Interferon gamma (IFNgamma) is a highly pleotropic pro-inflammatory and anti-viral cytokine that mediates its effects by binding to a receptor complex composed of interferon gamma receptors 1 and 2 (IFNGR1 and IFNGR2). Using gene synteny analysis, we identified a distinct isoform of the zebrafish IFNGR1. The two zebrafish IFNGR1 called here IFNGR1-1 and IFNGR1-2 were used to identify the respective cDNA sequences of the goldfish IFNGR1-1 and IFNGR1-2. Analysis of protein sequences revealed that all fish IFNGR1 species have potential JAK1 and STAT1 docking sites. Phylogenetically, teleost IFNGR1 proteins grouped separately from those of higher vertebrates. Q-PCR analysis revealed that while the constitutive mRNA levels of the two zebrafish IFNGR1 isoforms were comparable in different tissues examined, the goldfish IFNGR1-1 tissue expression was substantially higher than that of IFNGR1-2. Q-PCR analysis of goldfish immune cell populations revealed highest expression of both receptor isoforms in monocytes. Incubation of goldfish macrophages with recombinant goldfish IFNgamma2 (rgIFNgamma2) up-regulated expression of both IFNGR1-1 and IFNGR1-2, while treatment of cells with rgTNFalpha2 only increased the expression of IFNGR1-1. Treatment with rgTGFbeta resulted in more modest increases in expression of both receptor isoforms only after prolonged treatment. In vitro binding studies indicated that rgIFNGR1-1 bound to rgIFNgamma1 but not rgIFNgamma2, while the rgIFNGR1-2 bound to rgIFNgamma2. Thus, unlike mammals that have a single IFNGR1, cyprinid fish have two distinct IFNGR1 isoforms that preferentially bind corresponding ligands, IFNgamma1 and IFNgamma2, respectively, suggesting that the type II interferon system of these fish species is distinct from that of higher vertebrates.