We have used immunocytochemistry and in situ hybridization to examine the expression of photoreceptor specific genes in retinas of normal dogs and those affected with hereditary cone degeneration (cd), a rare autosomal recessive disorder that selectively affects cones. In the cd retina, cone disease begins early in life; cones are lost by extrusion of the nucleus into the inner segment, and later by displacement of the nucleus, surrounded by a thin rim of cytoplasm, into the interphotoreceptor space. Two micrometer sections from the superior and inferior retinal meridians, extending from the optic disk to the ora serrata, were used for in situ hybridization with a bovine rod opsin and human red/green cone opsin cRNA probes, or were reacted with antibodies directed against photoreceptor-specific proteins and visualized with appropriate biotinylated antibodies. Antibodies against the following proteins were used: alpha- and beta 3-transducins, phosducin, alpha/beta- and gamma-phosphodiesterases, COS-1, and OS-2, opsin, S-antigen and IRBP. Immunoreactivity or hybridization labeling was evaluated in unstained sections; cone pathology was judged in adjacent Toluidine Blue-stained sections. With these methods it was possible to evaluate immunoreactivity or hybridization labeling and cone pathology at the single cell level. Both middle-(COS-1) and short-(OS-2) wavelength-sensitive cones were present in controls and cd affected retinae at 2.2 months, and distinct transcripts of the red/green cone pigment gene were identified in the majority of cones in both normal and affected retinas at this age. However, beta 3-transducin immunoreactivity was completely absent from cd-affected cone outer segments. Both cone types were present but in reduced numbers in older animals (11.5 and 17 months), and no reactivity to beta 3-transducin was noted. No differences were found with the other antibodies used. The specific absence of beta 3-transducin immunoreactivity from the cone outer segments suggests a potential involvement of the beta 3-transducin gene or gene product in the disease process.