Pore properties and ionic block of the rabbit epithelial calcium channel expressed in HEK 293 cells.
We have used the whole-cell patch-clamp technique to analyse the permeation properties and ionic block of the epithelial Ca2+ channel ECaC heterologously expressed in human embryonic kidney (HEK) 293 cells. Cells dialysed with 10 mM BAPTA and exposed to Ca2+-containing, monovalent cation-free solutions displayed large inwardly rectifying currents. Their reversal potential depended on the extracellular Ca2+ concentration, [Ca2+]o. The slope of the relationship between reversal potential and [Ca2+]o on a logarithmic scale was 21 +/- 4 mV, compared with 29 mV as predicted by the Nernst equation (n = 3-5 cells). Currents in mixtures of Ca2+ and Na+ or Ca2+ and Ba2+ showed anomalous mole fraction behaviour. We have described the current-concentration plot for Ca2+ and Na+ by a kinetic permeation model, i.e. the "step" model. Extracellular Mg2+ blocked both divalent and monovalent currents with an IC50 of 62 +/- 9 microM(n = 4) in Ca2+-free conditions and 328 +/- 50 microM (n = 4-9) in 100 microM Ca2+ solutions. Mono- and divalent currents through ECaCs were blocked by gadolinium, lanthanum and cadmium, with a blocking order of Cd2+ >> Gd3+ > La3+. We conclude that the permeation of monovalent and divalent cations through ECaCs shows similarities with L-type voltage-gated Ca2+ channels, the main differences being a higher Ca2+ affinity and a significantly higher current density in micromolar Ca2+ concentrations in the case of ECaCs.
Vennekens
R
R
Department of Physiology, Campus Gasthuisberg, KULeuven, Leuven, Belgium.
Prenen
J
J
Hoenderop
J G
JG
Bindels
R J
RJ
Droogmans
G
G
Nilius
B
B
D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D002412:Cations / Q000494:pharmacology; D002413:Cations, Divalent / Q000037:antagonists & inhibitors / Q000378:metabolism; D002414:Cations, Monovalent / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D004553:Electric Conductivity; D004622:Embryo, Mammalian / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D007477:Ions; D007668:Kidney / Q000166:cytology / Q000196:embryology*; D011817:Rabbits; D012964:Sodium / Q000494:pharmacology
D015220:Calcium Channels; D002412:Cations; D002413:Cations, Divalent; D002414:Cations, Monovalent; D007477:Ions; D012964:Sodium