G-protein coupling and signalling of Y1-like neuropeptide Y receptors in SK-N-MC cells.
We have studied [125I]neuropeptide Y-binding sites and neuropeptide Y-mediated second messenger responses in human SK-N-MC neuroblastoma cells with special reference to the role of G-proteins. Neuropeptide Y stimulated two second messenger responses in SK-N-MC cells, inhibition of cAMP accumulation and mobilization of Ca2+ from intracellular stores. Both effects were completely abolished by pretreatment with pertussis toxin. Binding of [125I]neuropeptide Y to intact cells or SK-N-MC cell membranes was rapid, reversible, characterized by high affinity and low capacity, and had pharmacological characteristics of a homogeneous population of Y1-like neuropeptide Y receptors. In permeabilized cells, [125I] neuropeptide Y binding was inhibited by GTP gamma S in a concentration-dependent manner. Saturation experiments in the absence and presence of GTP gamma S demonstrated a reduction in the number of high-affinity [125I]neuropeptide Y-binding sites without a decrease in affinity of the remaining sites. Pretreatment of intact cells with pertussis toxin completely abolished the inhibition of [125I]neuropeptide Y binding by GTP gamma S. Moreover, pertussis toxin treatment reduced the number of high-affinity [125I]neuropeptide Y binding sites. We conclude that the agonist ligand [125I]neuropeptide Y identifies functional neuropeptide Y receptors in SK-N-MC cells; however, the number of specific [125I]neuropeptide Y-binding sites may not necessarily reflect the number of neuropeptide Y receptors, because the former is affected by the functional state of cellular G-proteins.
Feth
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Department of Pediatrics, University of Essen, Federal Republic of Germany.
Rascher
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Michel
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D002118:Calcium / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D007457:Iodine Radioisotopes; D007700:Kinetics; D009447:Neuroblastoma / Q000378:metabolism / Q000648:ultrastructure; D009478:Neuropeptide Y / Q000378:metabolism; D037342:Pertussis Toxin; D017476:Receptors, Neuropeptide Y; D017981:Receptors, Neurotransmitter / Q000378:metabolism* / Q000502:physiology; D015290:Second Messenger Systems / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D014407:Tumor Cells, Cultured / Q000648:ultrastructure; D010566:Virulence Factors, Bordetella / Q000378:metabolism / Q000494:pharmacology
D007457:Iodine Radioisotopes; D009478:Neuropeptide Y; D017476:Receptors, Neuropeptide Y; D017981:Receptors, Neurotransmitter; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000242:Cyclic AMP; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D002118:Calcium