Using whole-cell patch clamp recording on medial prefrontal cortical slices of rats aged 17-33 postnatal days, we demonstrated the glycine-induced strychnine-sensitive outward currents. The amplitude of the peak current increased with the concentrations of glycine with an EC50 of 74.7 mu M. Application of 1 mu M strychnine alone to cells caused a slight inward current without blocking the sIPSCs, indicating that GlyRs in the mPFC are activated by an endogenous ligand that can be released tonically. Glycine reversibly depressed firing rate in cells from both layer 6 and layer 3, with significantly greater inhibition on the former than the latter (EC50 12.9 vs 85.6 mu M). Glycine hyperpolarized membrane potential in cells of both layer 6 and layer 3 depending on its concentrations, with an IC50 of 99.1 and 207.2 mu M, respectively. We propose that GlyRs participate in a novel inhibitory mechanism in mPFC, modulating neuronal activity. This finding further supports an important role of GlyR in cortical function and dysfunction. (C) 2014 Elsevier Inc. All rights reserved.