Human fetal mesencephalic neural precursor cells (NPCs) may serve as a continuous source of dopaminergic neurons for regenerative therapies in Parkinson's disease. Whole-cell recordings of ligand- and voltage-gated ion channels demonstrated the expression of functional ionotropic glutamate-receptors as well as the ability of differentiated NPCs to generate action potentials. The present study provides functional and molecular analysis of NPCs in respect to the expression of glutamate receptors. After 3 weeks of differentiation 90% of NPCs (n=56/62) responded to glutamate (mean amplitude 195 pA, peak currents up to 900 pA). The concentration-response plots of differentiated NPCs yielded an EC50 of 2.2 mM for glutamate (hillslope = 0.9, n = 20) and an EC50 of 36mM for NMDA (hillslope=0.8, n=16). NMDA- and AMPA-evoked currents could be antagonized by coapplication of memantine and CNQX, respectively. Quantitative real-time PCR showed a predominant expression of subunits NR2A and NR2B (NMDA), GluR2 (AMPA), GluR7 (Kainat), and mGlu3 (metabotropic glutamate receptor). All investigated NMDA-induced currents were blocked by the NR2B-subunit specific antagonist ifenprodil indicating functional expression of NR2B-containing receptors. In fura-2 imaging experiments, NMDA-induced calcium signals could be suppressed by memantine (100 mM) and ifenprodil (100 mM). Blocking of voltage-gated calcium channels by verapamil (100 mM) completely abolished AMPA-evoked calcium responses but only partially inhibited NMDA-induced signals, confirming the expression of calcium-impermeable, GluR2-containing AMPA receptors. These data suggest that differentiated NPCs derived from human fetal midbrain tissue acquire essential glutamate receptor properties during in vitro maturation.