title: CFTR mediated chloride secretion in the avian renal proximal tubule.
abstract: In primary cell cultures of the avian (Gallus gallus) renal proximal tubule parathyroid hormone and cAMP activation generate a Cl(-)-dependent short circuit current (I(SC)) response, consistent with net transepithelial Cl(-) secretion. In this study we investigated the expression and physiological function of the Na-K-2Cl (NKCC) transporter and CFTR chloride channel, both associated with Cl(-) secretion in a variety of tissues, in these proximal tubule cells. Using both RT-PCR and immunoblotting approaches, we showed that NKCC and CFTR are expressed, both in proximal tubule primary cultures and in a proximal tubule fraction of non-cultured (native tissue) fragments. We also used electrophysiological methods to assess the functional contribution of NKCC and CFTR to forskolin-activated I(SC) responses in filter grown cultured monolayers. Bumetanide (10 μM), a specific blocker of NKCC, inhibited forskolin activated I(SC) by about 40%, suggesting that basolateral uptake of Cl(-) is partially mediated by NKCC transport. In monolayers permeabilized on the basolateral side with nystatin, forskolin activated an apical Cl(-) conductance, manifested as bidirectional diffusion currents in the presence of oppositely directed Cl(-) gradients. Under these conditions the apical conductance appeared to show some bias towards apical-to-basolateral Cl(-) current. Two selective CFTR blockers, CFTR Inhibitor 172 and GlyH-101 (both at 20 μM) inhibited the forskolin activated diffusion currents by 38-68%, with GlyH-101 having a greater effect. These data support the conclusion that avian renal proximal tubules utilize an apical CFTR Cl(-) channel to mediate cAMP-activated Cl(-) secretion.
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, DE 19716, USA. Laverty@udel.edu
authors.forename: Gary
authors.initials: G
authors.lastname: Laverty
authors.forename: Ashley
authors.initials: A
authors.lastname: Anttila
authors.forename: Jenava
authors.initials: J
authors.lastname: Carty
authors.forename: Varudhini
authors.initials: V
authors.lastname: Reddy
authors.forename: Jamie
authors.initials: J
authors.lastname: Yum
authors.forename: Sighvatur S
authors.initials: SS
authors.lastname: Arnason
mesh_terms: D000818:Animals; D002645:Chickens / Q000235:genetics / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D061251:Primary Cell Culture; D028021:Sodium-Potassium-Chloride Symporters / Q000235:genetics / Q000378:metabolism*
chemical_list: D002712:Chlorides; D028021:Sodium-Potassium-Chloride Symporters; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP