The perforated-patch technique was used to study the response of human bronchial cells to extracellular nucleotides. ATP or UTP (100 μm) elicited a complex response consisting of a large transient membrane current increase followed by a relatively small sustained level. These two phases were characterized by different current kinetics. Throughout the transient phase (2–3 min) the membrane current (I p ) displayed slow activation and deactivation kinetics at depolarizing and hyperpolarizing potentials respectively. At steady-state (I s ) the relaxation at hyperpolarizing potential disappeared whereas at positive membrane potentials the current became slightly deactivating. The I s amplitude was dependent on the extracellular Ca2+ concentration, being completely inhibited in Ca2+-free medium. Cell pre-incubation with the membrane-permeable chelating agent BAPTA/AM prevented completely the response to nucleotides, thus suggesting that both I p and I s were dependent on intracellular Ca2+. The presence of a hypertonic medium during nucleotide stimulation abolished I s leaving I p unchanged. On the contrary, niflumic acid, a blocker of Ca2+-activated Cl− channels, prevented completely I p without reducing significantly I s . 1,9-dideoxyforskolin fully inhibited I s but also reduced I p . Replacement of extracellular Cl− with aspartate demonstrated that the currents activated by nucleotides were Cl− selective. I p resulted five times more Cl− selective than I s with respect to aspartate. Taken together, our results indicate that ATP and UTP activate two types of Cl− currents through a Ca2+-dependent mechanism.
The Journal of Membrane Biology – Springer Journals
Published: Apr 1, 1997
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