Dichlorobenzamil inhibits stimulated bone resorption in vitro

We have previously proposed that stimulated release of Ca from bone requires Na-Ca exchange. 3',4'-Dichlorobenzamil (DCB), an amiloride analog, has been shown to directly block Na-Ca exchange in cardiac tissue. To further test our hypothesis we have characterized the effect of DCB on basal and stimulated bone resorption from neonatal mouse calvaria in vitro. DCB inhibition of resorption from bones stimulated with 1 nM PTH was dose dependent. The IC50 was about 7 microM, and complete inhibition occurred at 10 microM. DCB alone inhibited basal Ca release at 10 microM. Amiloride, which is less potent as an inhibitor of Na-Ca exchange, had no effect on PTH-stimulated resorption at concentrations lower than 0.1 mM, but inhibited basal resorption at 10 microM. Stimulated Ca release was inhibited either by continuous treatment with DCB plus PTH for 72 h or by a short pretreatment with DCB alone, followed by removal of DCB before addition of PTH. At least 9-h pretreatment with DCB was necessary to block the subsequent response to PTH. The inhibitory effect of DCB pretreatment could be prevented if PTH was present together with DCB during pretreatment periods of 24 h or less. This reversibility suggests that the inhibition by DCB is not simply a toxic effect of the drug. DCB also inhibited resorption stimulated by 1,25-dihydroxyvitamin D3 or prostaglandin E2, which indicates that the effect of DCB is beyond the level of specific hormone-receptor interaction. Thus, the data are consistent with a role for Na-Ca exchange in the process of hormonally stimulated bone resorption.