EFFECT OF MEDIUM CONDUCTIVITY AND COMPOSITION ON THE UPTAKE OF PROPIDIUM IODIDE INTO ELECTROPERMEABILIZED MYELOMA CELLS

The effects of ionic composition and conductivity of the medium on ele ctropermeabilization of the plasma membrane of mammalian cells were st udied. Temporal and spatial uptake of propidium iodide (PI) into field -treated cells was measured by means of flow cytometry, spectrofluorim etry and confocal laser scanning microscopy. Murine myeloma cells were electropulsed in iso-osmolar solutions. These contained 10-100 mu g m l(-1) PI at different conductivities (0.8-14 mS cm(-1)) and ionic stre ngths, adjusted by varying concentrations of K+, Na+, Cl- and SO42-. F ield-induced incorporation of PI into reversibly permeabilized cells w as (almost) independent of ionic composition and strength (at a fixed medium conductivity), but increased dramatically with decreasing mediu m conductivity at a fixed field strength. The time-course of PI uptake (which apparently reflected the resealing process of the membrane) co uld be fitted by a single-exponential curve (relaxation time about 2 m in in the absence of Ca2+) and was independent of medium conductivity and composition. These and other data suggested that the expansion of the 'electroleaks' during the breakdown process is field-controlled an d depends, therefore, on the (conductivity-dependent) discharging proc ess of the permeabilized membrane. The threshold field strength for dy e uptake increased with increasing K+ concentration (about 0.6 kV cm(- 1) in K+-free, NaCl-containing medium and about 0.9 kV cm(-1) in 30 mM KCl-containing medium). Also, the spatial uptake pattern of PI shifte d from an asymmetric permeation through the cell hemisphere facing the anode to a more symmetric uptake through both hemispheres. These resu lts suggested that the generated potential is superimposed on the (K+- dependent) resting membrane potential. Taking this into account, the b reakdown voltage of the membrane was estimated to be about 1 V.