Permeation and gating properties of the novel epithelial Ca2+ channel

The recently cloned epithelial Ca2+ channel (ECaC) constitutes the Ca2+ influx pathway in 1,25-dihydroxyvitamin Ds-responsive epithelia, We have combined patch-clamp analysis and fura-a fluorescence microscopy to functionally characterize ECaC heterologously expressed in HEK293 cells. The intracellular Ca2+ concentration in ECaC-expressing cells was closely correlated with the applied electrochemical Ca2+ gradient, demonstrating the distinctive Ca2+ permeability and constitutive-activation of ECaC, Cells dialyzed with 10 mn 1,2-bis(2-aminoph- enoxy)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 timecourse, This inactivation was less pronounced if Ba2+ or Sr2+ replaced Ca2+ and was absent in Ca2+-free solutions, ECaC showed an anomalous mole fraction behavior. The permeability ratio P-CA:P-NA calculated from the reversal potential at 30 mM [Ca2+], was larger than 100, The divalent cation selectivity profile is Ca2+ > Mn2+ > Ba2+ similar to Sr2+. Repetitive stimulation of ECaC-expressing cells induced a decay of the current response, which was greatly reduced if Ca2+ was replaced by Ba2+ and was virtually abolished if [Ca2+], was lowered to 1 nar. In conclusion, ECaC is a Ca2+ selective channel, exhibiting Ca2+-dependent autoregulatory mechanisms, including fast inactivation and slow down-regulation.