cAMP-dependent mobilization of intracellular Ca2+ stores by activation of ryanodine receptors in pancreatic beta-cells - A Ca2+ signaling system stimulated by the insulinotropic hormone glucagon-like peptide-1-(7-37)

Glucagon-like peptide-1 (GLP-1) is an intestinally derived insulinotropic h ormone currently under investigation for use as a novel therapeutic agent i n the treatment of type 2 diabetes mellitus, In vitro studies of pancreatic islets of Langerhans demonstrated that GLP-1 interacts with specific beta- cell G protein-coupled receptors, thereby facilitating insulin exocytosis b y raising intracellular levels of cAMP and Ca2+. Here we report that the st imulatory influence of GLP-1 on Ca2+ signaling results, in part, from cAMP- dependent mobilization of ryanodine-sensitive Ca2+ stores. Studies of human , rat, and mouse beta-cells demonstrate that the binding of a fluorescent d erivative of ryanodine (BODIPY FL-X ryanodine) to its receptors is specific , reversible, and of high affinity. Rat islets and BTC3 insulinoma cells ar e shown by reverse transcriptase polymerase chain reaction analyses to expr ess mRNA corresponding to the type 2 isoform of ryanodine receptor-intracel lular Ca2+ release channel (RYR2), Single-cell measurements of [Ca2+](i) us ing primary cultures of rat and human beta-cells indicate that GLP-1 facili tates Ca2+-induced Ca2+ release (CICR), whereby mobilization of Ca2+ stores is triggered by influx of Ca2+ through L-type Ca2+ channels, In these cell s, GLP-1 is shown to interact with metabolism of D-glucose to produce a fas t transient increase of [Ca2+](i). This effect is reproduced by 8-Br-cAMP, but is blocked by a GLP-1 receptor antagonist (exendin-(9-39)), a cAMP anta gonist ((Rp)-cAMPS), an L-type Ca2+ channel antagonist (nimodipine), an ant agonist of the sarco(endo)plasmic reticulum Ca2+ ATPase (thapsigargin), or by ryanodine, Characterization of the CICR mechanism by voltage clamp analy sis also demonstrates a stimulation of Ca2+ release by caffeine. These find ings provide new support for a model of beta-cell signal transduction where by GLP-1 promotes CICR by sensitizing intracellular Ca2+ release channels t o the stimulatory influence of cytosolic Ca2+.