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Free fatty acids (FFAs) have pleiotropic effects on the pancreatic β-cell. Although acute exposure to FFAs stimulates glucose-stimulated insulin secretion (GSIS), prolonged exposure impairs GSIS and causes apoptosis. FFAs exert their effects both via intracellular metabolism and interaction with the FFA receptor 1 (FFAR1/GPR40). Here we studied the role of FFAR1 in acute and long-term effects of palmitate on GSIS and insulin content in isolated human islets by using the FFAR1 agonist TAK-875 and the antagonist ANT203. Acute palmitate exposure potentiated GSIS approximately 3-fold, whereas addition of the antagonist decreased this potentiation to approximately 2-fold. In the absence of palmitate, the agonist caused a 40% increase in GSIS. Treatment with palmitate for 7 days decreased GSIS to 70% and insulin content to 25% of control level. These negative effects of long-term exposure to palmitate were ameliorated by FFAR1 inhibition and further aggravated by additional stimulation of the receptor. In the absence of extracellularly applied palmitate, long-term treatment with the agonist caused a modest increase in GSIS. The protective effect of FFAR1 inhibition was verified by using FFAR1-deficient MIN6 cells. Improved β-cell function by the antagonist was paralleled by the decreased apoptosis and lowered oxidation of palmitate, which may represent the potential mechanisms of protection. We conclude that FFAR1 in the pancreatic β-cell plays a substantial role not only in acute potentiation of GSIS by palmitate but also in the negative long-term effects of palmitate on GSIS and insulin content.

Prolonged exposure to palmitate impairs β-cell function and mass. One of the proposed mechanisms is alteration in ceramide (Cer) generation. In the present study, exposure to palmitate induced the level of palmitoyl transferase and Cer synthases, enzymes of the Cer de novo and salvage pathways, and doubled total Cer levels, which was associated with decreased insulin secretion and augmented apoptosis in MIN6 cells and human islets. By inhibiting enzymes of the pathways pharmacologically with myriocin (ISP-1) or fumonisin B1 or by small interfering RNA (siRNA), we showed that Cer(14:0), Cer(16:0), Cer(20:1), and Cer(24:0) species, generated by the salvage pathway, are linked to the harmful effect of palmitate on β-cells. Oleate attenuates negative effects of palmitate on β-cells. When oleate was included during culture of MIN6 cells with palmitate, the palmitate-induced up-regulation of the enzymes of the de novo and salvage pathways was prevented resulting in normalized levels of all Cer species except Cer(20:1). Our data suggest that enhanced Cer generation in response to elevated palmitate levels involves both de novo and salvage pathways. However, the negative effects of palmitate on β-cells are attributed to generation of Cer species Cer(14:0), Cer(16:0), and Cer(24:0) via acylation of sphingosine.

During the development of type I diabetes, the plasma insulin pattern changes. Because the islet secretory pattern has been implicated in this phenomenon, insulin release was measured from female nonobese diabetic (NOD) mouse islets isolated at different ages. Islets from 5-week-old mice were used as controls because they had no infiltrating mononuclear cells and insulin release rose almost 9-fold with maintained oscillatory frequency when the glucose concentration was raised from 3 to 11 mM. Islets isolated from 13- and 25-week-old mice were infiltrated with mononuclear cells. In these islets, increase in the glucose concentration from 3 to 11 mM only doubled insulin release. However, despite the cellular infiltration, insulin release was pulsatile. Islets from 13-week-old mice had reduced glucose oxidation rate. Culture of such islets for 7 days at 11.1 mM glucose causes a decrease in the number of mononuclear cells infiltrating the islets, which in the present study was accompanied by a normalization of both glucose oxidation and glucose-induced insulin release. In the presence of the mitochondrial substrate alpha-keto-isocaproate (5 mM) both control and infiltrated islets responded with pronounced insulin pulses with similar amplitudes. The results suggest that the deranged plasma insulin pattern observed during the development of type I diabetes may be related to decrease in the insulin pulse amplitude rather than loss of the pulsatile release from the islets.

Oscillatory signaling and insulin release were studied in isolated pancreatic islets and β-cells obtained from human cadaveric organ donors. Taking advantage of Sr2+ as an analog for Ca2+, it was possible to demonstrate glucose-induced rhythmic activity in individual β-cells identified by immunostaining. Glucose-induced slow oscillations of Sr2+ (frequency, 0.1–1.0/min) were sometimes seen at a sugar concentration as low as 3 mm. Addition of 20 nm glucagon resulted in a broadening of the oscillations or in their transformation into sustained elevation. Moreover, the presence of glucagon resulted in the appearance of short transients of Sr2+, which disappeared after exposure to the intracellular Ca2+-adenosine triphosphatase inhibitor thapsigargin. Digital image analyses indicated that slow oscillations can be synchronized among cells in small aggregates and intact islets. The rhythmic activity in the glucose-stimulated β-cell had its counterpart in pulsatile insulin release when single islets were perifus...

The problem of how glucose affects the intracellular (La3+-nondisplaceable) calcium content of pancreatic beta-cells was approached by combining measurements of 45Ca in ob/ob-mouse islets loaded to isotopic equilibrium with determinations of calcium using electrothermal atomic absorption spectroscopy. Whereas short term changes of the glucose concentration induced marked alterations of insulin release, the islet content of intracellular 45Ca was remarkably stable. The chronic actions of glucose differed from the acute ones in being readily demonstrable and sometimes resulting even in a suppression of the calcium content. Thus, after 7 days of culture in 20 mM glucose, the amount of intracellular calcium was actually lower than when the islets were cultured at 5.5 mM glucose. The long term effect of glucose in suppressing the islet content of intracellular calcium was associated with degranulation and loss of immunoreactive insulin, indicated both from staining of the beta-cells and measurements of the extracted hormone by RIA. The previously unknown ability of glucose to suppress the islet content of intracellular calcium may consequently result from mobilization of the secretory granules.