Mechanism of response of potential-sensitive dyes studied by time-resolved fluorescence

Abstract

The mechanism of response of two potential-sensitive dyes, diOC₂(5) (3,3-diethyloxadicarbocyanine iodide) and oxonol V (bis-[3-phenyl-5-oxoisoxazol-4-yl]pentamethine oxonol), were studied by using steady-state and time-resolved fluorescence techniques. The lipid concentration dependence of the Δψ(membrane potential)-induced change in total fluorescence intensity was quite different for these two dyes. Time-resolved fluorescence measurements showed that the fluorescence decay of these dyes in membranes could be resolved into at least three exponentials.-induced changes in the levels of these three populations were also measured under a variety of conditions. In the case of diOC₂(5) an inside negative Δψincreased the levels of the bound forms. This shows that diOC₂(5) responds to mainly by an on-off mechanism whereby Δψ perturbs the membrane-water partition coefficient of the dye. The Δψ-induced changes approached zero when the dye was totally membrane bound. In contrast, the Δψ induced response of oxonol V increased with increased membrane binding. An inside negative Δψ decreased the level of the bound form with a longer lifetime. This shows that the mechanism of response of oxonol V is a Δψ-induced shift in the equilibrium between bound forms of the dye.