The increase in serum FSH that accompanies female reproductive aging occurs before changes in estradiol (E2). A decrease in negative feedback from inhibin A (a product of the dominant follicle and corpus luteum) and/or inhibin B (secreted by developing follicles) may explain the rise in FSH with age. To test the hypothesis that decreases in inhibin A or inhibin B occur at an age at which the first increase in follicular phase FSH is evident, daily blood samples were obtained across the menstrual cycle from younger (<35 yr; n = 23) and older (35-46 yr; n = 21) cycling women. These cross-sectional studies were complemented by longitudinal data in 3 women studied at a 10-yr interval. In the early follicular phase, mean inhibin B was lower in older cycling women (88 +/- 7 vs. 112 +/- 10 pg/mL; P < 0.05) and FSH was higher (13.0 +/- 0.5 vs. 11.2 +/- 0.7 IU/L in older vs. younger, respectively; P < 0.04). In the mid- and late follicular phases, inhibin B was also lower in the older women (117 +/- 9 vs. 146 +/- 10 and 85 +/- 8 vs. 117 +/- 11 pg/mL; P < 0.04), whereas E2 was higher (105 +/- 14 vs. 68 +/- 5 and 240 +/- 27 vs. 163 +/- 9 pg/mL; P < 0.02), and no differences in FSH were observed in the two groups at these times. In women studied longitudinally, FSH and inhibin B varied inversely in the follicular phase. In the early luteal phase, mean inhibin B was lower in the older group (64 +/- 6 vs. 94 +/- 12 pg/mL; P < 0.03), and FSH was higher (12.5 +/- 1.0 vs. 9.7 +/- 0.6 IU/L; P < 0.03). In the mid- and late luteal phases, inhibin B was also lower in older subjects (21 +/- 2 vs. 33 +/- 5 and 22 +/- 2 vs. 36 +/- 6 pg/mL; P < 0.02). No difference in inhibin A, E2, or progesterone was observed across the luteal phase, between the two groups. However, in all subjects studied longitudinally, increased age was associated with a decrease in inhibin A, inhibin B, and progesterone in the absence of changes in E2. Our conclusions were: 1) reproductive aging is accompanied by decreases in both inhibin B and inhibin A; 2) the decrease in inhibin B precedes the decrease in inhibin A and occurs in concert with an increase in E2, suggesting that inhibin B negative feedback is the most important factor controlling the earliest increase in FSH with aging; 3) these studies suggest that the decrease in inhibin B is the earliest marker of the decline in follicle number across reproductive aging.