Multiple types of chloride channels in bovine pulmonary artery endothelial cells.
We have characterized two different types of Cl- currents in calf pulmonary artery endothelial (CPAE) cells by using a combined patch-clamp and Fura-2 microfluorescence technique to measure simultaneously ionic currents and the intracellular Ca2+ concentration, [Ca2+]i. Exposure of CPAE cells to 28% hypotonic solution induces cell swelling without a change in membrane capacitance and [Ca2+]i, and concomitantly activates a current. This current, I(Cl, vol), is closely correlated with the changes in cell volume and shows a modest outward rectification. It slowly inactivates at potentials more positive than +60 mV but is time- and voltage-independent at other potentials. Increase in [Ca2+]i by different maneuvers, such as application of vasoactive agonists (ATP), ionomycin, or loading of the cells directly with Ca2+ also activates a Cl- current, I(Cl, Ca). This current slowly activates at positive potentials, inactivates quickly at negative potentials and shows strong outward rectification. A time-independent component of the current activated by elevation of [Ca2+]i alone can be inhibited by cell shrinking by exposing the cells to hypertonic solution, indicating that an increase in [Ca2+]i also co-activates I(Cl, vol). Forskolin or cAMP never activated a current in CPAE cells, which indicates the lack of cAMP-activated channels in these cells. There is also no evidence for the existence of voltage-gated Cl- channels in resting, nonstimulated cells. Challenging a cell with elevated [Ca2+]i and hypotonic solutions activated I(Cl, vol) on top of I(Cl, Ca), suggesting that I(Cl, Ca) and I(Cl, vol) are different channels. We conclude that CPAE cells do not express voltage-gated (ClC-type) or cAMP-gated (CFTR-type) Cl- channels, but activate large Cl- currents after volume (mechanical?) or chemical (Ca2+) stimulation.
Nilius
B
B
KU Leuven, Laboratorium voor Fysiologie, Campus Gasthuisberg, Belgium. bernd.nilius@med.kuleuven.ac.be
Szücs
G
G
Heinke
S
S
Voets
T
T
Droogmans
G
G
D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D000242:Cyclic AMP / Q000494:pharmacology; D004553:Electric Conductivity; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D015640:Ion Channel Gating; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism*
D018118:Chloride Channels; D000242:Cyclic AMP; D002118:Calcium