Calcyon is a 24 kD protein recently cloned from a human brain cDNA library and shown to interact with the dopamine receptor 1 (D1R) of D1-like receptors. This interaction shifts the effector coupling of D1R to stimulate a calcium signaling pathway, without influencing the D1R-adenylyl-cAMP pathway. To obtain more knowledge about the potential role of calcyon in the brain, we cloned rat calcyon cDNA and studied its distribution in the brain. Northern blot analysis and RT-PCR revealed that rat calcyon mRNA was expressed only in the brain. With the use of the in situ hybridization technique, we studied rat calcyon mRNA distribution in the brain and related it to the distribution of D1R and dopamine receptor 5 (D5R) mRNAs. Prominent calcyon mRNA signals were found in several brain regions, including hippocampus, hypothalamus, cerebellum, and medial prefrontal cortex. Less abundant calcyon mRNA expression was observed in the dorsal striatum region, where D1R mRNA is highly expressed and where D1R/cAMP-DARPP-32 signaling pathway is of great functional importance. The strongest expression of D5R mRNA was found in the hippocampus and cerebellum, where D1R mRNA expression was relatively low. In conclusion, rat calcyon appears to be a brain specific protein. There is a certain overlap between calcyon mRNA distribution and that of the D1R and D5 mRNAs, indicating that calcyon might be associated not only with D1R but also with D5R.