Regulation of aromatase gene expression in purified rat Leydig cells has not yet been investigated. Therefore, using a highly specific quantitative RT-PCR method, we have measured the amount of cytochrome P450 aromatase (P450arom) mRNA and aromatase activity in mature rat Leydig cells submitted to various treatments during 24 h. Estradiol production was enhanced in a dose-related manner in the presence of testosterone, the maximum (28% increase) being obtained with 200 ng/ml. Related to the P450arom mRNA levels, a decrease was observed in the presence of low concentrations (50 and 100 ng/ml) of testosterone, then a 20% increase of the amount of transcripts was recorded for the higher concentrations (200-500 ng/ml). The same result was obtained in the presence of 5alpha-dihydrotestosterone (an androgen resistant to aromatase activity). The addition of ovine LH (oLH; 0.1-50 ng/ml) to the Leydig cell culture medium induced a dose-related augmentation of estradiol output up to 10 ng/ml oLH, although a decrease was observed with 50 ng/ml when compared with maximal values. mRNA levels slightly decreased in the presence of low concentrations (0.1-1 ng/ml) of oLH, an effect that was abolished by the addition of testosterone; mRNA levels were increased by oLH (5-10 ng/ml) 35 and 75% respectively in the absence and presence of testosterone (when compared with Leydig cells incubated without treatment). With 50 ng/ml oLH, a large augmentation (twofold) of the P450arom mRNA level either without or with testosterone was observed. Dibutyryl cyclic AMP (1 mM) mimicked the effect of oLH. The half-life of the P450arom mRNAs was twofold increased in the presence of testosterone and oLH when compared with the half-life in the absence of treatment (5.8+/-0.6 h). Taken together, our data have demonstrated that, in freshly isolated Leydig cells from mature rat testes, the regulation of aromatase expression and enzymatic activity is under LH (through cyclic AMP) and steroid control; moreover seminiferous tubule-secreted factor(s) are also involved. Therefore, rat Leydig cell aromatase is controlled at both transcriptional and post-transcriptional steps by endocrine and/or locally produced modulators.

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