The rate of uptake of aspartate into the rat hepatoma cell line H4-II-E-C3 is very much higher than that exhibited by normal rat hepatocytes. Using an RT-PCR-based strategy, a glutamate transporter resembling mouse liver GLT-1A has been cloned from H4-II-E-C3 cells. Northern blotting confirmed that relatively high levels of mRNA for GLT-1A are expressed in hepatoma cells compared with negligible levels in rat hepatocytes. To our knowledge, this is the first report of the cloning of a high-affinity glutamate transporter from a transformed cell line and also the first demonstration of functional expression of GLT-1 outside the central nervous system.