Direct Substrate Delivery Into Mitochondrial Fission–Deficient Pancreatic Islets Rescues Insulin Secretion

Abstract

In pancreatic b-cells, mitochondrial bioenergetics control glucose-stimulated insulin secretion. Mitochondrial dynamics are generally associated with quality control, maintaining the functionality of bioenergetics. By acute pharmacological inhibition of mitochondrial fission protein Drp1, we demonstrate in this study that mitochondrial fission is necessary for glucose-stimulated insulin secretion in mouse and human islets. We confirm that genetic silencing of Drp1 increases mitochondrial proton leak in MIN6 cells. However, our comprehensive analysis of pancreatic islet bioenergetics reveals that Drp1 does not control insulin secretion via its effect on proton leak but instead via modulation of glucose-fueled respiration. Notably, pyruvate fully rescues the impaired insulin secretion of fission-deficient b-cells, demonstrating that defective mitochondrial dynamics solely affect substrate supply upstream of oxidative phosphorylation. The present findings provide novel insights into how mitochondrial dysfunction may cause pancreatic b-cell failure. In addition, the results will stimulate new thinking in the intersecting fields of mitochondrial dynamics and bioenergetics, as treatment of defective dynamics in mitochondrial diseases appears to be possible by improving metabolism upstream of mitochondria.The development of type 2 diabetes is associated with mitochondrial dysfunction in pancreatic b-cells. b-Cells have developed highly coordinated mechanisms that link glucose sensing with metabolic signaling cascades that direct the secretion of sufficient insulin to maintain glucose homeostasis (1). The mechanism underlying glucose-stimulated insulin secretion (GSIS) from b-cells involves glucose uptake by specific glucose transporters followed by glucose catabolism through glycolysis yielding pyruvate, which in turn is further catabolized via the tricarboxylic acid cycle. Both glycolysis and tricarboxylic acid cycle turnover generate reducing power that is used by the mitochondrial respiratory chain to produce ATP through oxidative phosphorylation. The resultant increase in the cytosolic ATP/ADP ratio closes ATP-sensitive K + (K ATP ) channels, thus depolarizing the plasma membrane. Depolarization triggers opening of voltagegated Ca 2+ channels, and the influx of Ca 2+ ions consequently leads to exocytosis of insulin-containing granules from b-cells (2). This sequence of events illustrates that b-cell mitochondria exert strong control over GSIS (3), as mitochondrial energy-transducing processes dictate how fast and efficiently glucose is converted to ATP. Mitochondrial dysfunction impairs glucose-insulin secretion coupling and ultimately promotes b-cell apoptosis and death, one of the key features of type 2 diabetes (4).Mitochondria are dynamic organelles that are constantly remodeled through the opposing action of fission and fusion proteins (5). Until recently, the influence of mitochondrial dynamics over energy transduction has been underappreciated. The continuous changes in mitochondrial morphology are controlle...

Control of pancreatic β-cell bioenergetics

“…This notion is further supported by the beta cells' responsiveness to pyruvate [33,36,41] that can enter insulinoma cells when added as sodium pyruvate and primary beta cells when given as methyl-pyruvate. We find that INS-1E cells increase their respiratory activity to the same extent in response to 5 mM pyruvate as to 20 mM glucose (Fig.…”

“…Beta cell respiratory analyses generally offer clear evidence for the strong control of ATP flux by ATP supply: basal oxygen uptake in insulinoma cells as well as in rodent and human islets is increased 1.5 to 3-fold by glucose [21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36] . From oligomycin-sensitive respiration we calculate that glucose raises the ATP supply flux in fuel-deprived INS-1E cells up to almost 6 times ( Fig.…”

“…2A). The glucose effect on ATP supply is comparably strong because most of the respiratory rise is coupled to phosphorylation [25,36] , whilst the proportion of basal respiration used to make ATP is exceptionally low [28,33,[37][38][39] . Glucose increases ATP-synthesis-coupled respiration more than it raises respiration linked to mitochondrial proton leak, which means it stimulates both the rate and coupling efficiency of oxidative phosphorylation [26,36] .…”

Glycemic Variability Promotes Both Local Invasion and Metastatic Colonization by Pancreatic Ductal Adenocarcinoma

“…Mitochondrial respiration was determined by oxygen consumption rate, and glycolysis rate was determined by extracellular acidification rate as previously described. 40 Briefly, 2 × 10 4 hypoglycemic or hyperglycemic cells were seeded in XF96 culture microplates (101085-004; Agilent Technologies Deutschland GmbH, Waldbronn, Germany) overnight at cultivation condition. For basal cellular respiration measurement, cells were treated sequentially with 2,4-DNP (0.1 mmol/L), 2DG (100 mmol/L), oligomycin (1.5 μg/mL), and eventually rotenone/antimycin A (2.5 μmol/L and 2.5 μmol/L, respectively).…”

Cellular models for beta‐cell function and diabetes gene therapy

“…These cells are being widely used for preliminary investigations on beta‐cell biology and drug targets. For example, recent investigations include exploration of dopamine receptor signalling, lipidomic profiling, antidiabetic actions of dogfish glucagon analogues, high‐throughput screening for inhibitors of beta‐cell apoptosis, Ca 2+ channel clustering with insulin granules in diabetes, mitochondrial dynamics and bioenergetics as well as involvement of neuronal K + Cl − cotransporter in insulin secretion . These investigations lay a foundation for further studies on understanding of beta‐cell biology in health and disease.…”