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.