Indirect immunofluorescence and antibodies to tubulin were used to visualize the distribution of microtubules (MT) in short-term primary cultures of retinal pigment epithelial (RPE) cells isolated from the eyes of control RCS rats and those with inherited retinal degeneration. At all stages of cell spreading in vitro, the pigment granules (PG) in these cells remained tightly clustered in the perinuclear region, surrounded by numerous MT. Initially, in relatively round cells, the perinuclear region was encompassed by a closely woven ring of circularly arranged MT. At later stages of spreading, the ring had disappeared and MT running in various directions could be observed. Cells that were plated singly assumed at first a discoid, then stellate, shape during spreading, whereas, those plated in small groups formed colonies of wedge-shaped cells. In well-spread single cells or colonies, a brightly fluorescent zone that appeared to contain the highest density of MT within the cells was located between the PG-containing perinuclear region and the spreading edges of the cells. A single prominent star-shaped structure from which a number of MT radiated was situated among the PG in the perinuclear region of each cell; we believe this structure corresponds to the centrioles. It was generally well-separated from the regions of highest MT density. No differences in cell shape or in the distribution of MT, centriole-containing regions or PG in spreading or fully spread cells were detected when RPE cells from normal and dystrophic RCS rats were compared. Thus, the distribution of MT and their assembly during cell spreading appear normal in RPE cells of the dystrophic rat.