Size Distribution and Hormonal Responsiveness of Dispersed Rabbit Luteal Cells during Pseudopregnancy1

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Abstract
Due to the evidence of two distinct steroidogenic cell types in corpora lutea of large domestic animals, cells of the rabbit corpus luteum were characterized with respect to cell diameters, relative abundance, steroidogenic capacity and responsiveness to hormones. Pseudopregnancy was induced in New Zealand rabbits by injection of 30-160 IU pregnant mare''s serum gonadotropin (PMSG) followed in 2-4 days by an i.m. injection of 20-35 .mu.g gonadotropin-releasing hormone (GnRH). Corpora lutea were obtained 2, 5 and 9 days after injection of GnRH and dissociated into single cell suspensions. Suspended steroidogenic cells were incubated (2 b, 37.degree. C) in medium 199 alone or in medium containing ovine luteinizing hormone (oLH) (100 ng/ml), or isoproterenol (100 .mu.M). Media were collected and assayed for progesterone content. Secretion of progesterone (.hivin.x .+-. SE, n = 4) was stimulated (p < 0.05) by oLH on each day: Day 2 = 1.7 .+-. 0.2-fold; Day 5 = 3.5 .+-. 0.4-fold; and Day 9 = 3.1 .+-. 0.6-fold stimulation above controls. Isoproterenol also stimulated (p < 0.05) secretion of progesterone by suspended luteal cells on Days 2 and 9. Microscopic examination of cell suspensions stained for 3.beta.-hydroxysteroid dehydrogenase (3.beta.HSD) activity provided identification of cells with steroidogenic capacity. The diameters (.hivin.x .+-. SE) for steroidogenic cells increased (p < 0.05) from Days 2 to 9 (Day 2 = 15.2 .+-. 0.2 .mu.m; Day 5 = 22.4 .+-. 0.4 .mu.m; Day 9 = 28.3 .+-. 1.6 .mu.m). The large cell to small cell ratio increased from 0.01 on Day 2 to 2.03 on Day 9. Cells collected on Day 9 were separated by elutriation into two size classes (small .ltoreq. 22 .mu.m and large > 22 .mu.m). Secretion of progesterone was stimulated (p < 0.05) in small cells incubated with oLH (100 ng/ml; 14.7-fold increase) or isoproterenol (100 .mu.m; 6.3-fold increase); secretion of progesterone in large cells was not significantly enhanced by either secretagogue. Thus, the corpus luteum of the Day 9 pseudopregnant rabbit contains steroidogenic cells of heterogeneous sizes, and small and large cell fractions respond differently to oLH and isoproterenol. These results suggest that two populations of steroidogenic luteal cells exist in the rabbit corpus luteum; these populations may differ in the mechanisms whereby the secretion of progesterone is regulated.