Lysophosphatidylcholine cell depolarization: increased membrane permeability for use in the determination of cell...

Current techniques for the determination of cellular membrane potentials based on the uptake of a radiolabeled lipophilic cation, [3H]triphenylmethylphosphonium, and the cyanine dye, DiOC5(3), were analyzed in terms of the proportions of these probes which are accumulated due to potential-dependent and potential-independent forces. Measurements were made of probe uptake in two model systems: rabbit type II pneumocytes and human promyelocytic HL60 cells. For both cell types, the membrane potential-independent component of triphenylmethylphosphonium uptake was found to be a function of several variables, including the length of exposure of the cells to the transport facilitator tetraphenylboron, the concentration of tetraphenylboron, and the integrity of the cell membrane. To accurately determine the magnitude of the potential-independent component of probe uptake by type II and HL60 cells, the cell-permeabilizing agent lysophosphatidylcholine was used. The ability of lysophosphatidylcholine to depolarize cell membranes and accurately predict membrane potential-independent accumulation was found to be equal to or superior to several other techniques commonly used to achieve membrane depolarization (e.g. gramicidin, valinomycin plus high external potassium). Lysophosphatidylcholine cell treatment was found to be a simple, rapid, and accurate technique to increase cell membrane permeability and allow equilibration of intra- and extracellular ions. The method is shown to be useful for determining membrane potential-independent accumulation of both radiolabeled and fluorescent probes of membrane potential.