Lead increases free Ca2+ concentration in cultured osteoblastic bone cells: simultaneous detection of intracellular free Pb2+ by 19F NMR

Abstract

Lead (Pb) has been shown to perturb Ca-mediated cellular processes. However, to date, a direct effect of Pb on intracellular free Ca2+ concentration ([Ca2+]i) has not been demonstrated. 19F NMR in combination with 1,2-bis(2-amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid (5F-BAPTA) was used to simultaneously measure [Ca2+]i and intracellular free Pb2+ concentration ([Pb2+]i) in the rat osteoblastic bone cell line ROS 17/2.8. The basal concentration of [Ca2+]i in ROS 17/2.8 cells was measured to be 128 +/- 24 nM. Treatment with Pb2+ at 5 and 25 microM produced sustained 50% and 120% increases in [Ca2+]i, respectively, over a time course of 5 hr. At a medium Pb2+ concentration of 25 microM, the entry of Pb2+ into ROS 17/2.8 cells yielded measurable [Pb2+]i in cultured cells. Collectively, these findings advance the hypothesis that Pb toxicity is mediated, in part, through perturbations in [Ca2+]i.