DIRECT EFFECTS OF DIAZOXIDE ON MITOCHONDRIA IN PANCREATIC B-CELLS ANDON ISOLATED LIVER-MITOCHONDRIA

1 The direct effects of diazoxide on mitochondrial membrane potential, Ca2+ transport, oxygen consumption and ATP generation were investigat ed in mouse pancreatic B-cells and rat liver mitochondria. 2 Diazoxide , at concentrations commonly used to open adenosine 5'-triphosphate (A TP)-dependent K+-channels (K-ATP channels) in pancreatic B-cells (100 to 1000 mu M), decreased mitochondrial membrane potential in mouse int act perifused B-cells, as evidenced by an increase of rhodamine 123 fl uorescence. This reversible decrease of membrane potential occurred at non-stimulating (5 mM) and stimulating (20 mM) glucose concentrations . 3 A decrease of mitochondrial membrane potential in perifused B-cell s was also caused by pinacidil, but no effect could be seen with levcr omakalim (500 mu M each). 4 Measurements by a tetraphenylphosphonium-s ensitive electrode of the membrane potential of rat isolated liver mit ochondria confirmed that diazoxide decreased mitochondrial membrane po tential by a direct action. Pretreatment with glibenclamide (2 mu M) d id not antagonize the effects of diazoxide. 5 In Fura 2-loaded B-cells perifused with the Ca2+ channel blocker, D 600, a moderate, reversibl e increase of intracellular Ca2+ concentration could be seen in respon se to 500 mu M diazoxide. This intracellular Ca2+ mobilization may be due to mitochondrial Ca2+ release, since the reduction of membrane pot ential of isolated liver mitochondria by diazoxide was accompanied by an accelerated release of Ca2+ stored in the mitochondria. 6 In the pr esence of 500 mu M diazoxide, ATP content of pancreatic islets incubat ed in 20 nM glucose for 30 min was significantly decreased by 29%. How ever, insulin secretion from mouse perifused islets induced by 40 mM K + in the presence of 10 mM glucose was not inhibited by 500 mu M diazo xide, suggesting that the energy-dependent processes of insulin secret ion distal to Ca2+ influx were not affected by diazoxide at this conce ntration. 7 The effects of diazoxide on oxygen consumption and ATP pro duction of liver mitochondria varied depending on the respiratory subs trates (5 mM succinate, 10 mM alpha-ketoisocaproic acid, 2 mM tetramet hyl phenylenediamine plus 5 mM ascorbic acid), indicating an inhibitio n of respiratory chain complex II. Pinacidil, but not levcromakalim, i nhibited alpha-ketoisocaproic acid-fuelled ATP production. 8 In conclu sion, diazoxide directly affects mitochondrial energy metabolism, whic h may be of relevance for stimulus-secretion coupling in pancreatic B- cells.