Swelling-activated K+ fluxes in vascular endothelial cells: role of intracellular Ca2+.
Swelling of bovine aortic endothelial cells activates Ca(2+)-dependent K+ channels. To determine the role of Ca2+ in this response, we examined the effect of cell swelling on intracellular Ca2+ concentration ([Ca2+]i), and the role of [Ca2+]i in swelling-activated K+ efflux. Basal [Ca2+]i, measured by fura 2 fluorescence, was 62 nM and increased by 36 nM in hypotonic medium (220 mosmol/l) compared with a 277 nM increase in response to extracellular ATP. In cells loaded with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid (BAPTA), the increases induced by swelling and by ATP were reduced to 13 and 20 nM, respectively. Exposure to hypotonic medium (220 mosmol/kg) or to the Ca2+ ionophore A-23187 stimulated a furosemide-insensitive 86Rb efflux consistent with activation of K+ channels. The swelling-activated efflux was inhibited 16% by 5 mM tetraethylammonium and 24% by 23 mM tetrabutylammonium, but not by 100 microM quinidine, a pattern similar to that previously observed for swelling-activated K+ channels in cell-attached patches. The effects of A-23187 and hypotonic swelling on 86Rb efflux were completely additive, suggesting Ca(2+)-independent activation by cell swelling. Removal of Ca2+ from the external medium or loading of cells with BAPTA to buffer intracellular Ca2+ blocked the activation of 86Rb efflux by A-23187, but not by hypotonic swelling. Hypertonic medium (440 mosmol/kg by the addition of sucrose) attenuated the increased 86Rb efflux in response to A-23187. We conclude that the activation of K+ efflux in swollen endothelial cells occurs independently of changes in [Ca2+]i.(ABSTRACT TRUNCATED AT 250 WORDS)
Perry
P B
PB
Department of Medicine, Emory University School of Medicine, Atlanta, Georgia 30322.
O'Neill
W C
WC
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D002417:Cattle; D002478:Cells, Cultured; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000502:physiology*
D015221:Potassium Channels; D000001:Calcimycin; D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D011188:Potassium; D002118:Calcium