Stimulation of glucagon-like peptide-1 secretion by muscarinic agonist in a murine intestinal endocrine cell line.
Studies on the cholinergic regulation of intestinal L-cells have been focused on the release of enteroglucagon, but the signal transduction pathways were not defined. These were here investigated by using as index the release of immunoreactive glucagon-like peptide-1 (GLP-1) from the endocrine cell line STC-1, that has been shown to contain proglucagon mRNA transcripts. Abundant GLP-1 immunoreactivity was revealed in STC-1 cells at immunocytochemistry and by RIA. The cell content was 4927 +/- 689 pg/10(6) cells, as measured with antiserum 199D that recognizes specifically the C-terminal amidated forms of GLP-1. The secretion of GLP-1 over a 2-h incubation period amounted to 1.4 +/- 0.3% of the total GLP-1 cell content and was significantly increased by 10 microM forskolin and 100 nM 12-O-tetradecanoylphorbol 13-acetate to 206% and 574% of control values, respectively. The cholinergic agonist carbachol stimulated GLP-1 secretion in a concentration-dependent manner, maximal release was observed at 1 mM carbachol (228% of the control value). Binding of the muscarinic antagonist [N-methyl-]scopolamine ([3H]NMS) on cell homogenates was time dependent, specific, and saturable. Scatchard analysis revealed one class of receptors (Kd, 14 pM; binding capacity, 20 fmol/mg protein). Carbachol (0.1 microM to 1 mM) dose dependently displaced [3H] NMS binding and increased the intracellular calcium concentration without modification of adenylate cyclase activity. The order of potency of different antagonists, showing a preferential affinity for M1, M2, and M3 muscarinic receptor subtypes, to inhibit [3H]NMS binding, the carbachol-induced increase in intracellular calcium, and carbachol-stimulated GLP-1 secretion, was as follows: atropine (nonselective) > 4-diphenylacetoxy-N-methylpiperidine methiodide (M3) > pirenzepine (M1) > AF-DX 116 (M2). The results of the present study, therefore, demonstrate that secretion of GLP-1 induced by cholinergic agonist depends on muscarinic M3-subtype receptors in the endocrine intestinal cell line STC-1. This system may prove useful to study the cellular mechanisms of GLP-1 secretion.
Abello
J
J
Institut National de la Santé et de la Recherche Médicale U-45, Hôpital Edouard Herriot, Lyon, France.
Ye
F
F
Bosshard
A
A
Bernard
C
C
Cuber
J C
JC
Chayvialle
J A
JA
D002118:Calcium / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D004702:Endocrine Glands / Q000187:drug effects / Q000378:metabolism*; D005934:Glucagon / Q000378:metabolism*; D052216:Glucagon-Like Peptide 1; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000187:drug effects; D007700:Kinetics; D018727:Muscarinic Antagonists; D019832:N-Methylscopolamine; D010277:Parasympathomimetics / Q000494:pharmacology*; D010446:Peptide Fragments / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000502:physiology*; D012602:Scopolamine Derivatives / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D018727:Muscarinic Antagonists; D010277:Parasympathomimetics; D010446:Peptide Fragments; D011498:Protein Precursors; D011976:Receptors, Muscarinic; D012602:Scopolamine Derivatives; D005576:Colforsin; D052216:Glucagon-Like Peptide 1; D002217:Carbachol; D005934:Glucagon; D000242:Cyclic AMP; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D019832:N-Methylscopolamine