In the corpus luteum, prostaglandin F2α (PGF2α) appears to be a physiological agent with both antisteroidogenic and luteolytic actions. It is hypothesized that the antisteroidogenic action of PGF2α is through altered transport of cholesterol to the mitochondrial cytochrome P450 side chain cleavage enzyme (P450scc). However, the effect of PGF2α on the cholesterol transport protein, sterol carrier protein-2 (SCP2; 13.2 kDa) expression, requires analysis. In this study, the decline in serum progesterone following PGF2α injection was examined in parallel with altered ovarian SCP2, P450scc, and 3β-hydroxysteroid dehydrogenase (3β-HSD) protein and mRNA levels. Rats (28 days old) were treated with 8 IU pregnant mare serum gonadotropin to induce follicular development and ovulation. Ten days after ovulation, animals were treated +/- PGF2α (single injection; 250 μg). Ovarian SCP2, P450scc, and 3β-HSD mRNA levels were examined at 0 (t 0 ), 4, and 8 hours post-PGF2α treatment using Western and Northern blot analysis. SCP2 mRNA levels were also examined using a ribonuclease protection assay (RPA) which detects a 429 bp SCP2-mRNA specific sequence. Results indicated that serum progesterone was significantly reduced 8 hours after PGF2α-injection (30.8 ± 4.5 ng/ml, n=14 vs. 130.2 ± 8.0 ng/ml control, n=6, p<0.001). The decline in progesterone paralleled a 50-60% reduction in 3β-HSD protein and mRNA levels by 4 hours post-PGF2α (n=6). P450scc expression was also reduced at 4 hours (44-54%) after which both protein and mRNA levels increased 1.7- and 2-fold, respectively at 8 hours post-PGF2α relative to t 0 (p<0.02; n=6). In contrast, the 0.8 kb SCP2-specific mRNA transcript was reduced to 50-80% of the pre-PGF2α treatment level (p<0.01, n=3). SCP2 RPA analysis indicated that SCP2 mRNA levels were reduced 65 and 85% by 4 and 8 hours post-PGF2α compared to t 0 ovarian tissue (p<0.001, n=4). Consistent with the loss of SCP2 mRNA expression, Western blot analysis indicated that a 15 kDa SCP2 immunoreactive protein (presumably the pro-SCP2 form) was significantly reduced or absent in the PGF2α treated animals (p<0.04; n=3). These results are the first to demonstrate that ovarian SCP2 expression is significantly altered following PGF2α-treatment and this study confirms that PGF2α alters ovarian cholesterol transport capacity as part of its antisteroidogenic action.