Internalization of glucagon-like peptide-1(7-36)amide in rat insulinoma cells.
Glucagon-like peptide-1(7-36)amide [GLP-1(7-36)amide] is supposed to be an important physiologic incretin. Recently, high affinity receptors for GLP-1(7-36)amide have been demonstrated on rat insulinoma-derived RINm5F cells. The present study examined the internalization and degradation of the GLP-1-receptor complex. Internalization of the peptide was time- and temperature-dependent. At 37 degrees C binding and internalization was rapid. At 60 min 35% of 125I-labeled GLP-1(7-36)amide was internalized. Incubation in the presence of increasing concentrations of non-labeled GLP-1(7-36)amide resulted in a decrease of internalization of 125I-labeled peptide indicating that this process is saturable. Incubation in the presence of 0.2 mM chloroquine, an inhibitor of intracellular hormone degradation, resulted in intracellular accumulation of 125I-GLP-1(7-36)amide. HPLC-supported analysis of cell content after internalization of 125I-GLP-1(7-36)amide during a 60-min incubation period at 37 degrees C revealed an elution profile showing two maxima of radioactivity: one represented intact labeled GLP-1(7-36)amide, the other an intracellular degradation product of the peptide. Chloroquine caused a 5-fold increase of the peak representing intact 125I-GLP-1(7-36)amide thus demonstrating inhibition of degradation of labelled peptide. Furthermore, a 4-fold increase of the other peak occurred possibly mirroring a delay of release of degradation products by chloroquine. It was excluded that chloroquine is able to interfere with GLP-1(7-36)amide-binding to its receptor.
Göke
R
R
Dept. of Internal Medicine, Philipps University of Marburg, Federal Republic of Germany.
Richter
G
G
Göke
B
B
Trautmann
M
M
Arnold
R
R
D007516:Adenoma, Islet Cell / Q000378:metabolism*; D000818:Animals; D002738:Chloroquine / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D005934:Glucagon / Q000378:metabolism*; D052216:Glucagon-Like Peptide 1; D000067757:Glucagon-Like Peptide-1 Receptor; D004763:Glucagon-Like Peptides; D007340:Insulinoma / Q000378:metabolism*; D007700:Kinetics; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D010455:Peptides / Q000032:analysis / Q000378:metabolism*; D051381:Rats; D011956:Receptors, Cell Surface / Q000378:metabolism; D018027:Receptors, Glucagon; D013696:Temperature; D014407:Tumor Cells, Cultured
C000593835:Glp1r protein, rat; D000067757:Glucagon-Like Peptide-1 Receptor; D010455:Peptides; D011956:Receptors, Cell Surface; D018027:Receptors, Glucagon; D004763:Glucagon-Like Peptides; D002738:Chloroquine; D052216:Glucagon-Like Peptide 1; D005934:Glucagon; C055454:glucagon-like peptide 1 (1-36)amide