Abstract
Activities of the steroidogenic enzymes involved in functional luteolysis and PGF2α induced luteolysis were determined in PMSG-hCG primed immature rats to elucidate the luteolytic effects of PGF2α. Plasma progesterone (P4), plasma 20α dihydroprogesterone (20αOHP4), in vitro production of pregnenolone (P5) from endogenous cholesterol in ovarian mitochondria (8,000×g pellet; Mt), activities of 3β hydroxysteroid dehydrogenase (313HSD) in ovarian microsome (105,000×g pellet; Ms) and 20α hydroxysteroid dehydrogenase (20αHSD) in ovarian cytosol (105,000×g supernatant; Sup) were determined. For the investigation of intramitochondrial transport of cholesterol, measurement of free cholesterol (FCh) of Mt and the Lineweaver-Burk plotting for cholesterol side-chain cleavage enzyme (CSCC), prepared by osmotic shock and sonication of Mt, were carried out. Functional life span of the rat corpus luteum was estimated as 12 days post hCG treatment from plasma P4. As plasma P4 decreased, concomitant increase in plasma 20αOHP4 was observed. In vitro production of P5 in Mt correlated well with plasma P4 levels, indicating cholesterol side-chain cleavage reaction is the rate limiting step involved in ovarian steroidogenesis. In contrast, both values of Km and Vmax of CSCC did not change between day 7, on which the highest value of plasma P4 observed, and day 12, on which functional luteolysis was ascertained from plasma P4. Both FCh of Mt and the activity of 3βHSD in Ms remained unchanged during the functional life of the corpus luteum. The activity of 20αHSD in Sup increased from day 10 post hCG treatment in accordance with plasma 20αOHP4. These results indicate that 1) decrease in availability of FCh to CSCC within mitochondria and 2) increase in catabolism of P4 into inactive progestin, 20αOHP4, may play key roles in the functional luteolysis. To compare the events observed in functional luteolysis with PGF2α induced luteolysis, the animals were treated either with PGF2α (1 mg/rat; s.c.), cycloheximide (5 mg/rat; i.p.; CX), or vehicle on day 7 post hCG treatment, and sacrified 1 hour later for the analyses mentioned above. Both plasma P4 and in vitro production of Ps in Mt decreased significantly with PGF2a or CX treatment, whereas the FCh of Mt and the activity of CSCC remained unchanged with PGF2α treatment. Both plasma 20αOHP4 and 20αHSD in Sup increased with PGF2α treatment. However, CX suppressed both of them. These results seem to fit with the findings observed in the functional luteolysis, and therefore, it is suggested that PGF2α might be involved in luteolysis through the blockade of intramitochondrial cholesterol transport and the facilitation of catabolism of P4 into 20αOHP4.
