A clonal rat pancreatic delta cell line (Rin14B) expresses a high number of galanin receptors negatively coupled to a pertussis-toxin-sensitive cAMP-production pathway.
Galanin, an ubiquitous neuropeptide, was recently shown to inhibit somatostatin release by the rat islet tumor cell line, Rin-m. By using the clonal pancreatic delta cell line Rin14B, originating from Rin-m cells, we were able to identify the presence of one type of specific galanin-binding site of high affinity (Kd = 1.6 nM; maximal binding capacity = 270 fmol/mg protein) and high specificity for the peptide. Binding of 125I-galanin to these receptors was time-dependent and highly sensitive to guanine nucleotides. Using the cross-linker disuccinimidyl tartrate, covalent linking of the galanin receptor to 125I-galanin in membranes from Rin14B cells, followed by SDS/PAGE analysis of membrane proteins, indicated that the galanin receptor is a protein of 54 kDa. 0.1-100 nM galanin also exerted a marked inhibitory effect on the cAMP-production system under basal conditions, as well as in the presence of the pancreatic peptide glucagon. At a maximal dose, galanin induces a 90-100% decrease of basal and glucagon-stimulated cAMP production levels, with a median inhibition concentration (IC50) of 3 nM galanin. The direct inhibitory effect of galanin on the adenylate cyclase activity in Rin14B cell membranes was also demonstrated (IC50 = 3 nM galanin). The inhibitory effect of galanin on the basal and glucagon-stimulated cAMP production in Rin14B cells was reversed by pertussis toxin. The toxin was also shown to specifically ADP-ribosylate a protein of 41 kDa in membranes from Rin14B cells. Taken together, these data show that the pancreatic delta cell line Rin14B expresses high affinity galanin receptors negatively coupled to a pertussis-toxin-sensitive cAMP-production system.
Amiranoff
B
B
INSERM U178, Villejuif, France.
Lorinet
A M
AM
Laburthe
M
M
D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D002999:Clone Cells / Q000187:drug effects / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism*; D005934:Glucagon / Q000494:pharmacology; D007457:Iodine Radioisotopes; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D037342:Pertussis Toxin; D051381:Rats; D044088:Receptors, Galanin; D011964:Receptors, Gastrointestinal Hormone / Q000096:biosynthesis*; D013388:Succinimides / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D037361:Adenylate Cyclase Toxin; D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D044088:Receptors, Galanin; D011964:Receptors, Gastrointestinal Hormone; D013388:Succinimides; D010566:Virulence Factors, Bordetella; C018052:disuccinimidyl tartarate; D005934:Glucagon; D000242:Cyclic AMP; D037342:Pertussis Toxin; D000262:Adenylyl Cyclases