Permeation of Chinese hamster ovary cells by glycerol: mechanism and kinetics.

When monolayer cultures of Chinese hamster ovary cells were exposed to 3H-glycerol ranging in concentration from 0.1 microM to 200 mM, glycerol in flux was found to increase in direct proportion to the extracellular concentration of glycerol. Other experiments indicated that the same relationship existed at concentrations in excess of 1.0 M. Similarly, glycerol efflux was found to vary in direct proportion to the intracellular concentration of glycerol. In neither case could influx or efflux be saturated. Glycerol influx was not affected by depletion of ATP, alkylation by parachloromercuribenzene sulfonic acid, or exposure to persantine. Altering the pH or temperature also had little effect. Attempts at demonstrating countertransport of glycerol were negative. These data indicate that glycerol probably passes through the membrane by a non-mediated process. For cells in monolayer, the kinetics of influx and efflux are biphasic. Similar biphasic kinetics are observed with cells in suspension culture. A close fit to the data may be obtained by adding together two first-order curves. The pair of curves for influx are clearly different from the pair for efflux. The fit provided by the two pairs of first-order functions suggested that glycerol might diffuse into and out of two intracellular compartments. However, the experimental data do not agree with the predicted behavior of a two-compartment system. As a result, the exact nature of the diffusion limiting steps which are described by the first-order equations remains undefined.