IDENTIFICATION OF THYROTROPIN-RELEASING-HORMONE RECEPTOR IN THE RAT TESTIS

We have recently documented the expression of preprothyrotropin-releasing hormone (TRH) gene in murine, human and rat testis. Moreover, we have localized TRH to rat Leydig cells immunohistochemically, and found that both prepro TRH mRNA and TRH levels are developmentally regulated in the rat testis. To investigate the potential roles of TRH in testicular function, characterization of TRH receptors (TRH-R) in this tissue was undertaken. Recently, a cDNA encoding murine TRH-R has been isolated, making possible cloning of a rat TRH-R cDNA from the anterior pituitary gland. This cDNA was used for detection of TRH-R gene expression in the rat testis by Northern blot analysis and reverse transcription-polymerase chain reaction (RT-PCR). TRH receptor assays were also performed with (H-3)MeHisTRH as the radioactive ligand. In Northern blot analysis, a single and specific hybridization band, approximately 3.8 kb in size, was identified in whole testis RNA, identical in size with that found in the anterior pituitary gland. The concentration of IRH-R mRNA in the testis was approximately 10% of that in the pituitary. TRH-R mRNA was also detected by RT-PCR in Metrizamide gradient-purified Leydig cells. TRH receptor binding assays revealed the presence of specific, high affinity binding sites with a Kd of 1.6 x 10(-8) M in the testis. Such TRH binding was inhibited by chlordiazepoxide, a specific antagonist of TRH receptor binding. We conclude that TRH may exert local, probably autocrine, actions in the testis via a transmembrane receptor very similar or identical to that in pituitary.