Depolarization-mediated inhibition of Ca(2+) entry in endothelial cells.
The effect of extracellular Cl(-) in regulating ACh-induced Ca(2+) entry into freshly isolated rabbit aortic endothelial cells was studied using Ca(2+)-sensitive fluorescence microscopy and patch-clamp electrophysiology. After ACh caused transient Ca(2+) release in Ca(2+)-free medium, readdition of 3 mM Ca(2+) to the bath maintained Ca(2+) entry. Removal of extracellular Cl(-) abolished the plateau phase in Ca(2+) signal and inhibited divalent cation entry. However, in the presence of the K(+) ionophore valinomycin, removal of Cl(-) had no effect on the Ca(2+) plateau. Under current-clamp conditions, substitution of gluconate for Cl(-) induced membrane depolarization. Under voltage clamp, with CsCl in the pipette, ACh activated a slowly developing Cl(-) current, which was blocked by SITS and 5-nitro-2-(3-phenylpropylamino)benzoic acid. Varying the membrane potential by utilizing different extracellular K(+) concentrations in the presence of 5 microM valinomycin demonstrated that depolarization blocked ACh-stimulated Mn(2+) entry. These data suggest that ACh-induced Ca(2+) entry in freshly isolated endothelial cells requires the presence of extracellular Cl(-) to maintain a polarized membrane potential.
Wang
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Vancouver Vascular Biology Research Center, Department of Pharmacology, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3.
van Breemen
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D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000502:physiology; D002712:Chlorides / Q000502:physiology; D004553:Electric Conductivity; D004594:Electrophysiology; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D005110:Extracellular Space / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology; D011188:Potassium / Q000378:metabolism; D011817:Rabbits; D014634:Valinomycin / Q000494:pharmacology
D002712:Chlorides; D014634:Valinomycin; D000109:Acetylcholine; D011188:Potassium; D002118:Calcium