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Permeation and gating properties of the novel epithelial Ca(2+) channel Vennekens R; Hoenderop JG; Prenen J; Stuiver M; Willems PH; Droogmans G; Nilius B; Bindels RJJ Biol Chem 2000[Feb]; 275 (6): 3963-9The recently cloned epithelial Ca(2+) channel (ECaC) constitutes the Ca(2+) influx pathway in 1,25-dihydroxyvitamin D(3)-responsive epithelia. We have combined patch-clamp analysis and fura-2 fluorescence microscopy to functionally characterize ECaC heterologously expressed in HEK293 cells. The intracellular Ca(2+) concentration in ECaC-expressing cells was closely correlated with the applied electrochemical Ca(2+) gradient, demonstrating the distinctive Ca(2+) permeability and constitutive activation of ECaC. Cells dialyzed with 10 mM 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid displayed large inward currents through ECaC in response to voltage ramps. The corresponding current-voltage relationship showed pronounced inward rectification. Currents evoked by voltage steps to potentials below -40 mV partially inactivated with a biexponential time course. This inactivation was less pronounced if Ba(2+) or Sr(2+) replaced Ca(2+) and was absent in Ca(2+)-free solutions. ECaC showed an anomalous mole fraction behavior. The permeability ratio P(Ca):P(Na) calculated from the reversal potential at 30 mM [Ca(2+)](o) was larger than 100. The divalent cation selectivity profile is Ca(2+) > Mn(2+) > Ba(2+) approximately Sr(2+). Repetitive stimulation of ECaC-expressing cells induced a decay of the current response, which was greatly reduced if Ca(2+) was replaced by Ba(2+) and was virtually abolished if [Ca(2+)](o) was lowered to 1 nM. In conclusion, ECaC is a Ca(2+) selective channel, exhibiting Ca(2+)-dependent autoregulatory mechanisms, including fast inactivation and slow down-regulation.|Calcium Channels/*metabolism[MESH]|Calcium/metabolism[MESH]|Cations, Divalent/metabolism[MESH]|Cell Line[MESH]|Chelating Agents/pharmacology[MESH]|Fura-2[MESH]|Humans[MESH]|Ion Channel Gating/*physiology[MESH]|Microscopy, Fluorescence[MESH]|Patch-Clamp Techniques[MESH]|TRPV Cation Channels[MESH] |
