabilizing drugs decrease central nervous system cyclic AMP signaling. We report here that chronic, but not acute treatment with lithium chloride in human neuroblastoma SH-SY5Y cells, inhibits phosphorylation of cyclic AMP responsive element binding protein and cyclic AMP responsive element DNA binding induced by the adenylyl cyclase activator forskolin, but has no effect on constitutive expression of cyclic AMP responsive element binding protein. These results are consistent with an effect of lithium to blunt the cyclic AMP signal transduction pathway. Such an effect is not shared by the other commonly prescribed mood stabilizer, sodium valproate. Our results suggest that cyclic AMP responsive element binding protein regulated gene expression may be relevant to the long-term prophylactic effect of lithium. Furthermore, sodium valproate, which is also effective in bipolar disorder, would appear to act on other pathways to bring about its therapeutic effects.
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PMID:Cyclic AMP responsive element binding protein phosphorylation and DNA binding is decreased by chronic lithium but not valproate treatment of SH-SY5Y neuroblastoma cells. 1036 32

Nitric oxide (NO) donors inhibit hormone- and forskolin-stimulated adenylyl cyclase activity in purified plasma membrane preparations from N18TG2 neuroblastoma cells. Northern blot analyses indicate that the predominant isoform of adenylyl cyclase in N18TG2 cells is the type VI. Our experiments eliminate all the known regulatory proteins for this isoform as possible targets of NO. NO decreases the Vmax of the enzyme without altering the Km for ATP. Occupancy of the substrate-binding site protects the enzyme from the inhibitory effects of NO, suggesting that the conformation of the enzyme determines its sensitivity. The inhibition is reversed by reducing agents, implicating a Cys residue(s) as the target for nitric oxide and an S-nitrosylation as the underlying modification. These findings implicate NO as a novel cellular regulator of the type VI isoform of adenylyl cyclase.
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PMID:Adenylyl cyclase, a coincidence detector for nitric oxide. 1038 85

Pituitary adenylyl cyclase activating polypeptide (PACAP-27), forskoline and carbachol increased type A atrial natriuretic peptide receptor (NPR-A) density, as well as NPR-A mRNA level, in the human neuroblastoma NB-OK-1 cell line. TPA did not have any effect per se, but blunted the effect of PACAP-27 on both NPR-A density and NPR-A mRNA. The half-life of the NPR-A mRNA was not modified by any of the agents tested. Our data support an original transcriptional upregulation of human NPR-A in response to cAMP-induced agents, and in response to carbachol.
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PMID:Human A-type ANP receptor upregulation by PACAP and carbachol in neuroblastoma cells. 1041 13

Recent evidence supporting a role for phosphoinositides in the endocytosis of phospholipase C-coupled receptors has prompted an investigation of whether there exists a similar requirement for the internalization of adenylyl cyclase-linked receptors. When 1321N1 astrocytoma cells, which possess both muscarinic cholinergic receptors (mAChRs) that couple to phospholipase C and beta-adrenergic receptors (beta(2)-ARs) linked to adenylyl cyclase, were pretreated with wortmannin (WT) at a concentration known to inhibit phosphatidylinositol 4-kinase activity, the labeling of both phosphatidylinositol 4-phosphate and phosphatidylinositol 4, 5-bisphosphate (PIP(2)) was reduced. Stimulation of phosphoinositide breakdown by activation of mAChRs in WT-pretreated cells led to a further depletion of PIP(2). As previously demonstrated for SH-SY5Y neuroblastoma, inclusion of WT inhibited the endocytosis of mAChRs in 1321N1 cells by >85%. In contrast, the internalization of beta(2)-ARs was only partially ( approximately 30%) prevented. However, when the concentration of PIP(2) was further reduced by exposure of WT-pretreated 1321N1 cells to a muscarinic agonist, the endocytosis of beta(2)-ARs was substantially inhibited (>70%). Lower concentrations of WT (100 nM) that were sufficient to fully inhibit phosphatidylinositol 3-kinase activity had no effect on either phosphoinositide synthesis or receptor endocytosis. The results indicate that the agonist-induced endocytosis of an adenylyl cyclase-linked receptor such as the beta(2)-AR, like that of the phospholipase C-coupled mAChR, is dependent on the synthesis of phosphoinositides and, in particular, that of PIP(2).
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PMID:Inhibition of beta(2)-adrenergic and muscarinic cholinergic receptor endocytosis after depletion of phosphatidylinositol bisphosphate. 1041 68


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