The hypothalamus is involved in many physiological functions in teleosts. To accelerate the molecular analysis of hypothalamic functions, a list of transcripts expressed in the hypothalamus of adult tilapia, Oreochromis mossambicus, was compiled using the expressed sequence tag (EST) strategy. Of 161 clones, 130 clones were unique genes and 31 clones were found to be redundant. Of 130 unique genes, 32.3% (42/130 ESTs) were identified as known genes and 67.7% (88/130 ESTs) as unknown genes. The functional categorization of the known genes was analyzed. Bioinformatic analysis revealed that 62 of 88 unknown genes (62/130 ESTs, 47.7%) showed a significant homology to neither nucleotide nor translated peptide sequences in the public database. These genes might be particularly expressed in the tilapia hypothalamus.