GABA B receptors associate with G i/o -proteins that regulate voltage-gated Ca 2+ channels and thus the intracellular Ca 2+ concentration ([Ca 2+ ]i), there is also reported cross-regulation of phospholipase C. These associations have been studied extensively in the brain and also shown to occur in non-neural cells (e.g. human airway smooth muscle). More recently GABA B receptors have been observed in chick retinal pigment epithelium (RPE). The aims were to investigate whether the GABA B receptor subunits, GABA B1 and GABA B2 , are co-expressed in cultured human RPE cells, and then determine if the GABA B receptor similarly regulates the [Ca 2+ ]i of RPE cells and if phospholipase C is involved. Human RPE cells were cultured from five donor eye cups. Evidence for GABA B1 and GABA B2 mRNAs and proteins in the RPE cell cultures was investigated using real time polymerase chain reaction, western blots and immunofluorescence. The effects of the GABA B receptor agonist baclofen, antagonist CGP46381, a G i/o -protein inhibitor pertussis toxin, and the phospholipase C inhibitor U73122 on [Ca 2+ ]i in cultured human RPE were demonstrated using Fluo-3. Both GABA B1 and GABA B2 mRNA and protein were identified in cell cultures of human RPE; antibody staining was co-localized to the cell membrane and cytoplasm. One-hundred micromolars of baclofen caused a transient increase in the [Ca 2+ ]i of RPE cells regardless of whether Ca 2+ was added to the buffer. Baclofen-induced increases in the [Ca 2+ ]i were attenuated by pre-treatment with CGP46381, pertussis toxin, and U73122. GABA B1 and GABA B2 are co-expressed in cell cultures of human RPE. GABA B receptors in RPE regulate the [Ca 2+ ]i via a G i/o -protein and phospholipase C pathway.