We previously demonstrated that the inhibitory neurotransmitter glycine induced membrane currents in glial cells from rat spinal cord. In this present study, the patch-clamp technique was combined with the reverse transcription-mediated PCR to analyze the glycine receptor-subunit expression in individual glial cells of rats age 3-18 days. Using the patch-clamp technique in the whole-cell configuration, glial cells were identified by their membrane current pattern and tested for responsiveness to glycine. Subsequently, the cytoplasm was harvested followed by reverse transcription of total cytoplasmic RNA. Subunit-specific cDNA fragments were amplified and analyzed by agarose gel electrophoresis, Southern blotting, and sequencing. In all cell types investigated, transcripts of the alpha1 subunit, but not of alpha 2 or alpha 3 subunits, were detected. In addition, about one-half the glial cells analyzed contained beta-subunit mRNA. These results illustrate that glial cells of rat spinal cord express functional glycine receptors in contrast to cultured glial cells. Glial cells are in intimate contact with synaptic regions making it likely that these nonneuronal receptors may be activated during glycinergic transmission and may trigger yet unknown responses in the glial cells.