Plasminogen Activators (PAs) and their Inhibitors (PAIs) are predicted to be involved in the restructuring events that characterize the testis throughout development. We here demonstrated that PAI-3 or Protein C Inhibitor (PCI) was expressed in a sexually dimorphic fashion during mouse gonad genesis, whereas PAIs -1 and -2 exhibited no sex-differences. PCI transcripts accumulated rapidly in the male gonad, from 12.5 days postcoitum (dpc) onward. Western blot and immunohistochemistry (IHC) analyses confirmed that male but not female fetal gonads produced PCI, and that Leydig cells are the site of PCI synthesis. The occurrence of testicular target proteases for PCI, i.e. Protein C (PC), urokinase- and tissue-type PA was further tracked using RT-PCR, plasminogen zymography and/or IHC. PC and tissue-type PA had no variations between sexes. By contrast urokinase-PA and its receptor (uPAR, which dictates the site and extent of proteolysis) exhibited sex-differences from 13.5-14.5 dpc. At that time, uPAR expression was restricted to Leydig cells. At earlier ages, uPAR was uniformly and widely distributed in the gonads of both sexes. In adult testes, PCI and uPAR immunoreactivities were also present in Leydig cells. In addition, PCI, PC and uPAR had a germinal origin. Collectively, these results support the hypothesis that PCI may contribute to the proteolysis equilibrium within the testis, by acting in tandem with urokinase in Leydig cells and with PC and/or urokinase in spermatogenic cells. Of importance now is to determine how much this role is linked to the phenotype of sterility reported elsewhere in male mice deleted for pci.