title: P2Y₁ receptor-dependent diacylglycerol signaling microdomains in β cells promote insulin secretion.
abstract: Diacylglycerol (DAG) controls numerous cell functions by regulating the localization of C1-domain-containing proteins, including protein kinase C (PKC), but little is known about the spatiotemporal dynamics of the lipid. Here, we explored plasma membrane DAG dynamics in pancreatic β cells and determined whether DAG signaling is involved in secretagogue-induced pulsatile release of insulin. Single MIN6 cells, primary mouse β cells, and human β cells within intact islets were transfected with translocation biosensors for DAG, PKC activity, or insulin secretion and imaged with total internal reflection fluorescence microscopy. Muscarinic receptor stimulation triggered stable, homogenous DAG elevations, whereas glucose induced short-lived (7.1 ± 0.4 s) but high-amplitude elevations (up to 109 ± 10% fluorescence increase) in spatially confined membrane regions. The spiking was mimicked by membrane depolarization and suppressed after inhibition of exocytosis or of purinergic P2Y₁, but not P2X receptors, reflecting involvement of autocrine purinoceptor activation after exocytotic release of ATP. Each DAG spike caused local PKC activation with resulting dissociation of its substrate protein MARCKS from the plasma membrane. Inhibition of spiking reduced glucose-induced pulsatile insulin secretion. Thus, stimulus-specific DAG signaling patterns appear in the plasma membrane, including distinct microdomains, which have implications for the kinetic control of exocytosis and other membrane-associated processes.
authors.affiliation: Department of Medical Cell Biology, Uppsala University, Biomedical Centre, Uppsala, Sweden.
authors.forename: Anne
authors.initials: A
authors.lastname: Wuttke
authors.forename: Olof
authors.initials: O
authors.lastname: Idevall-Hagren
authors.forename: Anders
authors.initials: A
authors.lastname: Tengholm
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004075:Diglycerides / Q000378:metabolism*; D005089:Exocytosis / Q000187:drug effects; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D051379:Mice; D011493:Protein Kinase C / Q000378:metabolism*; D058472:Receptors, Purinergic P2Y1 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
chemical_list: C011439:1,2-diacylglycerol; D004075:Diglycerides; D007328:Insulin; D058472:Receptors, Purinergic P2Y1; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D005947:Glucose; D002118:Calcium