Nucleosides are important modulators of neural activity and survival. In the present study, we produced primary cultures of rat cortical astrocytes, as described earlier [1], and examined the expression of nucleoside transporters at transcript level by reverse transcription-polymerase chain reaction (RT-PCR) and by real time PCR; the sequence of the primers and the expected sizes of the products are presented in the Table 1. RT-PCR identified the mRNAs for the rat equilibrative nucleoside transporters (rENT)1, rENT2 and recently characterized rENT3, as well as for the rat concentrative nucleoside transporters (rCNT)2 and rCNT3, while the band corresponding to rCNT1 was faint (Figure 1). Real time PCR was used to explore the amount of mRNA for these transporters, relative to the amount of mRNA for the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase(GAPDH). Results have revealed that mRNA encoding rENT3, a transporter mainly located in lysosomes, was the most abundant, with relative expression of 1.47±0.23 (mean±SEM, n=4); the amounts of mRNA for rENT2 and rENT1 were 0.65±0.26 (n=3) and 0.19±0.06 (n=4), respectively. Amounts of mRNA for the two purine-preferring concentrative transporters, rCNT2 and rCNT3 were also relatively high, 1.22±0.23 (n=4) and 1.11±0.19 (n=3), respectively, while the mRNA for the pyrimidine-preferably rCNT1 was scarce, 0.14±0.07 (n=3) relative to mRNA for GAPDH. These results demonstrate for the first time that rat astrocytes in primary culture express all six nucleoside transporters; however, relative expression indicates that concentrative transport of purine nucleosides across the cellular membrane, as well as the equlibrative transport within the intracellular compartment, may be important for homeostasis of nucleosides and nucleotides in these cells.