Fluctuation of surface charge in membrane pores

Surface charge in track-etched polyethylene terephthalate (PET) membranes with narrow pores has been probed with a fluorescent cationic dye (3,3'-diethyloxacarbocyanine iodide (diO-C-2-(3))) using confocal microscopy. Staining of negatively charged PET membranes with diO-C-2-(3) is a useful measure of surface charge for the following reasons: 1) the dye inhibits K+ currents through the pores and reduces their selectivity for cations; 2) it inhibits [H-3]-choline(+) transport and promotes Cl-36(-) transport across the membrane in a pH- and ionic-strength-dependent fashion; and 3) staining of pores by diO-C2-(3) is reduced by low pH and by the presence of divalent cations such as Ca2+ and Zn. Measurement of the time dependence of cyanine staining of pores shows fluctuations of fluorescence intensity that occur on the same time scale as do fluctuations of ionic current in such pores. These data support our earlier proposal that fluctuations in ionic current across pores in synthetic and biological membranes reflect fluctuations in the surface charge of the pore walls in addition to molecular changes in pore proteins.