Recent studies have demonstrated that bone is highly innervated and contains neuromediators that have functional receptors on bone cells. However, no data exist concerning the quantitative changes of innervation during bone loss associated with estrogen withdrawal. To study the involvement of nerve fibers in the regulation of bone remodeling, we have evaluated the modifications of innervation in a classical in vivo model of osteopenia in rats, ovariectomy (OVX). Skeletal innervation was studied by immunocytochemistry using antibodies directed against specific neuronal markers, neurofilament 200 (NF200) and synaptophysin (SY), and the neuromediator glutamate (Glu). Sciatectomy, another model of bone loss due to limb denervation and paralysis, was used to validate our quantitative image analysis technique of immunostainings for nerve markers. Female wistar rats aged of 12 weeks were sham-operated (SHAM) or ovariectomized (OVX). Bone mineral density measurement and bone histomorphometry analysis of tibiae 14 days postsurgery demonstrated a significant bone loss in OVX compared with SHAM. We observed an important reduction of nerve profile density in tibiae of OVX animals compared with SHAM, while innervation density in skin and muscles was similar for OVX and control rats. Quantitative image analysis of immunostainings demonstrated a significant decrease of the percentage of immunolabelling per total bone volume of NF200, SY, and Glu in both the primary and secondary spongiosa of OVX rats compared with SHAM. These data indicate for the first time that ovariectomy- induced bone loss in rat tibiae is associated with a reduction in nerve profile density, suggesting a functional link between the nervous system and the bone loss after ovariectomy.