ACTH stimulates insulin secretion from MIN6 cells and primary mouse and human islets of Langerhans.
It has previously been suggested that ACTH and ACTH-related peptides may act as paracrine modulators of insulin secretion in the islets of Langerhans. We have, therefore, examined the expression and function of the ACTH receptor (the melanocortin 2 receptor, MC2-R) in human and mouse primary islet tIssue and in the MIN6 mouse insulinoma cell line. Mouse MC2-R mRNA was detected in both MIN6 cells and mouse islet tIssue by PCR amplification of cDNA. In perifusion experiments with MIN6 pseudo-islets, a small, transient increase in insulin secretion was obtained when ACTH(1-24) (1 nM) was added to medium containing 2 mM glucose (control) but not when the medium glucose content was increased to 8 mM. Further investigations were performed using static incubations of MIN6 cell monolayers; ACTH(1-24) (1 pM-10 nM) provoked a concentration-dependent increase in insulin secretion from MIN6 monolayer cells that achieved statistical significance at concentrations of 1 and 10 nM (150 +/- 13.6% basal secretion; 187 +/- 14.9% basal secretion, P<0.01). Similar responses were obtained with ACTH(1-39). The phosphodiesterase inhibitor IBMX (100 microM) potentiated the responses to sub-maximal doses of ACTH(1-24). Two inhibitors of the protein kinase A (PKA) signaling pathway, Rp-cAMPS (500 microM) and H-89 (10 microM), abolished the insulin secretory response to ACTH(1-24) (0.5-10 nM). Treatment with 1 nM ACTH(1-24) caused a small, statistically significant increase in intracellular cAMP levels. Secretory responses of MIN6 cells to ACTH(1-24) were also influenced by changes in extracellular Ca2+ levels. Incubation in Ca2+-free buffer supplemented with 0.1 mM EGTA blocked the MIN6 cells' secretory response to 1 and 10 nM ACTH(1-24). Similar results were obtained when a Ca2+ channel blocker (nitrendipine, 10 microM) was added to the Ca2+-containing buffer. ACTH(1-24) also evoked an insulin secretory response from primary tIssues. The addition of ACTH(1-24) (0.5 nM) to perifusions of mouse islets induced a transient increase in insulin secretion at 8 mM glucose. Perifused human primary islets also showed a secretory response to ACTH(1-24) at basal glucose concentration (2 mM) with a rapid initial spike in insulin secretion followed by a decline to basal levels. Overall the results demonstrate that the MC2-R is expressed in beta-cells and suggest that activation of the receptor by ACTH initiates insulin secretion through the activation of PKA in association with Ca2+ influx into beta-cells.
Al-Majed
H T
HT
Centre for Reproduction, Endocrinology and Diabetes, GKT School of Biomedical Sciences, King's College London, Guy's Campus, London SE1 1UL, UK.
Jones
P M
PM
Persaud
S J
SJ
Sugden
D
D
Huang
G C
GC
Amiel
S
S
Whitehouse
B J
BJ
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002460:Cell Line; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007340:Insulinoma / Q000378:metabolism; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D007546:Isoquinolines / Q000494:pharmacology; D051379:Mice; D009568:Nitrendipine / Q000494:pharmacology; D019899:Paracrine Communication; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D044103:Receptor, Melanocortin, Type 2 / Q000235:genetics / Q000378:metabolism; D013268:Stimulation, Chemical; D013449:Sulfonamides; D013873:Thionucleotides / Q000494:pharmacology
D002121:Calcium Channel Blockers; D007328:Insulin; D007546:Isoquinolines; D010726:Phosphodiesterase Inhibitors; D012333:RNA, Messenger; D044103:Receptor, Melanocortin, Type 2; D013449:Sulfonamides; D013873:Thionucleotides; C016957:adenosine-3',5'-cyclic phosphorothioate; D000324:Adrenocorticotropic Hormone; D009568:Nitrendipine; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D015056:1-Methyl-3-isobutylxanthine