Insulin secretion is highly sensitive to desorption of plasma membrane cholesterol

Abstract

Cholesterol-rich clusters of SNARE (soluble NSF attachment protein receptor) proteins have been implicated as being important for exocytosis. Here we demonstrate the significance of cholesterol for normal biphasic insulin secretion in mouse beta cells by removal of cholesterol from the plasma membrane using methyl-beta-cyclodextrin (MBCD). Maximal inhibition of insulin secretion in static incubations was achieved using 0.1 mM MBCD. In in situ pancreatic perfusion measurements, both first and second phase insulin secretions were reduced by approximately 50% (P<0.05). This was accompanied by a reduced number of docked large dense core vesicles (LDCVs) (approximately 40%; P<0.01) and a reduced exocytotic response (>50%; P<0.01). Further, subcellular fractionations demonstrated movement of the synaptosomal protein of 25 kDa (SNAP-25) from the plasma membrane to the cytosol after MBCD treatment. The inhibitory actions of MBCD were counteracted by subsequent addition of cholesterol. We hypothesize that desorption of cholesterol leads to the disturbance of a basic exocytotic mechanism partly due to migration of SNAP-25, and we conclude that insulin secretion is highly sensitive to changes in plasma membrane cholesterol.

Glucose-dependent docking and SNARE protein-mediated exocytosis in mouse pancreatic alpha-cell

“…However, earlier data from our group have demonstrated that similar result is obtained when using the smaller Fab-fragments of an antibody as when using the full-size antibody [35]. In this study we performed control experiments using IgG to ascertain that the antibody did not reduce the exocytotic response simply by steric hindrance.…”

“…Inc., MA, USA). Micrographs were analysed with respect to the intracellular granular distribution using similar methods that have been described elsewhere [35]. The apparent diameter of individual granules was determined with the image processing software Scion Image (NIH freeware) and the exact diameter was calculated as described elsewhere [24].…”

Cluster analysis of rat pancreatic islet gene mRNA levels after culture in low-, intermediate- and high-glucose concentrations

“…Gene mRNA levels are expressed relative to the level in G10, data are means±SE for three (real-time RT-PCR) and four (microarray) experiments. The mean signal intensity (microarray) and threshold cycle measured in G10 (real-time PCR performed with islet cDNA equivalent to 2 ng total RNA) are shown as rough indicators of gene mRNA levels *p<0.05, **p<0.01 vs islets cultured in G10 (one-way ANOVA and Newman-Keuls test) NA, not affected support of that hypothesis, cholesterol depletion and Srebp1c (also known as Srebf1) gene inactivation significantly altered beta cell function [41,42]. However, the fact that Srebp1c and Srebp2 overexpression are also deleterious to beta cell differentiation and function [43,44] suggests that both excessive and insufficient cholesterol availability is harmful to beta cells.…”

Distinct pathways of cholesterol biosynthesis impact on insulin secretion

“…On the other hand, insulin secretion was reduced in mouse pancreatic islets after inhibition of cholesterol biosynthesis downstream of farnesyl synthesis (Xia et al 2008). Moreover, depletion of cholesterol by MbCD has been also shown to reduce insulin secretion in mouse islets (Vikman et al 2009). Interestingly, high levels of cholesterol may also influence insulin secretion.…”