Nucleoside transporters play important physiological roles by regulating intra- and extra-cellular concentrations of purine and pyrimidine (deoxy)nucleosides. This review describes the biological function and activity of the two major families of membrane nucleoside transporters that exist in mammalian cells. These include equilibrative nucleoside transporters that transport nucleosides in a gradient-dependent fashion and concentrative nucleoside transporters that import nucleosides against a gradient by coupling movement with sodium transport. Particular emphasis is placed on describing the roles of nucleoside transport in normal physiological processes, including inflammation, cardiovascular function and nutrient transport across the blood-brain barrier. In addition, the role of nucleoside transport in pathological conditions such as cardiovascular disease and cancer are discussed. The potential therapeutic applications of manipulating nucleoside transport activities are discussed, focusing on nucleoside analogs as anti-neoplastic agents. Finally, we discuss future directions for the development of novel chemical entities to measure nucleoside transport activity at the cellular and organismal level.