Steroidogenic effect of 17 beta-estradiol on rabbit luteal cells in vitro: estrogen-induced maintenance of progesterone production.

Previous studies have established that 17 beta-estradiol is the principal luteotropic hormone in the rabbit. However, a direct effect of 17 beta-estradiol on rabbit luteal cell progesterone production has been difficult to show in vitro. The goal of this study was to develop a system in which the effect of estrogen on luteal cell progesterone production could be studied in vitro. To that end, a dissociated rabbit luteal cell preparation was developed using collagenase and the resultant isolated cells were studied using a perifusion system. Optimization of the cell digest procedure revealed that: inclusion of 2% bovine serum albumin in our optimal dissociation medium increased cell yield; and animals killed by cervical dislocation maintained more stable levels of progesterone during a 7-h perifusion compared to animals killed with barbituate-induced euthanasia (euthobarb). When dissociated luteal cells were perifused with medium, stable progesterone output (greater than 80% of initial levels) was observed for 5-6 h, after which medium progesterone concentrations declined. The inclusion of 17 beta-estradiol (10(-8) M) in the perifusion medium maintained progesterone output at control levels for up to 15 h. However, the maintenance of progesterone was not noted until after 5 h of perifusion, suggesting that the effect of estradiol may be time dependent. Thus, this investigation describes a rabbit luteal cell dissociation technique and perifusion system that may be used to examine the mechanism through which estradiol acts to maintain rabbit luteal progesterone production.