The transport properties of the rabbit peritoneal polymorphonuclear leukocyte (PMN) plasma membrane to Na+, K+, and Ca2+ have been characterized. The use of a silicone oil centrifugation technique provided a rapid and reliable method for measuring ion fluxes in these cells. Na+ and K+ movements across PMN membranes were found to be rapid. The value for the unifirectional steady-state fluxes (in meq/liter cell X min) were of the order of 3.0 for Na+ and 7.4 for K+. Ouabian inhibited both K+ influx and Na+ efflux, the latter being also dependent on the presence of extracellular potassium. The rate constant (in min-1) for 45Ca influx was found to be .05 and that for 45Ca efflux .04. The synthetic chemotactic factor formyl-methionyl-leucyl-phenylalanine (FMLP) was found to affect the fluxes of Na+, K+, and Ca2+ at concentrations as low as 10(-10)M. FMLP induced a large and rapid increase in the permeability of the PMN plasma membrane to 22Na. Smaller and delayed enhancements of 42K influx and 22Na efflux were also noted. Some evidence that the latter findings are a consequence of the increased 22Na influx is presented. 45Ca influx and efflux were also stimulated by FMLP. In the presence of 0.25 mM extracellular calcium, FMLP induced an increase in the steady-state level of cell-associated 45Ca. In the presence of .01 mM extracellular calcium, however, a transient decrease in the steady-state level of cell-associated 45Ca was induced by FMLP. The curves relating the concentration of FMLP to its effects on cation fluxes are very similar to those found for its enhancement of migration.