Modulation of sheep ruminal urea transport by ammonia and pH

Abstract

Ruminal fermentation products such as short-chain fatty acids (SCFA) and CO2 acutely stimulate urea transport across the ruminal epithelium in vivo, whereas ammonia has inhibitory effects. Uptake and signaling pathways remain obscure. The ruminal expression of SLC14a1 (UT-B) was studied using polymerase chain reaction (PCR). The functional short-term effects of ammonia on cytosolic pH (pHi) and ruminal urea transport across native epithelia were investigated using pH-sensitive microelectrodes and via flux measurements in Ussing chambers. Two variants (UT-B1 and UT-B2) could be fully sequenced from ovine ruminal cDNA. Functionally, transport was passive and modulated by luminal pH in the presence of SCFA and CO2, rising in response to luminal acidification to a peak value at pH 5.8 and dropping with further acidification, resulting in a bell-shaped curve. Presence of ammonia reduced the amplitude, but not the shape of the relationship between urea flux and pH, so that urea flux remained maximal at pH 5.8. Effects of ammonia were concentration dependent, with saturation at 5 mmol/l. Clamping the transepithelial potential altered the inhibitory potential of ammonia on urea flux. Ammonia depolarized the apical membrane and acidified pHi, suggesting that, at physiological pH (< 7), uptake of NH4 (+) into the cytosol may be a key signaling event regulating ruminal urea transport. We conclude that transport of urea across the ruminal epithelium involves proteins subject to rapid modulation by manipulations that alter pHi and the cytosolic concentration of NH4 (+). Implications for epithelial and ruminal homeostasis are discussed.

Short-chain fatty acids and acidic pH upregulate UT-B, GPR41, and GPR4 in rumen epithelial cells of goats

“…However, the present study has reported that SCFA and acidic pH upregulate mRNA expression and protein abundance of UT-B in rumen epithelial cells, whereas ammonia has the opposite effect. These findings are in good agreement with previous studies of urea transport (1,34), and, taken together, these data improve our understanding of how the recycling of urea and UT-B expression are regulated in ruminant animals. Second, the results of the present study suggest that diet modulates ruminal UT-B expression via rumen fermentation products (e.g., SCFA and acidic pH).…”

“…Ammonia has an inhibitory effect on the transport of urea in the rumen epithelium (1,34), and also on gene expression by bovine endometrial cells (19). In cultured rumen epithelial cells, ammonia was found to directly inhibit expression of UT-B mRNA and protein at pH 6.8, with or without SCFA administration, but not at pH 7.4.…”

“…Urea transport across the rumen epithelium is generally accepted to occur down a concentration gradient from blood to lumen by diffusion through transport proteins, such as urea transporter B (UT-B), or possibly aquaporins. In our laboratory's recent studies, rumen fermentation products, including short-chain fatty acids (SCFA) and ammonia, as well as pH, were found to rapidly modulate UT-B activity (1,34). It is possible that multiple mechanisms mediate regulation of urea transport, although these mechanisms are not fully characterized.…”

UT-B Urea Transporter Localization in the Bovine Gastrointestinal Tract

“…concentration specifically regulate urea transport (Lu et al 2014). Most recently, studies in goat rumen have confirmed that SCFAs and pH are the key factors in dietary regulation of UT-B expression (Lu et al 2015).…”

“…UT-B transporter proteins have previously been reported in the parotid salivary gland of both sheep and cows (Ludden et al 2009;Dix et al 2013). Furthermore, two distinct UT-B urea transporter isoforms, UT-B1 and UT-B2, are known to be expressed in the rumen of these species (Stewart et al 2005;Lu et al 2014). Importantly, ruminal UT-B expression increases with age, correlating to increases in blood urea nitrogen (Naeem et al 2012) and it has recently been shown to be upregulated by solid feed intake (Berends et al 2014).…”