Visualizing the osmotic water permeability of a lipid bilayer under measured bilayer tension using a moving membrane method

The water flux across the cell membrane is a fundamental physiological process ubiquitous to all cell types. For evaluating water permeability (Pf), volume changes of cells and vesicles have been employed to measure water flux. However, upon volume expansion, the membrane stretches, and Pf could vary with membrane tension changes. Moreover, local concentration polarization of solutes occurs at the membrane interface due to bulk water flux across the membrane (unstirred layer; UL), which hampers evaluation of Pf values. Here, we establish an accurate method for evaluating the osmotic Pf of lipid bilayers under constant bilayer tension and known unstirred layer. A lipid bilayer is formed across a glass capillary by docking monolayers at oil-water interfaces, and osmotically driven bulk water flux across the membrane yields translational membrane movements along the capillary (moving membrane method). Integrated electrical and image analyses allowed a quantitative evaluation of membrane tension, which remained constant during measurements. In parallel, the unstirred layer was quantified at the nearest vicinity of the membrane using the gramicidin channel as a sensor. Accordingly, Pf values without and with cholesterol were evaluated under known bilayer tension and unstirred layers. Pf of the gramicidin A channel was also measured.