Studies were designed to examine the expression and activity of four caspases, which contribute to the initial (caspases-2, -8 and -9) and final (caspase-3) events in apoptosis, in the rat CL during pregnancy (days, 7, 17, 19 and 21 of gestation), postpartum (day 1 and 4) and after injection (0, 8, 16, 24 and 36 h) of the physiological luteolysin, PGF-2. In addition, the temporal relationship of caspase expression/activity relative to steroid production and luteal regression was evaluated. During pregnancy, the activity of all four caspases was significantly greater on day 19, prior to a decline in CL progesterone (P) and CYP11A1 levels at day 21 of gestation. The levels of the caspase-3 active fragment (p17, measured by Western blot) also increased at day 19 and 21 of pregnancy. Immunohistochemical analyses detected specific staining for the caspases in luteal cells (large and small), as well as, in endothelial cells. However, the percentage of apoptotic cells did not increase in the CL until postpartum. Following PGF-2 injection, there was a significant decrease in CL P by 24 h, though the activity of all four caspases did not increase until 36 h post treatment. The active p17 fragment of caspase-3 also significantly increased at 36 h post- PGF-2. These results suggest that an increase in the activity of caspase-2, -8, -9 and -3 is associated with the early events of natural luteolysis at the end of pregnancy. Also the exogenous administration of the luteolysin PGF-2 may regulate members of the caspase family.