MECHANISM OF RESPONSE OF POTENTIAL-SENSITIVE DYES STUDIED BY TIME-RESOLVED FLUORESCENCE

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 DELTApsi (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. DELTApsi-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 DELTApsi increased the levels of the bound forms. This shows that diOC2(5) responds to DELTApsi mainly by an ''on-off '' mechanism whereby DELTApsi perturbs the membrane-water partition coefficient of the dye. The DELTApsi-induced changes approached zero when the dye was totally membrane bound. In contrast, the DELTApsi-induced response of oxonol V increased with increased membrane binding. An inside negative DELTApsi decreased the level of the bound form with a longer lifetime. This shows that the mechanism of response of oxonol V is a DELTApsi-induced shift in the equilibrium between bound forms of the dye.