Light-microscopic studies of 3T3 cell plasma membrane alterations mediated by melittin

Various light microscopic techniques were used to study the effect of melittin, a major toxic constituent of honey bee venom, on plasma membranes of 3T3 mouse fibroblasts. Bright-field light microscopy and Trypan Blue dye exclusion were used to demonstrate changes in membrane permeability after exposure to melittin. Differential interference contrast (DIC) microscopy showed that membrane vesiculation induced by melittin was dose dependent. Using both fluorescent lipid and glycoprotein markers, we found that membrane vesicles were primarily composed of lipids. A sequence of events associated with vesicle formation was depicted by DIC and fluorescence microscopy. Confocal laser scanning fluorescence microscopy demonstrated a translocation of membrane glycoproteins from the plasma membrane to the cytosol following melittin treatment. The significance of membrane vesiculation and translocation of membrane glycoproteins in damaged cells is discussed.