Fig. 11

Effect of sulfonylureas on the protein composition of detergent-insoluble glycolipid-enriched rafts (DIGs). (140; G. Müller, S. Welte, S. Wied, W. Frick, manuscript submitted). Isolated rat adipocytes were incubated with increasing concentrations of different sulfonylureas for 4 hr at 30°C and then lysed by incubation in the presence of 1% TX-100 for 1 hr on ice. After centrifugation (25,000X g, 15 min), the detergent-insoluble pellet fraction was subjected to sucrose density gradient centrifugation. DIGs were recovered from the 8/20%-interface of the gradient, collected by centrifugation (100,000X g, 1 hr), solubilized in sample buffer, separated by SDS-PAGE, and analyzed for the presence of typical caveolar components by immunoblotting (IB; chemiluminescence detection) with specific antibodies. Phosphorimages and chemiluminescence images of a typical experiment (repeated three times with similar results) are shown. IRS, insulin receptor substrate; Gce, cAMP-binding ectoprotein; SU, sulfonylureas.