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The cellular uptake of ammonia affects the intracellular pH (pHi) of polar and non-polar cells. A predominant uptake of NH3 and its intracellular protonation tend to alkalinise the cytoplasm, whereas a predominant uptake of NH4(+) acidifies the cytoplasm by reversing this reaction. Hence, the well-known absorption of ammonia across the rumen epithelium probably causes a change in the pHi. The magnitude and direction of this change in pHi (acid or alkaline) depends on the relative transport rates of NH3 and NH4(+). Consequently, the intracellular availability of protons will influence the activity of the Na(+)-H(+) exchanger, which could affect transepithelial Na(+) transport. The aim of the present study has been to test this possible interaction between ruminal ammonia concentrations and Na(+) transport. The term ammonia is used to designate the sum of the protonated (NH4(+)) and unprotonated (NH3) forms. Isolated ruminal epithelium of sheep was investigated by using the Ussing-chamber technique in vitro. The present results indicate that ammonia inhibits Na(+) transport across the rumen epithelium of hay-fed sheep, probably by binding intracellular protons and thus inhibiting Na(+)-H(+) exchange. By contrast, ammonia stimulates Na(+) transport in concentrate-fed and urea-fed sheep, which develop an adaptation mechanism in the form of an increased metabolism of ammonia in the rumen mucosa and/or an increased permeability of rumen epithelium to the charged ammonium ion (NH4(+)). Intracellular dissociation of NH4(+) increases the availability of protons, which stimulate Na(+)-H(+) exchange. This positive effect of ruminal ammonia on Na(+) absorption may significantly contribute to the regulation of osmotic pressure of the ruminal fluid, because intraruminal ammonia concentrations up to 40 mmol/l have been reported.