Electrochemical Measurement of Cholesterol Flip-Flop in Plasma Membrane at Single Cells.

Abstract

Here, a microelectrode approach is established to measure the flip-flop rate of cholesterol in plasma membranes at single living cells. The initial validation is performed in a modeled phospholipid bilayer positioned at an interconnecting hole between two compartments, in which cholesterol in one compartment diffuses into the other one through a flip-flop movement in the bilayer and is then detected by a cholesterol oxidase-modified microelectrode. As compared with the time (140 ± 28 s) for free cholesterol transport in absence of the bilayer, a prolonged time (702 ± 42 s) is needed to observe the current increase in the presence of the bilayer. The difference in the time (562 s) gives the estimated flip-flop time of cholesterol in the bilayer. The position of the microelectrode in contact with a living cell and the injection of cholesterol inside the cell are further applied to measure the cholesterol flip-flop in the plasma membrane. The average time (1183 ± 146 s) is obtained to observe an additional current increase at the microelectrode, which reflects the cholesterol flip-flop rate in plasma membranes in single living cells. All these results support the establishment of this microelectrode approach for the study of the cholesterol flip-flop process in lipid membranes.