Effects of Glucose on Mitochondrial Function of Insulin-Producing INS-1 Insulinoma Cells

Abstract

BACKGROUND: Highly differentiated INS-1 832/13 cells are widely used as a model for glucose-stimulated insulin secretion (GSIS) similar to pancreatic beta cells. In the current view of GSIS, glucose metabolism leads to pyruvate formation, which is oxidized by mitochondria generating ATP. Mitochondrial ATP transported to the cytosol in exchange for cytosolic ADP via adenine nucleotide translocators (ANT) closes KATP channels. KATP channel closing causes plasma membrane depolarization which in turn opens voltage-dependent Ca2+ channels, triggering exocytosis of insulin granules. Our AIM was to evaluate mitochondrial function in INS-1 cells in relation to glucose stimulation. METHODS: Respiration of INS-1 cells incubated with 0, 3 or 15 mM glucose was determined in a Seahorse XF24. Mitochondrial and plasma membrane polarization was assessed by confocal microscopy of TMRM and DiBAC4, respectively. RESULTS: Glucose maximally increased respiration and insulin secretion at 15 mM. GSIS was blocked by rotenone, a mitochondrial respiratory chain inhibitor. Mitochondrial polarization was maintained in the absence of glucose and remained constant with increasing glucose to 15 mM. Blocking of mitochondrial ATP synthase by oligomicin inhibited respiration, depolarized mitochondria and hyperpolarized the plasma membrane. Mitochondrial depolarization after oligomycin was prevented by NIM811, an inhibitor of the mitochondrial permeability transition (MPT). Tolbutamide, a KATP channel blocker, reversed hyperpolarization of the plasma membrane. CONCLUSIONS: These findings indicate that mitochondrial function in INS-1 cells is preserved in the absence of glucose and that an increase of mitochondrial membrane potential is not required for GSIS.