Palmitate potentiation of glucose-induced insulin release: a study using 2-bromopalmitate.
ABSTRACT The mechanisms whereby fatty acids (FA) potentiate glucose-induced insulin secretion from the pancreatic beta cell are incompletely understood. In this study, the effects of palmitate on insulin secretion were investigated in isolated rat islets. Palmitate did not initiate insulin secretion at nonstimulatory glucose concentrations, but markedly stimulated insulin release at concentrations of glucose > or = 5.6 mmol/L. At concentrations of palmitate > or =0.5 mmol/L, the important determinant of the potency of the FA was its unbound concentration. At total concentrations < or = 0.5 mmol/L, both the total and unbound concentrations appeared important. Surprisingly, 2-bromopalmitate did not affect palmitate oxidation, but significantly diminished palmitate esterification into cellular lipids. Neither methyl palmitate, which is not activated into a long-chain acyl-CoA ester, nor 2-bromopalmitate affected glucose-stimulated insulin release. Further, 2-bromopalmitate partly inhibited the potentiating effect of palmitate. These results support the concept that FA potentiation of insulin release is mediated by FA-derived signals generated in the esterification pathway.
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ABSTRACT: Exposure of insulin-producing cells to elevated levels of the free fatty acid (FFA) palmitate results in the loss of β-cell function and induction of apoptosis. The induction of endoplasmic reticulum (ER) stress is one mechanism proposed to be responsible for the loss of β-cell viability in response to palmitate treatment; however, the pathways responsible for the induction of ER stress by palmitate have yet to be determined. Protein palmitoylation is a major posttranslational modification that regulates protein localization, stability, and activity. Defects in, or dysregulation of, protein palmitoylation could be one mechanism by which palmitate may induce ER stress in β-cells. The purpose of this study was to evaluate the hypothesis that palmitate-induced ER stress and β-cell toxicity are mediated by excess or aberrant protein palmitoylation. In a concentration-dependent fashion, palmitate treatment of RINm5F cells results in a loss of viability. Similar to palmitate, stearate also induces a concentration-related loss of RINm5F cell viability, while the monounsaturated fatty acids, such as palmoleate and oleate, are not toxic to RINm5F cells. 2-Bromopalmitate (2BrP), a classical inhibitor of protein palmitoylation that has been extensively used as an inhibitor of G protein-coupled receptor signaling, attenuates palmitate-induced RINm5F cell death in a concentration-dependent manner. The protective effects of 2BrP are associated with the inhibition of [(3)H]palmitate incorporation into RINm5F cell protein. Furthermore, 2BrP does not inhibit, but appears to enhance, the oxidation of palmitate. The induction of ER stress in response to palmitate treatment and the activation of caspase activity are attenuated by 2BrP. Consistent with protective effects on insulinoma cells, 2BrP also attenuates the inhibitory actions of prolonged palmitate treatment on insulin secretion by isolated rat islets. These studies support a role for aberrant protein palmitoylation as a mechanism by which palmitate enhances ER stress activation and causes the loss of insulinoma cell viability.AJP Endocrinology and Metabolism 03/2012; 302(11):E1390-8. · 4.51 Impact Factor
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ABSTRACT: Free fatty acids (FFAs) exert complex actions on pancreatic β-cells. Typically, an initial potentiation of insulin release is followed by a gradual impairment of β-cell function, the latter effect being of possible relevance to hyperlipidaemia in type 2 diabetes mellitus. The molecular actions of FFAs are poorly understood. The present study investigated the acute effects of saturated FFAs on electrophysiological responses of rat pancreatic β-cells. Membrane potential and KATP channel activity were recorded using the perforated patch technique. Volume-regulated anion channel (VRAC) activity was assessed from conventional whole-cell recordings. Cell volume regulation was measured using a video-imaging technique. Addition of octanoate caused a transient potentiation of glucose-induced electrical activity, followed by a gradual hyper-polarisation and a prolonged inhibition of electrical activity. Octanoate caused an initial increase in VRAC activity followed by a secondary inhibition coinciding with increased KATP channel activity. Similar effects were observed with palmitate and 2-bromopalmitate whereas butyrate was virtually ineffective. Octanoate and palmitate also exerted a dual effect on electrical activity evoked by tolbutamide. Octanoate significantly attenuated cell volume regulation in hypotonic solutions, consistent with VRAC inhibition. It is concluded that medium and long chain FFAs have a dual action on glucose-induced electrical activity in rat pancreatic β-cells: an initial stimulatory effect followed by a secondary inhibition. These effects appear to be the result of reciprocal actions on VRAC and KATP channel currents, and could contribute towards the stimulatory and inhibitory actions of FFAs on pancreatic β-cell function.The Journal of Physiology 01/2011; 589(Pt 6):1307-16. · 4.38 Impact Factor
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ABSTRACT: Hepatic gluconeogenesis tightly controls blood glucose levels in healthy individuals, yet disorders of fatty acids (FAs) oxidation are characterized by hypoglycemia. We studied the ability of free-FAs to directly inhibit gluconeogenesis, as a novel mechanism that elucidates the hypoglycemic effect of FAs oxidation defects. Primary rat hepatocytes were pre-treated with FAs prior to gluconeogenic stimuli with glucagon or dexamethasone and cAMP. Pre-treatment with 1 mM FAs (mixture of 2:1 oleate:palmitate) for 1 hour prior to gluconeogenic induction, significantly decreases the induced expression of the gluconeogenic genes phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6pase) as well as the induced glucose production by the cells. The inhibitory effect of FAs upon gluconeogenesis is abolished when pre-treatment is elongated to 18 hours, allowing clearance of FAs into triglycerides by the cells. Replacement of palmitate with the non-metabolic fatty acid 2-bromopalmitate inhibits esterification of FAs into triglycerides. Accordingly, the increased exposure to unesterified-FAs allows their inhibitory effect to be extended even when pre-treatment is elongated to 18 hours. Similar changes were caused by FAs to the induction of peroxisome-proliferator-activated receptor-γ coactivator 1α (PGC1α) expression, indicating this transcriptional coactivator as the mediating link of the effect. This inhibitory effect of FAs upon gluconeogenic induction is shown to involve reduced activation of cAMP response element-binding (CREB) transcription factor. The present results demonstrate that free-FAs directly inhibit the induced gluconeogenic response in hepatocytes. Hence, high levels of free-FAs may attenuate hepatic gluconeogenesis, and liver glucose output.Lipids in Health and Disease 06/2012; 11:66. · 2.31 Impact Factor