MULTIPLE TYPES OF CHLORIDE CHANNELS IN BOVINE PULMONARY-ARTERY ENDOTHELIAL-CELLS

We have characterized two different types of Cl- currents in calf pulm onary 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). Exposur e of CPAE cells to 28% hypotonic solution induces cell swelling withou t a change in membrane capacitance and [Ca2+](i), and concomitantly ac tivates a current, This current, I-Cl,I-vol, is closely correlated wit h 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 acti vates a Cl- current, I-Cl,I-Ca. This current slowly activates at posit ive potentials, inactivates quickly at negative potentials and shows s trong outward rectification, A time-independent component of the curre nt activated by elevation of [Ca2+](i) alone can be inhibited by cell shrinking by exposing the cells to hypertonic solution, indicating tha t an increase in [Ca2+](i) also co-activates I-Cl,I-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, nonstimula ted cells, Challenging a cell with elevated [Ca2+](i) and hypotonic so lutions activated I-Cl,I-vol on top of I-Cl,I-Ca, suggesting that I-Cl ,I-Ca and I-Cl,I-vol are different channels, We conclude that CPAE cel ls do not express voltage-gated (ClC-type) or cAMP-gated (CFTR-type) C l- channels, but activate large Cl- currents after volume (mechanical? ) or chemical (Ca2+) stimulation.