Evaluation of in vitro culture conditions to demonstrate pregnancy-dependent changes in luteal function in the pig

This study was designed to evaluate the most effective in vitro culture conditions for study of pregnancy-dependent changes in LH-stimulated steroidogenesis in porcine luteal tissue. Mixed luteal cells recovered from the CL of cyclic (Day 12) and pregnant (Day 12 and 16) pigs were incubated either in Eagle's minimum essential medium (MEM) or in MEM supplemented with different concentrations of porcine or calf serum or BSA, or were incubated in a defined medium (DM) containing 2 micrograms/ml insulin, 40 ng/ml hydrocortisone, 5 micrograms/ml transferrin, and 10(-7) M thyroxine. Cultures were treated with various doses of porcine LH (0.1, 1, 10, 100 ng/ml). Additionally, large and small luteal cells of Day 12 cyclic and pregnant pigs were incubated for 4 h in medium 199 supplemented with 2% porcine serum in the absence or presence of various concentrations of LH. Overall, progesterone production by the luteal cells of Day 12 pregnant pigs was significantly (p 0.05) higher than that by cells of the cyclic pigs. However, the luteal cells of pregnant Day 16 pigs released significantly (p 0.05) less progesterone than the other groups. LH had a stimulatory effect (p 0.001 ) on progesterone release by mixed luteal cells that was independent of the type of medium supplementation or reproductive status. Porcine serum, calf serum. and BSA (p 0.001, p 0.001, and p 0.05, respectively) also stimulated progesterone production. However, the doses of porcine and calf serum or BSA that stimulated progesterone release were dependent on the reproductive status of the pig. Progesterone production by large luteal cells always exceeded (p 0.001 ) that of small cells and was stimulated by LH (p 0.007). Small luteal cells of both cyclic and pregnant (Day 12) gilts did not respond to LH treatment, and small luteal cells of cyclic pigs produced significantly (p 0.04) more progesterone than those of pregnant pigs