1.S100B is a calcium-binding protein expressed and secreted by astrocytes, which has been implicated in glial-neuronal communication. Extracellular S100B appears to protect hippocampal neurons against toxic concentrations of glutamate. Here we investigated a possible autocrine role of S100B in glutamate uptake activity. 2. Astrocyte cultures were prepared of hippocampi from neonate Wistar rats. [(3)H] Glutamate uptake was measured after addition of S100B protein, antibody anti-S100B or TRTK-12, a peptide that blocks S100B activity mediated by the C-terminal region. 3.Antibody anti-S100B addition decreased glutamate uptake measured 30 min after medium replacement, without affecting cell integrity or viability. Moreover, low levels of S100B (less than 0.1 ng/mL) stimulated glutamate uptake measured immediately after medium replacement. 4. This finding reinforces the importance of astrocytes in the glutamatergic transmission, particularly the role of S100B neuroprotection against excitotoxic damage.