In vitro studies on bone resorption in neonatal mouse calvariae using a modified dissection technique giving four samples of bone from each calvaria

Abstract

Bone resorption in a modified bone culture system, based on incubation of small fragments from neonatal mouse calvarial bones, has been studied. Four bone fragments were dissected out from each mouse calvaria and were thereafter cultured in CMRL 1066 medium in plastic multiwell dishes. Bone resorption was assessed by 45Ca release from prelabeled bones. The rate of bone resorption in response to parathyroid hormone (PTH) was less in the anterior part of the calvaria compared to the posterior part. After removing the anterior region, four parietal bone fragments that showed identical basal and PTH-stimulated release of 45Ca could be dissected out from each mouse. Excretion of lactate dehydrogenase and beta-glucuronidase was the same in bones cultured submerged or on grids. Uptake of [3H]thymidine in bones cultured submerged was 54% of [3H]thymidine uptake in bones cultured on grids. Dose-response curves, established by using parietal bone fragments, showed that the sensitivity and the magnitude of the increase in 45Ca release seen after stimulation with PTH, prostaglandin E2, and 1 alpha-hydroxyvitamin D3 were the same for bones cultured submerged or on grids. The 45Ca release in response to stimulation with PTH, prostaglandin E2, and 1 alpha-OHD3 was the same in calvarial fragments cultured submerged and those previously obtained with calvarial halves cultured on grids. Thus, even though the rate of DNA synthesis was slower in bones cultured submerged, the rate and the magnitude of resorption were the same in bones cultured on grids or submerged. These data show that it is possible to perform studies on bone resorption with small fragments of neonatal mouse parietal bones.