Epac2‐dependent mobilization of intracellular Ca2+ by glucagon‐like peptide‐1 receptor agonist exendin‐4 is disrupted in β‐cells of phospholipase C‐ɛ knockout mice

Calcium can be mobilized in pancreatic β‐cells via a mechanism of Ca2+‐induced Ca2+ release (CICR), and cAMP‐elevating agents such as exendin‐4 facilitate CICR in β‐cells by activating both protein kinase A and Epac2. Here we provide the first report that a novel phosphoinositide‐specific phospholipase C‐ɛ (PLC‐ɛ) is expressed in the islets of Langerhans, and that the knockout (KO) of PLC‐ɛ gene expression in mice disrupts the action of exendin‐4 to facilitate CICR in the β‐cells of these mice. Thus, in the present study, in which wild‐type (WT) C57BL/6 mouse β‐cells were loaded with the photolabile Ca2+ chelator NP‐EGTA, the UV flash photolysis‐catalysed uncaging of Ca2+ generated CICR in only 9% of the β‐cells tested, whereas CICR was generated in 82% of the β‐cells pretreated with exendin‐4. This action of exendin‐4 to facilitate CICR was reproduced by cAMP analogues that activate protein kinase A (6‐Bnz‐cAMP‐AM) or Epac2 (8‐pCPT‐2′‐O‐Me‐cAMP‐AM) selectively. However, in β‐cells of PLC‐ɛ KO mice, and also Epac2 KO mice, these test substances exhibited differential efficacies in the CICR assay such that exendin‐4 was partly effective, 6‐Bnz‐cAMP‐AM was fully effective, and 8‐pCPT‐2′‐O‐Me‐cAMP‐AM was without significant effect. Importantly, transduction of PLC‐ɛ KO β‐cells with recombinant PLC‐ɛ rescued the action of 8‐pCPT‐2′‐O‐Me‐cAMP‐AM to facilitate CICR, whereas a K2150E PLC‐ɛ with a mutated Ras association (RA) domain, or a H1640L PLC‐ɛ that is catalytically dead, were both ineffective. Since 8‐pCPT‐2′‐O‐Me‐cAMP‐AM failed to facilitate CICR in WT β‐cells transduced with a GTPase activating protein (RapGAP) that downregulates Rap activity, the available evidence indicates that a signal transduction ‘module’ comprised of Epac2, Rap and PLC‐ɛ exists in β‐cells, and that the activities of Epac2 and PLC‐ɛ are key determinants of CICR in this cell type.