Molecular mechanisms of muscarinic acetylcholine receptor-stimulated increase in cytosolic free Ca2+ concentration and ERK1/2 activation in the MIN6 pancreatic b-cell line

Abstract Muscarinic acetylcholine receptor (mAChR) activation of pancreatic b-cells elevates intracellular Ca2? and potentiates glucose-stimulated insulin secretion. In addition, it activates a number of signaling molecules, including ERK1/2, whose activation has been shown to play an important role in regulating pancreatic b-cell function and mass. The aim of this work was to determine how mAChR activation elevates intracellular Ca2? concentration ([Ca2?]i) and activates ERK1/2 in the pancreatic b-cell line MIN6.We demonstrate that agonist-stimulated ERK1/2 activation is dependent on the activation of phospholipase C and an elevation in [Ca2?]i, but is independent of the activation of diacylglycerol-dependent protein kinase C isoenzymes. Using a pharmacological approach, we provide evidence that agonist-induced increases in [Ca2?]i and ERK activity require (1) IP3 receptor-mediated mobilization of Ca2? from the endoplasmic reticulum, (2) influx of extracellular Ca2? through store-operated channels, (3) closure of KATP channels, and (4) Ca2? entry via L-type voltage-operated Ca2? channels. Moreover, this Ca2?-dependent activation of ERK is mediated via both Ras-dependent and Ras-independent mechanisms. In summary, this study provides important insights into the multifactorial signaling mechanisms linking mAChR activation to increases in [Ca2?]i and ERK activity.