Membrane Perfusion of Hydrophobic Substances Around Channels Embedded in the Contact Bubble Bilayer

Abstract

In fluidic biomembranes, lipids and membrane proteins diffuse restlessly, and lipid compositions change steadily. To mimic dynamic behavior of the biomembranes, a method for introducing rapid changes in the constituents in the lipid bilayer was developed. In contact bubble bilayers (CBB), as a water-in-oil droplet bilayer system, the bilayer hydrophobic interior is contiguous with the bulk oil phase. Making use of this geometrical feature as an access route, hydrophobic substances were administered into the bilayer. Polytheonamide B, a cytotoxic hydrophobic peptide, was applied, and oriented incorporation and relevant single-channel current recordings were enabled. Nystatin was pre-loaded in the CBB, and sterol perfusion exhibited slow development of the macroscopic current. On the contrary, the reconstituted KcsA potassium channels immediately attenuate the channel activity when cholesterol was applied. This oil-phase route in the CBB allows rapid perfusion of hydrophobic substances around the bilayer-embedded channels during continuous recordings of channel currents.