Insulin-like growth factor-I (IGF-I) is a key factor in bone remodeling. In osteoblasts, IGF-I synthesis is enhanced by parathyroid hormone and prostaglandin E 2 (PGE 2 ) through cAMP-activated protein kinase. In rats, estrogen loss after ovariectomy leads to a rise in serum IGF-I and an increase in bone remodeling, both of which are reversed by estrogen treatment. To examine estrogendependent regulation of IGF-I expression at the molecular level, primary fetal rat osteoblasts were co-transfected with the estrogen receptor (hER, to ensure active ER expression), and luciferase reporter plasmids controlled by promoter 1 of the rat IGF-I gene (IGF-I P1), used exclusively in these cells. As reported, 1 M PGE 2 increased IGF-I P1 activity by 5-fold. 17-Estradiol alone had no effect, but dose-dependently suppressed the stimulatory effect of PGE 2 by up to 90% (ED 50 ϳ0.1 nM). This occurred within 3 h, persisted for at least 16 h, required ER, and appeared specific, since 17␣-estradiol was 100 -300-fold less effective. By contrast, 17-estradiol stimulated estrogen response element (ERE)-dependent reporter expression by up to 10-fold. 17-Estradiol also suppressed an IGF-I P1 construct retaining only minimal promoter sequence required for cAMP-dependent gene activation, but did not affect the 60-fold increase in cAMP induced by PGE 2 . There is no consensus ERE in rat IGF-I P1, suggesting novel downstream interactions in the cAMP pathway that normally enhances IGF-I expression in skeletal cells. To explore this, nuclear extract from osteoblasts expressing hER were examined by electrophoretic mobility shift assay using the atypical cAMP response element in IGF-I P1. Estrogen alone did not cause DNA-protein binding, while PGE 2 induced a characteristic gel shift complex. Co-treatment with both hormones caused a gel shift greatly diminished in intensity, consistent with their combined effects on IGF-I promoter activity. Nonetheless, hER did not bind IGF-I cAMP response element or any adjacent sequences. These results provide new molecular evidence that estrogen may temper the biological effects of hormones acting through cAMP to regulate skeletal IGF-I expression and activity.Although postmenopausal osteoporosis is one of the most prevalent age-related skeletal disorders, our understanding of the role of estrogen, which has a critical role in maintaining bone mass, remains incomplete. The principal laboratory animal model of postmenopausal osteoporosis is the ovariectomized (OVX) 1 rat. One consistent observation with this model is an increase in the rate of bone remodeling after OVX-induced estrogen depletion (1-5). Furthermore, consistent with a cause and effect relationship, estrogen administration re-establishes a reduced rate of bone remodeling in the OVX rat.Bone remodeling consists of two opposing events, i.e. bone resorption and formation (6). Skeletal integrity in adults relies on closely coupled remodeling where there is a balance between these catabolic and anabolic processes. Net bone loss in postmenopausal o...