We investigated whether intestinal calcium absorption and serum 1,25-dihydroxycholecalciferol (calcitriol) concentrations are higher in women during lactation and after weaning to compensate for calcium lost in breast milk. Measurements were obtained at 4.6 mo postpartum in 24 lactating women and 24 nonlactating women, at 9.6 mo postpartum in 24 lactating women (2.6 mo after complete weaning) and 24 nonlactating women. One-half of the women in each group were randomly assigned to receive 1 g supplemental Ca/d as calcium carbonate. Fractional calcium absorption was measured by using stable isotopic tracers 42Ca and 44Ca. Fractional absorption was 0.32+/-0.02 (+/-SEM) in both lactating and nonlactating women, but was higher in lactating women after weaning (0.37+/-0.02) compared with nonlactating postpartum control subjects (0.31+/-0.02). These effects were independent of calcium intake. Changes in serum calcitriol paralleled changes in fractional absorption. There were no differences in calcitriol concentrations between lactating and nonlactating women, but calcitriol was greater in women after weaning compared with postpartum control subjects. Lactating women who had resumed menses had higher fractional absorption and serum calcitriol than did lactating women who had not. Serum calcium and phosphorus concentrations were greater in lactating compared with nonlactating women; there were no differences between groups after weaning. We conclude that lactation stimulates increases in fractional calcium absorption and serum calcitriol, but the responses are only apparent after weaning or the resumption of menses.