Stimulation of glucagon-like peptide-1 secretion by muscarinic agonist in a murine intestinal endocrine cell line.

Abstract

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.

Peptide YY, Glucagon-Like Peptide-1, and Neurotensin Responses to Luminal Factors in the Isolated Vascularly Perfused Rat Ileum

“…The GLP-1 assay was performed as recently described (17,18). Briefly, antiserum against GLP-1-(7-36) amide was obtained in a rabbit by immunization with synthetic GLP-1-(7-36) amide conjugated to BSA and was used at a final dilution of 1:300,000.…”

Regulation of Glucagon-Like Peptide-1 Synthesis and Secretion in the GLUTag Enteroendocrine Cell Line1

“…An alternative model, the isolated canine L cell (25)(26)(27), provides a more homogeneous cell population, but this culture system requires centrifugal elutriation to prepare and is therefore relatively expensive. A secretin tumor cell line (STC-1) that secretes GLP-1 has also been used as an L cell model (28); however, STC-1 cells are poorly differentiated and multipotential. Hence, the similarity of this intestinal S cell model to the GLP-1-producing L cell is unclear.…”

“…Finally, as bethanechol acts through muscarinic receptors only, whereas carbachol binds to both muscarinic and nicotinic receptors, the present findings suggest a role for a muscarinic receptor in the regulation of GLP-1 secretion. Although the secretin tumor (STC-1) cell line is not a true model of the L cell, studies using these cells have indicated an involvement of the M3 receptor subtype in modulating GLP-1 release (28). Further investigations to determine the receptor subtype expressed by the GLUTag cells are therefore clearly warranted.…”

TRPA1 channel activation induces cholecystokinin release via extracellular calcium

“…In this study, we show that TRPA1 channel is expressed in the human and mouse duodenum. In order to investigate the possible function we used mouse intestinal neuroendocrine cell line STC-1 as a well-established model for intestinal hormone release (CCK, GLP-1, GIP) [12][13][14][15] and show that activation of TRPA1 channels with mustard oil derived AITC elevates the intracellular Ca 2+ levels and stimulates cholecystokinin release.…”