title: Frequency-dependent mitochondrial Ca(2+) accumulation regulates ATP synthesis in pancreatic β cells.
abstract: Pancreatic β cells respond to increases in glucose concentration with enhanced metabolism, the closure of ATP-sensitive K(+) channels and electrical spiking. The latter results in oscillatory Ca(2+) influx through voltage-gated Ca(2+) channels and the activation of insulin release. The relationship between changes in cytosolic and mitochondrial free calcium concentration ([Ca(2+)]cyt and [Ca(2+)]mit, respectively) during these cycles is poorly understood. Importantly, the activation of Ca(2+)-sensitive intramitochondrial dehydrogenases, occurring alongside the stimulation of ATP consumption required for Ca(2+) pumping and other processes, may exert complex effects on cytosolic ATP/ADP ratios and hence insulin secretion. To explore the relationship between these parameters in single primary β cells, we have deployed cytosolic (Fura red, Indo1) or green fluorescent protein-based recombinant-targeted (Pericam, 2mt8RP for mitochondria; D4ER for the ER) probes for Ca(2+) and cytosolic ATP/ADP (Perceval) alongside patch-clamp electrophysiology. We demonstrate that: (1) blockade of mitochondrial Ca(2+) uptake by shRNA-mediated silencing of the uniporter MCU attenuates glucose- and essentially blocks tolbutamide-stimulated, insulin secretion; (2) during electrical stimulation, mitochondria decode cytosolic Ca(2+) oscillation frequency as stable increases in [Ca(2+)]mit and cytosolic ATP/ADP; (3) mitochondrial Ca(2+) uptake rates remained constant between individual spikes, arguing against activity-dependent regulation ("plasticity") and (4) the relationship between [Ca(2+)]cyt and [Ca(2+)]mit is essentially unaffected by changes in endoplasmic reticulum Ca(2+) ([Ca(2+)]ER). Our findings thus highlight new aspects of Ca(2+) signalling in β cells of relevance to the actions of both glucose and sulphonylureas.
authors.affiliation: Section of Cell Biology, Division of Diabetes Endocrinology and Metabolism, Department of Medicine, Imperial College London, SW7 2AZ, London, UK.
authors.forename: Andrei I
authors.initials: AI
authors.lastname: Tarasov
authors.forename: Francesca
authors.initials: F
authors.lastname: Semplici
authors.forename: Daliang
authors.initials: D
authors.lastname: Li
authors.forename: Rosario
authors.initials: R
authors.lastname: Rizzuto
authors.forename: Magalie A
authors.initials: MA
authors.lastname: Ravier
authors.forename: Patrick
authors.initials: P
authors.lastname: Gilon
authors.forename: Guy A
authors.initials: GA
authors.lastname: Rutter
mesh_terms: D000200:Action Potentials; D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000096:biosynthesis*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005260:Female; D005947:Glucose / Q000378:metabolism; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D007328:Insulin; C034703:mitochondrial calcium uniporter; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D005947:Glucose; D002118:Calcium