Uridine uptake was examined in Tetrahymena pyriformis GL-7 in defined medium under conditions where food vacuole formation is not a significant factor in solute acquisition by the cell. The results indicate the presence of a saturable mechanism which follows Michaelis-Menten kinetics. When corrected for diffusion the apparent Km for the carrier is 2.3 +/- 0.6 muM and the Vmax is 7.3 +/- 0.2 X 10(-7) nmoles/cell/min. It is evident from nucleotide pool analysis that most of the radioactivity of externally supplied [3H]uridine appears in UMP with the remainder in UTP. Uridine is apparently phosphorylated immediately upon entry into the cell and neither uridine-cytidine kinase activity nor RNA synthesis are rate-limiting in the uptake process. Uridine transport is competitively inhibited by a variety of ribo- and deoxyribonucleosides as well as several nucleoside analogs. Neither uracil nor ribose or deoxyribose are effective inhibitors of uridine transport indicating the carrier is specific for the nucleoside. There is little difference between the Ki values for ribo- as opposed to deoxyribonucleosides except in the case of deoxyguanosine which is much less effective as an inhibitor under the conditions of this study, than all the other nucleosides, including guanosine.