In order too examine whether there is species difference in the function of the thin ascending limb of Henle (tALH), segments isolated from rat and hamster kidneys were perfused in vitro and compared to results obtained by identical techniques from rabbits. The net water flux and the transtubular potential difference were not different from zero. The osmotic water permeability of the rat and hamster tALH was unmeasurably small. Isotopic and electrically determined permeabilities for Cl- and Na+ were quite high in both species, with Cl- being approximately twice as permeable as Na+. Urea permeability across the tALH of both species was moderate. Solute permeability to NaCl, urea, and raffinose (10(-5) cm sec-1) of the tALH of both species was determined by measuring changes in osmolality of the collected fluid when each solute was added to the bath at the same osmolality. They were 82.2, 38.3, and 6.9, respectively. The relative permeabilities to various ions were estimated from the deflection of the transtubular potential difference when NaCl in the bathing medium was replaced by various salts. The following sequence of permeabilities was obtained across the tALH from both the rat and hamster: Cl- greater than or equal to Br- greater than or equal to SCN- greater than or equal to I- greater than or equal to Na+ = K+ greater than Li+ greater than choline+ greater than or equal to HCO3- greater than acetate- greater than or equal to cyclamate-. These data indicate that membrane characteristics of the tALH of rats and hamsters are similar to the previously published values of rabbits. The conclusion is drawn that the principle of the passive equilibrating model of the countercurrent multiplication system is operative in other mammalian species besides the rabbit.