The effects of GBR-12909 (selective DA uptake inhibitor), zimelidine (selective 5-HT uptake inhibitor) and nisoxetine (selective NE uptake inhibitor) on the uptake of 30 nM [3H]DA into cultured rat astrocytes were examined. [3H]DA uptake was inhibited by approximately 50% by GBR-12909 or zimelidine in a concentration-dependent manner (100 nM to approximately 10 microM). Furthermore, the inhibition curves of GBR-12909 were biphasic, and uptake was completely inhibited by a high concentration of GBR-12909 (100 microM). [3H]DA uptake was also inhibited by approximately 50% by nisoxetine in a concentration-dependent manner (0.1 to approximately 100 nM), and nisoxetine was more potent than GBR-12909 or zimelidine. The inhibitory potencies were in the order nisoxetine > GBR-12909 > zimelidine. The uptake of [3H]DA under Na+-free conditions was approximately 50% of that under normal conditions. Thus, DA was taken up by both Na+-dependent and Na+-independent mechanisms. Nisoxetine (100 nM), zimelidine (100 microM) and GBR-12909 (10 microM) inhibited [3H]DA uptake into astrocytes only in the presence of Na+. On the other hand, this uptake was completely inhibited by a high concentration of GBR-12909 (100 microM) in the absence of Na+. The present data suggest that the Na+-dependent uptake of [3H]DA in cultured rat astrocytes may occur in the NE uptake system. Furthermore, astrocytes express the extraneuronal monoamine transporter (uptake2), which is an Na+-independent system, and this transporter is involved in the inactivation of centrally released DA.