To assess the direct effect of growth hormone and growth hormone-releasing hormone on gene expression of steroidogenic enzymes and production of progesterone, 17-hydroxyprogesterone (17-OHP) and estradiol, we cultured luteinizing granulosa cells with or without follicle-stimulating hormone (FSH), growth hormone and growth hormone-releasing hormone at different concentrations. Luteinizing granulosa cells were obtained from women undergoing an in vitro fertilization program in the Department of Obstetrics and Gynecology, S. Raffaele Scientific Institute, Milan, Italy. At a concentration of 1 microgram/ml, FSH significantly increased estradiol production (2.1 +/- 0.7-fold the control value; p < 0.05 vs. control) and progesterone production (3.5 +/- 2.0-fold the control value; p < 0.05 vs. control). Growth hormone was effective on estradiol, progesterone and 17-OHP at 1 microgram/ml, enhancing estradiol production (1.3 +/- 0.2-fold the control value; p < 0.05 vs. control), progesterone production (2.5 +/- 1.0-fold the control value; p < 0.05 vs. control), and 17-OHP (1.4 +/- 0.2-fold the control value; p < 0.05 vs. control). Growth hormone-releasing hormone increased estradiol production (1.5 +/- 1.2-fold the control value) and progesterone production (1.3 +/- 0.8-fold the control value), but not significantly. No effects by growth hormone-releasing hormone were seen on 17-OHP production. FSH, growth hormone and growth hormone-releasing hormone did not increase P450scc and P450 aromatase mRNAs, whereas FSH increased P450c17 mRNA to 150% at 100 ng/ml and 1 microgram/ml, growth hormone increased it to 230% at 100 ng/ml and to 200% at 1 microgram/ml, and growth hormone-releasing hormone increased it to 140% at 100 ng/ml and to 190% of control values at 1 microgram/ml. These results indicate a direct effect of growth hormone on steroidogenesis by increasing P450c17 mRNA accumulation and progesterone, 17-OHP and estradiol production.