Gangliosides released from tumor cells, as well as administered exogenously, suppress the immune responses by largely unknown mechanisms. We show here that a pretreatment of rat basophilic leukemia cells with isolated brain gangliosides inhibited the release of preformed secretory mediators from cells activated via FcepsilonRI but not Thy-1 glycoprotein. Exogenously administered gangliosides also affected the cell-substrate adhesion and the levels of polymeric filamentous actin in Ag-activated cells. Although the production of phosphoinositides was also decreased, enzymatic activity of phosphatidylinositol 3-kinase was not inhibited. Gangliosides had no or only marginal effect on the association of aggregated FcepsilonRI with glycosphingolipid-enriched membranes and on tyrosine phosphorylation of FcepsilonRI and the linker for activation of T cells. Though pretreatment with gangliosides did not inhibit the association of linker for activation of T cells with phospholipase C (PLC)gamma1 and PLCgamma2, tyrosine phosphorylation of these enzymes, as well as their enzymatic activities and association with detergent-insoluble signaling assemblies were reduced. This resulted in a decreased production of inositol 1,4,5-trisphosphate and an inhibition of Ca(2+) mobilization. The combined data support the concept that exogenously administered gangliosides interfere with those properties of glycosphingolipid-enriched membranes that are important for the formation of plasma membrane-associated signaling assemblies containing PLCgamma but not for initial tyrosine phosphorylation of FcepsilonRI subunits.