Membrane potential, chloride exchange, and chloride conductance in Ehrlich mouse ascites tumour cells

1. The steady-state tracer exchange flux of chloride was measured at 10-150 mM external chloride concentration, substituting either lactate or sucrose for chloride. The chloride flux saturates in both cases with a K 1/2 about 50 and 15 mM, respectively. 2. The inhibitory effect of other monovalent anions on the chloride transport was investigated by measuring the 36Cl- efflux into media where either bromide, nitrate, or thiocyanate had been substituted for part of the chloride. The sequence of increasing affinity for the chloride transport system was found to be: Br- less than Cl- less than SCN- = NO3-. 3. The chloride steady-state exchange flux in the presence of nitrate can be described by Michaelis-Menten kinetics with nitrate as a competitive inhibitor of the chloride flux. 4. The apparent activation energy (EA) was determined to be 67 +/- 6.2 kJ/mole, and was constant between 7 and 38 degrees C. 5. The membrane potential (Vm) was measured as a function of the concentration of external K+, substituting K+ for Na+. The transference number of K+ (tK) was estimated from the slope of Vm vs. log10 (K+)e, and tCl and tNa were calculated, neglecting current carried by ions other than Cl-, K+, and Na+. The diffusional net flux of K+ was calculated from the steady-state exchange flux of 42K+, assuming the flux ratio equation to be valid. From this value the K+ conductance and the Na+ and Cl- conductances were calculated. The experiments showed that GCl, GNa, and GK are all about 14 muS/cm2. 6. The net (conductive) chloride permeability derived from the chloride conductance was 4 x 10(-8) cm/sec compared with the apparent permeability of 6 x 10(-7) cm/sec as calculated from the chloride tracer exchange flux. These data suggest that about 95% of the chloride transport is mediated by an electrically silent exchange diffusion. 7. Comparable effects of phloretin (0.25 mM) on the net (conductive) permeability and the apparent permeability to chloride (about 80% inhibition) may indicate that the chloride exchange and conductance pathways are not completely separate and distinct modes of transport, but may involve common elements. The reduced chloride permeability in the presence of phloretin is estimated to be two orders of magnitude larger than the ground permeability of the cell membrane.