Abstract
BackgroundMany mechanistic steps underlying nutrient-stimulated insulin secretion (NSIS) are poorly understood. The influence of intracellular pH (pHi) on insulin secretion is widely documented, and can be used as an investigative tool. This study demonstrates previously unknown effects of pHi-alteration on insulin secretion in mouse islets, which may be utilized to correct defects in insulin secretion.MethodsDifferent components of insulin secretion in mouse islets were monitored in the presence and absence of forced changes in pHi. The parameters measured included time-dependent potentiation of insulin secretion by glucose, and direct insulin secretion by different mitochondrial and non-mitochondrial secretagogues. Islet pHi was altered using amiloride, removal of medium Cl-, and changing medium pH. Resulting changes in islet pHi were monitored by confocal microscopy using a pH-sensitive fluorescent indicator. To investigate the underlying mechanisms of the effects of pHi-alteration, cellular NAD(P)H levels were measured using two-photon excitation microscopy (TPEM). Data were analyzed using Student's t test.ResultsTime-dependent potentiation, a function normally absent in mouse islets, can be unmasked by a forced decrease in pHi. The optimal range of pHi for NSIS is 6.4–6.8. Bringing islet pHi to this range enhances insulin secretion by all mitochondrial fuels tested, reverses the inhibition of glucose-stimulated insulin secretion (GSIS) by mitochondrial inhibitors, and is associated with increased levels of cellular NAD(P)H.ConclusionsPharmacological alteration of pHi is a potential means to correct the secretory defect in non-insulin dependent diabetes mellitus (NIDDM), since forcing islet pHi to the optimal range enhances NSIS and induces secretory functions that are normally absent.
Highlights
Many mechanistic steps underlying nutrient-stimulated insulin secretion (NSIS) are poorly understood
In this study, using a more accurate modern technique of pHi-measurement [35], we have investigated the effects of pHi on different components of NSIS in mouse islets, and explored the possible mechanisms of such effects
All groups were first pre-incubated in basal glucose for one hour, and subsequently exposed to each condition denoted in the X axis for one hour, followed by sample collection for insulin assay
Summary
Many mechanistic steps underlying nutrient-stimulated insulin secretion (NSIS) are poorly understood. Nutrient-stimulated insulin secretion (NSIS) in the pancreatic β cell consists of three distinct components with different underlying mechanisms These components include a) an initial peak (first phase) triggered by Ca2+, b) augmentation of the Ca2+-triggered response (second phase), and c) a memory that persists after removal of the nutrient, causing enhancement of subsequent secretory responses (time-dependent potentiation) [1,2,3,4]. BMC Endocrine Disorders 2004, 4 http://www.biomedcentral.com/1472-6823/4/1 from glucose metabolism, which leads to membrane depolarization through closure of ATP-dependent K+ channels (KATP channels) and consequent entry of extracellular Ca2+ through voltage-gated Ca2+ channels This influx of Ca2+ triggers the release of a small pool of secretory granules, producing the initial peak of the insulin response [1]. Time-dependent potentiation (TDP), a positive memory induced during this acute response, magnifies subsequent secretory responses to all secretagogues [2,4,5,6]
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