NH 4+ secretion in the avian colon. An actively regulated barrier to ammonium permeation of the colon mucosa

Abstract

Experiments were designed to characterize an active, electrogenic transport of NH 4 + ions across the colonic epithelium of the domestic fowl ( Gallus gallus). Colonic segments were isolated and stripped of underlying muscle. The mucosal epithelia were mounted in Ussing chambers and voltage-clamped to measure the short-circuit currents ( I SC) associated with transport. Bilateral addition of NH 4 + caused a dose-dependent outward current (negative I SC), with a Km of 34 ± 8 mM and a maximal current response of 311 ± 47 µA cm − 2 (12 ± 2 µEq cm − 2 h − 1 ). A similar effect was seen with unilateral addition of NH 4 + to the serosal (s) side, but not with mucosal (m) addition. Pre-treatment with 10 − 4 M amiloride exposed a net outward (negative) I SC, and serosal NH 4 + addition further increased this outward current with a Km of 53 ± 24 mM. Decreasing the bath pH from 7.3 to 6.0 did not affect the I SC response to NH 4 +. Unidirectional NH 4 + flux measurements revealed a net secretory flux (8.8 ± 3.1 µmol cm − 2 h − 1 s–m, versus 2.6 ± 1.4 µmol cm − 2 h − 1 m–s). Furthermore, the secretory flux closely matched the resulting change in I SC with serosal NH 4 +, showing that the transepithelial flux of NH 4 + could account for the outward current response. Addition of 50 nM bafilomycin A to the mucosal solution completely eliminated serosal to mucosal NH 4 + transport, implicating an apical V-type H +-ATPase in this transport process. The I SC response to NH 4 + was partially inhibited by ouabain, a blocker of the Na +/K +-ATPase, but only minimally affected by bumetanide, an inhibitor of the serosal Na +–K +–2Cl − cotransporter. Active NH 4 + extrusion across the mucosal membrane, combined with low permeability to NH 3 in this tissue, allow for maintenance of steep ammonia gradients across the colonic epithelium and protection from ammonia toxicity. Furthermore, these studies indicate that the hen colon may be a useful new model system for the study of NH 4 + transport.