We studied the time frame of suppression and recovery of estradiol after injection with leuprolide acetate utilizing an ultrasensitive recombinant cell bioassay for estradiol in eight normal premenopausal women. Previous studies have shown suppression of gonadotropins and estradiol at 4 weeks after the depot injection, but no studies have shown the weekly time course of estradiol suppression or recovery. Four women received one 3.75 mg i.m. injection of leuprolide acetate and four received two 3.75 mg doses of leuprolide acetate 4 weeks apart. Estradiol, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels were measured weekly for 8 to 12 weeks. Estradiol was significantly suppressed to 26.6 +/- 19.3% of baseline values by week 3 after the initial dose of leuprolide acetate and suppressed to 2.7 +/- 3.1% of baseline values by week 4 (p < 0.01 versus baseline). The actual values were less than 14.7 pmol/l (4 pg/ml) in all women by week 4. Estradiol remained suppressed for 8 weeks after one dose of leuprolide acetate and remained suppressed for 6 weeks after a second dose administered 4 weeks later. LH and FSH followed a similar pattern, but only remained suppressed for 7 weeks after one dose of leuprolide acetate and for 6 weeks after two doses. Estradiol levels at baseline were significantly correlated with body mass index (BMI). We also studied one postmenopausal woman. Her baseline estradiol levels were 10.3 pmol/l (2.8 pg/ml) and were suppressed to 3.9 pmol/l (1.1 pg/ml) by 2 weeks after leuprolide acetate. In conclusion, estradiol was suppressed to postmenopausal levels by the end of the first month of treatment with leuprolide acetate, as determined by an ultrasensitive bioassay. Higher doses would need to be tested to determine whether greater suppression can be achieved. The hypothalamic-pituitary-gonadal axis begins to recover 7 weeks after one dose and 6 weeks after a second dose of leuprolide acetate. This confirms the adequacy of 4-week dosing to maintain estradiol and gonadotropin suppression in adult women treated with leuprolide acetate, but raises the question whether less frequent dosing may be possible in some situations, or whether higher doses may be needed in some situations for an even greater degree of estradiol suppression.