Evidence for the functional involvement of members of the TRP channel family in the uptake of Na+ and NH4+ by the ruminal epithelium

Abstract

Large quantities of protein are degraded in the fermentative parts of the gut to ammonia, which is absorbed, detoxified to urea, and excreted, leading to formation of nitrogenous compounds such as N2O that are associated with global warming. In ruminants, channel-mediated uptake of NH4+ from the rumen predominates. The molecular identity of these channels remains to be clarified. Ruminal cells and epithelia from cows and sheep were investigated using patch clamp, Ussing chamber, microelectrode techniques, and qPCR. In patch clamp experiments, bovine ruminal epithelial cells expressed a conductance for NH4+ that could be blocked in a voltage-dependent manner by divalent cations. In the native epithelium, NH4+ depolarized the apical potential, acidified the cytosol and induced a rise in short-circuit current (Isc) that persisted after the removal of Na+, was blocked by verapamil, enhanced by the removal of divalent cations, and was sensitive to certain transient receptor potential (TRP) channel modulators. Menthol or thymol stimulated the Isc in Na+ or NH4+ containing solutions in a dose-dependent manner and modulated transepithelial Ca2+ fluxes. On the level of messenger RNA (mRNA), ovine and bovine ruminal epithelium expressed TRPA1, TRPV3, TRPV4, TRPM6, and TRPM7, with any expression of TRPV6 marginal. No bands were detected for TRPV1, TRPV5, or TRPM8. Functional and molecular biological data suggest that the transport of NH4+, Na+, and Ca2+ across the rumen involves TRP channels, with TRPV3 and TRPA1 emerging as prime candidate genes. TRP channels may also contribute to the transport of NH4+ across other epithelia.