title: Nicotinic acid-adenine dinucleotide phosphate-sensitive calcium stores initiate insulin signaling in human beta cells.
abstract: Recent studies suggest a role for autocrine insulin signaling in beta cells, but the mechanism and function of insulin-stimulated Ca(2+) signals is uncharacterized. We examined Ca(2+)-dependent insulin signaling in human beta cells. Two hundred nanomolar insulin elevated [Ca(2+)](c) to 284 +/- 27 nM above baseline in approximately 30% of Fura-4F-loaded cells. Insulin evoked multiple Ca(2+) signal waveforms, 60% of which included oscillations. Although the amplitude of Ca(2+) signals was dose-dependent between 0.002 and 2,000 nM, the percentage of cells responding was highest at 0.2 nM insulin, suggesting the interaction of stimulatory and inhibitory pathways. Ca(2+)-free solutions did not affect the initiation of insulin-stimulated Ca(2+) signals, but abolished the second phase of plateaus/oscillations. Likewise, inositol 1,4,5-trisphosphate (IP(3)) receptor antagonists xestospongin C and caffeine selectively blocked the second phase, but not the initiation of insulin signaling. Thapsigargin and 2,5-di-tert-butylhydroquinone (BHQ) blocked insulin signaling, implicating sarcoplasmic/endoplasmic Ca(2+)-ATPase (SERCA)-containing Ca(2+) stores. Insulin-stimulated Ca(2+) signals were insensitive to ryanodine. Injection of the CD38-derived Ca(2+) mobilizing metabolite, nicotinic acid-adenine dinucleotide phosphate (NAADP), at nanomolar concentrations, evoked oscillatory Ca(2+) signals that could be initiated in the presence of ryanodine, xestospongin C, and Ca(2+)-free solutions. Desensitizing concentrations of NAADP abolished insulin-stimulated Ca(2+) signals. Insulin-stimulated Ca(2+) signals led to a Ca(2+)-dependent increase in cellular insulin contents, but not secretion. These data reveal the complexity of insulin signal transduction and function in human beta cells and demonstrate functional NAADP-sensitive Ca(2+) stores in a human primary cultured cell type.
authors.affiliation: Department of Internal Medicine, Renal Division, Barnes-Jewish Hospital, St. Louis, MO 63110, USA. jim@jimjohnson.ca
authors.forename: James D
authors.initials: JD
authors.lastname: Johnson
authors.forename: Stanley
authors.initials: S
authors.lastname: Misler
mesh_terms: D002110:Caffeine / Q000494:pharmacology; D015220:Calcium Channels; D020013:Calcium Signaling / Q000187:drug effects*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D004594:Electrophysiology; D006801:Humans; D006873:Hydroquinones / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin / Q000378:metabolism* / Q000494:pharmacology; D007424:Intracellular Fluid / Q000378:metabolism; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism; D047028:Macrocyclic Compounds; D009249:NADP / Q000031:analogs & derivatives* / Q000494:pharmacology*; D010080:Oxazoles / Q000494:pharmacology; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D015220:Calcium Channels; D006873:Hydroquinones; C506926:ITPR1 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin; D047028:Macrocyclic Compounds; D010080:Oxazoles; D018160:Receptors, Cytoplasmic and Nuclear; C108644:xestospongin A; C019359:2,5-di-tert-butylhydroquinone; D002110:Caffeine; D009249:NADP; C024376:NAADP; D019284:Thapsigargin; D000252:Calcium-Transporting ATPases