Fuel-induced amplification of insulin secretion in mouse pancreatic islets exposed to a high sulfonylurea concentration: role of the NADPH/NADP+ ratio

Abstract

Aims/hypothesis The aim of this study was to examine whether the cytosolic NADPH/NADP + ratio of beta cells serves as an amplifying signal in fuel-induced insulin secretion and whether such a function is mediated by cytosolic α-ketoglutarate. Methods Pancreatic islets and islet cells were isolated from albino mice by collagenase digestion. Insulin secretion of incubated or perifused islets was measured by ELISA. The NADPH and NADP + content of incubated islets was determined by enzymatic cycling. The cytosolic Ca 2+ concentration ([Ca 2+ ] c ) in islets was measured by microfluorimetry and the activity of ATP-sensitive K + channels in islet cells by patch-clamping. Results Both 30 mmol/l glucose and 10 mmol/l α-ketoisocaproate stimulated insulin secretion and elevated the NADPH/NADP + ratio of islets preincubated in the absence of fuel. The increase in the NADPH/NADP + ratio was abolished in the presence of 2.7 μmol/l glipizide (closing all ATP-sensitive K + channels). However, α-ketoisocaproate, but not glucose, still stimulated insulin secretion. That glipizide did not inhibit α-ketoisocaproate-induced insulin secretion was not the result of elevated [Ca 2+ ] c , as glucose caused a more marked [Ca 2+ ] c increase. Insulin release triggered by glipizide alone was moderately amplified by dimethyl α-ketoglutarate (which is cleaved to produce cytosolic α-ketoglutarate), but there was no indication of a signal function of cytosolic α-ketoglutarate. Conclusions/interpretationThe results strongly suggest that the NADPH/NADP + ratio in the beta cell cytosol does not serve as an amplifying signal in fuel-induced insulin release. The study supports the view that amplification results from the intramitochondrial production of citrate by citrate synthase and from the associated export of citrate into the cytosol.

Regulation of insulin secretion: a matter of phase control and amplitude modulation

“…1), but other messengers could also be involved [8,34,48]. Recent studies call for a (re)evaluation of the roles of cAMP [49], NADPH [48, 50, 51], citrate cycle intermediates exported to the cytosol [48, 51,52], AMP-activated protein kinase [53] and granule translocation by the cytoskeleton [18,54]. Mathematical modelling of the amplifying pathway may also help to identify the effector system [55,56].…”

Hexose-6-phosphate dehydrogenase: linking endocrinology and metabolism in the endoplasmic reticulum

“…(Dzyakanchuk et al 2008). It means that the in vivo set point of the NADPH-NADP C redox couple is much more reduced in the ER lumen than in the cytosol (Díaz-Flores et al 2006, Pollak et al 2007, Panten & Rustenbeck 2008. H6PD is more resistant to feedback inhibition by NADPH than its cytosolic counterpart glucose-6-phosphate dehydrogenase (Oka et al 1981), which allows the almost complete reduction of the ER luminal pyridine nucleotide pool.…”

Role of metabolically generated reactive oxygen species for lipotoxicity in pancreatic β‐cells

“…The metabolism of fatty acids is apparently of subordinate importance as a site of generation of a specific metabolic signal for fatty acid-mediated insulin secretion. It may be possible, however, that metabolites generated within mitochondrial β-oxidation and subsequent citric acid cycle, as assumed for the potentiation of glucose-induced insulin secretion through the amplifying pathway [32][33][34][35], contribute in a collaborative manner to the insulin-secretory capacity of fatty acids.In a sense the mechanism of fatty acid potentiation resembles the mechanisms of other potentiators of glucose-induced insulin secretion such as acetylcholine, bombesin or arginine vasopressin, which mediate their effects via interaction with a receptor in the β-cell plasma membrane coupled to signalling pathways that increase intracellular [Ca 2+ ] i through mobilization from the ER and increased Ca 2+ influx through L-type Ca 2+ channels [25]. In particular, in the phase of insulin resistance before diabetes manifestation, when an increased β-cell , by contrast, they enter the mitochondria in a carnitine-dependent manner catalysed by carnitine palmitoyltransferase (CPT).…”