Desensitization of neuromedin B receptors (NMB-R) on native and NMB-R-transfected cells involves down-regulation and internalization.
The receptor for neuromedin B (NMB-R), a mammalian bombesin-related peptide, is widely distributed in the central nervous system and gastrointestinal tract. While it is known that this receptor is coupled to phospholipase C, like many other phospholipase C-activating receptors, little is known about regulation of the NMB-R subsequent to agonist stimulation. We studied both native NMB-R on C-6 rat glioblastoma cells and wild type NMB-R cloned from rat esophageal muscle which was stably transfected into Balb/3T3 fibroblasts. Both cell types rapidly increased [3H]inositol phosphates and [Ca2+]i in response to 1 microM NMB, whereas preincubation with 3 nM NMB for 3 h markedly attenuated the ability of 1 microM NMB, but not 1 microM endothelin-1, to alter either cell type's biological activity. Prolonged exposure to 3 nM NMB caused a rapid decrease in the number of NMB-R, with the maximal receptor down-regulation seen at 24 h due to NMB-R internalization. After maximal down-regulation, removal of agonist resulted in a rapid restoration of NMB-R to the cell surface of both cell types. NMB-R recovery at 6 h was blocked by monensin, an inhibitor of receptor recycling, but was not affected by cycloheximide, a protein synthesis inhibitor. Resensitization to agonist paralleled the recovery of NMB-R in both cell types, and resensitization likewise was blocked by monensin. Our data demonstrate that the NMB-R undergoes rapid homologous desensitization consequent to agonist stimulation, which is mediated by receptor down-regulation and which, in turn, is regulated by internalization. During resensitization, NMB-R reappearance on the cell surface membrane is independent of protein synthesis and is due to a recycling from an intracellular site.
Benya
R V
RV
Digestive Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Kusui
T
T
Shikado
F
F
Battey
J F
JF
Jensen
R T
RT
D016475:3T3 Cells; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D015536:Down-Regulation; D016232:Endothelins / Q000494:pharmacology*; D004947:Esophagus / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism*; D007457:Iodine Radioisotopes; D007700:Kinetics; D051379:Mice; D008985:Monensin / Q000494:pharmacology; D009130:Muscle, Smooth / Q000378:metabolism; D015287:Neurokinin B / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D018004:Receptors, Bombesin / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D014316:Tritium; D014407:Tumor Cells, Cultured
D016232:Endothelins; D007295:Inositol Phosphates; D007457:Iodine Radioisotopes; D018004:Receptors, Bombesin; D014316:Tritium; D015287:Neurokinin B; C038393:neuromedin B; D008985:Monensin; D003513:Cycloheximide; D002118:Calcium