Thymic involution during pregnancy is believed to be a critical adaptive mechanism for regulation and control of the maternal immune system. These regulatory feedback mechanisms are important for the survival of the semiallogeneic fetus. In the present study, we examined the effects of GnRH on pregnancy-induced thymic involution by characterizing the expression patterns of prohibitin (PHB), an antiproliferative gene product, GnRH, and GnRH receptor (GnRH-R) proteins in the rat thymus and in mature splenic lymphocytes. GnRH agonist infusions in pregnant rats markedly attenuated pregnancy-induced thymic involution resulting in significant increases in thymic weight and thymocyte numbers. In addition, histological examination of the thymus revealed increase in cortical cellularity. Western blot analyses revealed a significant increase of total PHB protein content in thymi during pregnancy. Furthermore, distinct changes in PHB isoform expression were observed in the pregnant involuting thymi with greater expression of the basic PHB isoform. Basic isoform expression decreased in pregnant rats and was comparable with nonpregnant rat thymi upon GnRH agonist treatment. PHB is mainly expressed in mature cells of the thymic medulla, where it strongly colocalized with GnRH. We have observed GnRH-R immunoreactivity mainly in thymic medulla. Furthermore, as assessed by immunofluorescence double labeling with proliferating cell nuclear antigen, PHB was preferentially expressed in nonproliferating thymocytes. In this study, we demonstrated that GnRH, GnRH-R, and PHB show characteristic polarized expression in thymocytes. In addition, GnRH and PHB were coexpressed in mature splenic T cells. Our results suggest that PHB and GnRH are involved in thymic growth and may be important for maturation of T lymphocytes.