Gonadotropin releasing hormone (GnRH) is the key hypothalamic neurohormone that is critical in its role of reproduction in all vertebrates. There are currently twenty-four known forms of GnRH that have been identified, 14 in vertebrates and 10 in invertebrates. In tunicates, the primary structure of nine forms have been identified, all of which have been shown to stimulate gamete release. However, the distribution and function of the various GnRH peptides in tunicates have not been fully examined. Therefore, the objective of this study was to determine tissue specific expression of Ci-gnrh-1 and Ci-gnrh-2 in an adult tunicate, Ciona intestinalis, using reverse transcriptase-polymerase chain reaction (RT-PCR) and in situ hybridization. To examine the expression of the two GnRH genes, total RNA and genomic DNA were isolated from whole animals. Total RNA from neural tissue (cerebral ganglion and neural gland), testis, ovary, heart, and hepatic organ were also isolated. Results from RT-PCR indicated both forms are only expressed in the neural tissue. We extended these studies using fluorescent dual label in situ hybridization. GnRH expression was confirmed to be in the cerebral ganglion bordering the neural gland. These current data along with previous studies suggest that GnRH may be involved in reproduction in the protochordate.