Identification and characterisation of functional bombesin receptors in human astrocytes.
Reverse transcription polymerase chain reaction (RT-PCR) demonstrated the presence of bombesin BB2 receptor mRNA but not bombesin BB1 receptor or bombesin BB3 receptor mRNA in cultured human astrocytes. Neuromedin C hyperpolarised human astrocytes in whole-cell current and voltage clamp recordings and increased the intracellular free Ca(2+) ion concentration ([Ca(2+)](i)) in single astrocytes. Treatment with neuromedin C caused larger and more frequent increases in [Ca(2+)](i) than those triggered by neuromedin B, with 96% and 78% of cells responding, respectively. The stimulatory effects of neuromedin C were inhibited significantly by treatment with U73122 or the bombesin BB2 receptor antagonist [D-Phe(6), des-Met(14)]bombesin-(6-14) ethylester. A Fluorometric Imaging Plate Reader (FLIPR) was used to measure [Ca(2+)](i) in cell populations. Neuromedin C was approximately 50-fold more potent than neuromedin B in elevating [Ca(2+)](i) in astrocytes and Chinese hamster ovary (CHO) cells expressing human bombesin BB2 receptors (hBB2-CHO). However, in CHO cells expressing the bombesin BB1 receptor hBB1-CHO, neuromedin B was 32-fold more potent than neuromedin C. [D-Phe(6), des-Met(14)]bombesin-(6-14) ethylester was a partial agonist in hBB1-CHO cells (E(max)=55%) but was a noncompetitive antagonist in both hBB2-CHO cells and astrocytes. These studies report the first identification of functional bombesin receptors on cultured human astrocytes and have demonstrated that the bombesin BB2 receptor contributes significantly to astrocyte physiology.
Mason
Sarah
S
Pfizer Global Research and Development, Cambridge Laboratories, Cambridge University Forvie Site, Robinson Way, Cambridge CB2 2QB, UK.
Smart
Darren
D
Marshall
Ian C B
IC
McKnight
Alexander
A
Skepper
Jeremy N
JN
McNulty
Shaun
S
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D001667:Binding, Competitive; D001839:Bombesin / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007457:Iodine Radioisotopes; D008564:Membrane Potentials / Q000187:drug effects; D015287:Neurokinin B / Q000031:analogs & derivatives* / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010446:Peptide Fragments / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011869:Radioligand Assay; D018004:Receptors, Bombesin / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D007457:Iodine Radioisotopes; D010446:Peptide Fragments; D012333:RNA, Messenger; D018004:Receptors, Bombesin; C040794:neuromedin C; D015287:Neurokinin B; C038393:neuromedin B; D001839:Bombesin; D002118:Calcium