Time- and dose-related effects of estradiol and diethylstilbestrol on the morphology and function of the fetal rat testis in culture

The mechanisms underlying the action of estrogens in the fetal testis are still largely obscure. In particular, whether this action is direct or indirect remains largely unexplored. This study was aimed at investigating the effect of estradiol (E-2) and diethylstilbestrol (DES) on the testis from 14.5-day-old rat fetuses in culture, at concentrations ranging from 4 x 10(-10) M (K-d of E-2 for the estrogen receptors [ER]: 1-4 x 10(-10) M) to 4 x 10(-6) M (concentration previously shown in the literature to affect in vitro gonocyte proliferation). Exposure to DES and E-2 decreased gonocyte number, the effects of DES being much more drastic than those of E-2. Gonocyte number decreased in a concentration-dependent manner (day 3: -5%, -16%, and -80% at 4 x 10(-10) M, 4 x 10(-8) M, and 4 x 10(-6)M of DES, respectively), as well as in a time-dependent manner (at 4 x 10(-6) M DES: -31% on day 1, -60% on day 2, and -80% on day 3). This was due to a decrease in the gonocyte mitotic index and a dramatic increase in apoptosis. Importantly, in the presence of the anti-estrogen ICI 182.780 (ICI), the effect of DES was abolished. Sertoli cell number subsequently decreased (day 3), although the rate of apoptosis did not increase. These changes were less dramatic than those observed with gonocytes and were due to a decrease in Sertoli cell proliferation, which was not antagonized by ICI. Addition of 4 x 10(-6) M DES had no effect on basal Sertoli cell cyclic adenosine 5'-monophosphate (cAMP) levels or on follicle-stimulating hormone (FSH)-stimulated cAMP production after adjustment for Sertoli cell number. Finally, estrogens reduced both Leydig cell number (-26% on day 3, 4 x 10(-6) M DES) and basal and luteinizing hormone (LH)-stimulated testosterone production. The latter effects were antagonized by ICI. In conclusion: 1) E-2 and DES induce alterations in the germ line and in somatic cells; 2) gonocyte alteration was the first event detected, and the action of estrogens at this level was mediated by ER, as is the case in Leydig cells; and 3) these data should help us to understand estrogen effects on the fetus and should be considered in the context of the debate on environmental estrogens.