Speed of Accumulation of the Membrane Potential Indicator diS-C₃(3) in Yeast Cells

Abstract

The carbocyanine dye diS-C₃(3) (3,3’- dipropylthiacarbocyanine iodide), whose the steady-state fluorescence spectra were measured in yeast cell suspensions, belongs to the group of slow (Nernstian, or redistribution) dyes which report on membrane potential by their voltage-sensitive partition between the extracellular medium and the cytosol.^1–3 Since the emission spectrum shifts and the quantum yield of fluorescence increases upon binding of the dye in the cell, two fluorescence parameters,^4–5 the wavelength of emission maximum and the intensity of fluorescence at this wavelength, were used to monitor the redistribution of the dye inside/outside the cells. To demonstrate that the dye accumulation in cells, as revealed by observed fluorescence changes, is actually membrane-potential-driven we used the uncoupler CCCP (carbonyl cyanide 3-chlorophenylhydrazone) which drastically increases membrane permeability for protons and depolarizes the cell membrane.⁶