Characterization of functional neuropeptide Y receptors in a human neuroblastoma cell line.
We identified receptors for neuropeptide Y (NPY) on an established human neuroblastoma cell line, SK-N-MC, which are functionally coupled to adenylate cyclase through the inhibitory guanine nucleotide-binding protein of adenylate cyclase, Gi. Intact SK-N-MC cells bound radiolabeled NPY with a KD of 2 nM and contained approximately 83,000 receptors/cell. Unlabeled porcine and human NPY and structurally related porcine peptide YY (PYY) competed with labeled NPY for binding to the receptors. NPY inhibited cyclic AMP accumulation in SK-N-MC cells stimulated by isoproterenol, dopamine, vasoactive intestinal peptide, cholera toxin, and forskolin. NPY inhibited isoproterenol-stimulated cyclic AMP production in a dose-dependent manner, with half-maximal inhibition at 0.5 nM NPY. Porcine and human NPY and porcine PYY gave similar dose-response curves. NPY also inhibited basal and isoproterenol-stimulated adenylate cyclase activity in disrupted cells. Pertussis toxin treatment of the cells completely blocked the ability of NPY to inhibit cyclic AMP production and adenylate cyclase activity. The toxin catalyzed the ADP-ribosylation of a 41-kDa protein in SK-N-MC cells that corresponds to Gi. The receptors on SK-N-MC cells appeared to be specific for NPY, as other neurotransmitter drugs, such as alpha-adrenergic, dopaminergic, muscarinic, and serotonergic antagonists, did not compete for either NPY binding or NPY inhibition of adenylate cyclase. Thus, SK-N-MC cells may be a useful model for investigating NPY receptors and NPY-mediated signal transduction.
Gordon
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EA
Laboratory of Molecular and Cellular Neurobiology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892.
Kohout
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TA
Fishman
P H
PH
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism; D001667:Binding, Competitive; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis; D004298:Dopamine / Q000494:pharmacology; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D009447:Neuroblastoma / Q000378:metabolism*; D009478:Neuropeptide Y / Q000378:metabolism / Q000494:pharmacology; D019894:Peptide YY; D010455:Peptides / Q000378:metabolism; D037342:Pertussis Toxin; D017476:Receptors, Neuropeptide Y; D017981:Receptors, Neurotransmitter / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D009478:Neuropeptide Y; D010455:Peptides; D017476:Receptors, Neuropeptide Y; D017981:Receptors, Neurotransmitter; D010566:Virulence Factors, Bordetella; D019894:Peptide YY; D005576:Colforsin; D000246:Adenosine Diphosphate Ribose; D014660:Vasoactive Intestinal Peptide; D002772:Cholera Toxin; D000242:Cyclic AMP; D037342:Pertussis Toxin; D000262:Adenylyl Cyclases; D007545:Isoproterenol; D004298:Dopamine