CA2+-INDUCED AND SWELLING-INDUCED ACTIVATION OF ION CONDUCTANCES IN BRONCHIAL EPITHELIAL-CELLS

The present study was performed to examine Ca2+-dependent and cell-swelling-induced ion conductances in a polarized bronchial epithelial cell line (16HBE14(o)-). Whole-cell currents were measured in fast and slow whole-cell patch-clamp experiments in cells grown either on filters or on coated plastic dishes. In addition the transepithelial voltage (V-te) and resistance (R(te)) were measured in confluent monolayers. Resting cells had a membrane voltage (V-m) of -36 +/- 1.1 mV (n = 137) which was mainly caused by K+ and Cl- conductances and to a lesser extent by a Na+ conductance. V-te was apical-side-negative after stimulation. Equivalent short-circuit current (I-sc = V-te/R(te)) was increased by the secretagogues histamine (0.1 mmol/l), bradykinin (0.1-10 mu mol/l) and ATP (0.1-100 mu mol/l). The histamine-induced I-sc was blocked by either basolateral diphenhydramine (0.1 mmol/l, n = 4) or apical cimetidine (0.1 mmol/l, n = 4). In fast and slow whole-cell recordings ATP and bradykinin primarily activated a transient K+ conductance and hyperpolarized V-m. This effect was mimicked by the Ca2+ ionophore ionomycin (1 mu mmol/l, n = 11). Inhibition of the bradykinin-induced I-sc by the blocker HOE140 (1 mu mol/l, n = 3) suggested the presence of a BK2 receptor. The potency sequence of different nucleotide agonists on the purinergic receptor was UTP approximate to ATP > ITP > GTP approximate to CTP approximate to [beta,gamma-methylene] ATP approximate to 2-methylthio-ATP = 0 and was obtained in I-sc measurements and patch-clamp recordings. This suggests the presence of a P-2u receptor. Hypotonic cell swelling activated both Cl- and K+ conductances. The Cl- conductance was only slightly inhibited by 4,4'-di- isothiocyanatostilbene-2,2'-disulphonic acid (0.5 mmol/l 1, n = 3). These data indicate that 16HBE140- bronchial epithelial cells, which are known to express high levels of cystic fibrosis transmembrane conductance regulator protein, form a secretory epithelium. While hypotonic cell swelling activates both K+ and Cl- channels, the Ca2+-induced Cl- secretion is due mainly to activation of basolateral K+ channels.