Glucocorticoid receptor (GR) distribution in isolated rat hepatocytes and nonparenchymal hepatic stellate cells, Kupffer cells, and liver fibroblasts with and without dexamethasone treatment was investigated by immunostaining and confocal laser scanning microscopy. In addition, human liver fibroblasts, Hep3B and HepG2 cells were investigated. Subcellular distribution of GR immunostaining was assessed semiquantitatively by digital image analysis. Short-term incubation of rat liver cells with dexamethasone resulted in an increase of nuclear staining. The same was true for human liver fibroblasts. In contrast, predominant nuclear staining could be observed in untreated as well as in dexamethasone-treated Hep3B and HepG2 cells. By means of reverse-transcription polymerase chain reaction, it could be shown that messenger RNA of both known human GR isoforms, hGR α and nonhormone-binding hGR β, are present in human cells. Furthermore, dexamethasone binding indicates that hGR α protein is expressed in all human cells investigated. The data of this study show that GR is present in all cells investigated. Rat liver cells and human liver fibroblasts contain a translocating GR, suggesting that glucocorticoid action is receptor mediated in these cells. Nuclear localization of unliganded GR in Hep3B and HepG2 indicates that factors other than glucocorticoids may direct subcellular GR distribution.