TY - JOUR
T1 - The mitochondrial Ca2+ uniporter MCU is essential for glucose-induced atp increases in pancreatic β-cells
AU - Tarasov, Andrei I.
AU - Semplici, Francesca
AU - Ravier, Magalie A.
AU - Bellomo, Elisa A.
AU - Pullen, Timothy J.
AU - Gilon, Patrick
AU - Sekler, Israel
AU - Rizzuto, Rosario
AU - Rutter, Guy A.
PY - 2012/7/19
Y1 - 2012/7/19
N2 - Glucose induces insulin release from pancreatic β-cells by stimulating ATP synthesis, membrane depolarisation and Ca2+ influx. As well as activating ATP-consuming processes, cytosolic Ca2+ increases may also potentiate mitochondrial ATP synthesis. Until recently, the ability to study the role of mitochondrial Ca2+ transport in glucose-stimulated insulin secretion has been hindered by the absence of suitable approaches either to suppress Ca2+ uptake into these organelles, or to examine the impact on β-cell excitability. Here, we have combined patch-clamp electrophysiology with simultaneous real-time imaging of compartmentalised changes in Ca2+ and ATP/ADP ratio in single primary mouse β-cells, using recombinant targeted (Pericam or Perceval, respectively) as well as entrapped intracellular (Fura-Red), probes. Through shRNA-mediated silencing we show that the recently-identified mitochondrial Ca2+ uniporter, MCU, is required for depolarisation-induced mitochondrial Ca2+ increases, and for a sustained increase in cytosolic ATP/ADP ratio. By contrast, silencing of the mitochondrial Na+-Ca2+ exchanger NCLX affected the kinetics of glucose-induced changes in, but not steady state values of, cytosolic ATP/ADP. Exposure to gluco-lipotoxic conditions delayed both mitochondrial Ca2+ uptake and cytosolic ATP/ADP ratio increases without affecting the expression of either gene. Mitochondrial Ca2+ accumulation, mediated by MCU and modulated by NCLX, is thus required for normal glucose sensing by pancreatic β-cells, and becomes defective in conditions mimicking the diabetic milieu.
AB - Glucose induces insulin release from pancreatic β-cells by stimulating ATP synthesis, membrane depolarisation and Ca2+ influx. As well as activating ATP-consuming processes, cytosolic Ca2+ increases may also potentiate mitochondrial ATP synthesis. Until recently, the ability to study the role of mitochondrial Ca2+ transport in glucose-stimulated insulin secretion has been hindered by the absence of suitable approaches either to suppress Ca2+ uptake into these organelles, or to examine the impact on β-cell excitability. Here, we have combined patch-clamp electrophysiology with simultaneous real-time imaging of compartmentalised changes in Ca2+ and ATP/ADP ratio in single primary mouse β-cells, using recombinant targeted (Pericam or Perceval, respectively) as well as entrapped intracellular (Fura-Red), probes. Through shRNA-mediated silencing we show that the recently-identified mitochondrial Ca2+ uniporter, MCU, is required for depolarisation-induced mitochondrial Ca2+ increases, and for a sustained increase in cytosolic ATP/ADP ratio. By contrast, silencing of the mitochondrial Na+-Ca2+ exchanger NCLX affected the kinetics of glucose-induced changes in, but not steady state values of, cytosolic ATP/ADP. Exposure to gluco-lipotoxic conditions delayed both mitochondrial Ca2+ uptake and cytosolic ATP/ADP ratio increases without affecting the expression of either gene. Mitochondrial Ca2+ accumulation, mediated by MCU and modulated by NCLX, is thus required for normal glucose sensing by pancreatic β-cells, and becomes defective in conditions mimicking the diabetic milieu.
UR - http://www.scopus.com/inward/record.url?scp=84864070892&partnerID=8YFLogxK
U2 - 10.1371/journal.pone.0039722
DO - 10.1371/journal.pone.0039722
M3 - Article
C2 - 22829870
AN - SCOPUS:84864070892
SN - 1932-6203
VL - 7
JO - PLoS ONE
JF - PLoS ONE
IS - 7
M1 - e39722
ER -