FasL (TNFSF6, CD95L) is hypothesized to trigger testicular germ cell apoptosis that normally occurs during a distinct peripubertal period as well as in response to toxicant-induced Sertoli cell injury. To test this hypothesis, we evaluated the testis of FasL gene-deficient mice (FasL(-/-)) at two distinct developmental ages (postnatal day [PND] 28 and 44) and after toxicant-induced Sertoli cell injury. Testicular cross sections from peripubertal (PND 28) FasL(-/-) mice showed significant increases in the basal germ cell apoptotic index (AI; 20.58 +/- 4.59) as compared to the testis of C57BL/6J wild-type mice (5.16 +/- 0.08) and closely correlated with increased expression of TRAIL protein in the testis of FasL(-/-) mice. A limited, but significant, number of seminiferous tubules in the testis of PND 28 FasL(-/-) mice showed a severe loss of germ cells with only Sertoli cells present. In contrast, no apparent gross histological changes were observed in the testis of adult (PND 44) FasL(-/-) mice. However, PND 44 FasL(-/-) mice did show a 51% reduction in homogenization-resistant elongate spermatids as compared to age-matched C57BL/6J mice. Exposure of PND 28 FasL(-/-) mice to mono-(2-ethylhexyl) phthalate (MEHP), a well-described Sertoli cell toxicant, unexpectedly caused a rapid decrease in the germ cell AI that paralleled increased levels of the CFLAR (c-FLIP) protein, a known inhibitor of death receptor signaling. In contrast, MEHP treatment did not decrease c-FLIP levels in PND 28 C57BL/6J mice. Taken together, these findings indicate that FasL protein expression is required during the peripubertal period for the proper regulation of germ cell apoptosis that occurs normally during this period. The influence of FasL on the cellular regulation of c-FLIP protein levels appears to be a unique mechanism for modulating germ cell apoptosis after toxicant-induced Sertoli cell injury.