Extracellular Ca2+-sensing receptor is a promiscuous divalent cation sensor that responds to lead

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Abstract
The extracellular Ca2+-sensing receptor (CaR) responds to polycations, including Ca2+and neomycin. This receptor is a physiological regulator of systemic Ca2+metabolism and may also mediate the toxic effects of hypercalcemia. A number of divalent cations, including Pb2+, Co2+, Cd2+, and Fe2+, are toxic to the kidney, brain, and other tissues where the CaR is expressed. To determine which divalent cations can activate the CaR, we expressed the human CaR in HEK-293 cells and measured activation of phospholipase A2(PLA2) and the mitogen-activated protein kinase p42ERK in response to potential agonists for the receptor. HEK-293 cells expressing the nonfunctional mutant CaR R796W served as controls. Extracellular Ca2+, Ba2+, Cd2+, Co2+, Fe2+, Gd3+, Ni2+, Pb2+, and neomycin activated the CaR, but Hg2+and Fe3+did not. We analyzed the kinetics of activation of p42ERK and PLA2by the CaR in response to Ca2+, Co2+, and Pb2+. The EC50values ranged from ∼0.1 mM for Pb2+to ∼4.0 mM for Ca2+. The Hill coefficients were >3, indicating multiple cooperative ligand binding sites or subunits. Submaximal concentrations of Ca2+and Pb2+were additive for activation of the CaR. The EC50for Ca2+or Pb2+was reduced four- to fivefold by the presence of the other ion. These divalent cations also activated PLA2via the CaR in Madin-Darby canine kidney cells that stably express the CaR. We conclude that many divalent cations activate the CaR and that their effects are additive. The facts that the CaR is a promiscuous polycation sensor and that the effects of these ions are additive to activate it suggest that the CaR may contribute to the toxicity of some heavy metals such as Pb2+, Cd2+, Co2+, and Fe2+for the kidney and other tissues where it is expressed.