Under traditional management on the Qinghai–Tibetan Plateau, yaks (Bos grunniens) graze only on natural pasture without supplements and are forced to cope with sparse forage of low N content, especially in winter. In contrast, indigenous Tibetan yellow cattle (Bos taurus) require supplements during the cold season. We hypothesized that, in response to harsh conditions, yaks cope with low N intakes better than cattle. To test this hypothesis, a study of whole-body N retention and urea kinetics was conducted in 2 concurrent 4 × 4 Latin squares, with 1 square using yaks and 1 square using cattle. Four isocaloric forage–concentrate diets differing in N concentrations (10.3, 19.5, 28.5, and 37.6 g N/kg DM) were formulated, and by design, DMI were similar between species and across diets. Urea kinetics were determined with continuous intravenous infusion of ¹⁵N¹⁵N urea for 104 h, and total urine and feces were concomitantly collected. Urea production, urea recycling to the gut, and ruminal microbial protein synthesis all linearly increased (P < 0.001) with increasing dietary N in both yaks and cattle. Urinary N excretion was less (P = 0.04) and N retention was greater (P = 0.01) in yaks than in cattle. Urea production was greater in yaks than in cattle at the 3 lowest N diets but greater in cattle than in yaks at the highest N diet (species × diet, P < 0.02). Urea N recycled to the gut (P < 0.001), recycled urea N captured by ruminal bacteria (P < 0.001), and ruminal microbial protein production (P = 0.05) were greater in yaks than in cattle. No more than 12% of urea recycling was through saliva, with no difference between species (P = 0.61). Glomerular filtration rate was lower (P = 0.05) in yaks than in cattle. The higher urea recycling and greater capture of recycled urea by ruminal microbes in yaks than in cattle suggest that yaks use mechanisms to utilize dietary N more efficiently than cattle, which may partially explain the better survival of yaks than cattle when fed low-N diets.

The symbiotic relationship between the host and the rumen microbiome plays a crucial role in ruminant physiology. One of the most important processes enabling this relationship is urea nitrogen salvaging (UNS). This process is important for both maintaining ruminant nitrogen balance and supporting production of their major energy supply, bacterially-derived short chain fatty acids (SCFA). The key step in UNS is the trans-epithelial movement of urea across the ruminal wall and this is a highly regulated process. At the molecular level, the key transport route is via facilitative urea transporter (UT)-B2 localized to ruminal papillae epithelial layers. Additional urea transport through aquaporins (AQP), such as AQP3, are now also viewed as important. Long-term regulation of these ruminal urea transport proteins appears to mainly involve dietary fermentable carbohydrates; whereas, transepithelial urea transport is finely regulated by local conditions, such as CO₂ levels, pH and SCFA concentration. Although the key principles of ruminal urea transport physiology are now understood, there remains much that is unknown regarding the regulatory pathways. One reason for this is the limited number of techniques currently used in many studies in the field. Therefore, future research in this area that combines a greater range of techniques could facilitate improvements to livestock efficiency, and potentially, reductions in the levels of waste nitrogen entering the environment.

Urea nitrogen salvaging is a crucial mechanism that ruminants have evolved to conserve nitrogen. Facilitative urea transporter-B proteins are known to be involved in urea transport across the rumen epithelium and thus efficiently facilitate the urea nitrogen salvaging process. Recently, functional studies have suggested that aquaglyceroporin transporters might also play a significant role in ruminal urea transport and aquaporin-3 (AQP3) protein has previously been detected in rumen tissue. In this current study, we investigated the specific localization of AQP3 transporters in the bovine rumen. First, end-point reverse-transcription PCR experiments confirmed strong AQP3 expression in both bovine rumen and kidney. Immunoblotting analysis using 2 separate anti-AQP3 antibodies detected AQP3 protein signals at 25, 32, and 42-45 kDa. Further immunolocalization studies showed AQP3 protein located in all the layers of rumen epithelium, especially in the stratum basale, and in the basolateral membranes of kidney collecting duct cells. These data confirm that AQP3 transporters are highly abundant within the bovine rumen and appear to be located throughout the ruminal epithelial layers. The physiological significance of the multiple AQP3 proteins detected and their location is not yet clear, hence further investigation is required to determine their exact contribution to ruminal urea transport.

Urea nitrogen secreted from blood to rumen is a crucial factor shaping the symbiotic relationship between host ruminants and their microbial populations. Passage of urea across rumen epithelia is facilitated by urea transporter B (UT-B), but the long-term regulation of these proteins remains unclear. As ruminal function develops over a period of months, the developing rumen is an excellent model with which to investigate this regulation. Using rumen epithelium samples of calves from birth to 96 d of age, this study performed immunolocalization studies to localize and semi-quantify UT-B protein development. As expected, preliminary experiments confirmed that ruminal monocarboxylate transporter 1 (MCT1) short chain fatty acid transporter protein abundance increased with age (P < 0.01, n = 4). Further investigation revealed that ruminal UT-B was present in the first few weeks of life and initially detected in the basolateral membrane of stratum basale cells. Over the next 2 months, UT-B staining spread to other epithelial layers and semi-quantification indicated that UT-B abundance significantly increased with age (P < 0.01, n = 4 or 6). These changes were in line with the development of rumen function after the advent of solid feed intake and weaning, exhibiting a similar pattern to both MCT1 transporters and papillae growth. This study therefore confirmed age-dependent changes of in situ ruminal UT-B protein, adding to our understanding of the long-term regulation of ruminal urea transporters.

Rumen studies have focused almost exclusively on livestock species under strictly regimented diets. This means that the ruminal condition of free-living and free-feeding wildlife remains practically unstudied. Urea nitrogen salvaging, a process by which urea is passed into the rumen, to both provide a valuable source of nitrogen for bacterial growth and to buffer the potentially harmful acidic effects of bacterial short chain fatty acids, has remained unexplored in wild ruminants, such as deer. UT-B2 transporters are the key proteins reported to facilitate the transepithelial ruminal urea transport. In this study, we investigate the expression, abundance and localisation of urea transporters in the rumen of a semi-wild fallow deer (Dama dama) population. Physical measurements confirmed that males had larger rumen than females, while adults had longer papillae than juveniles. Initial RT-PCR experiments confirmed the expression of UT-B2, while immunolocalisation studies revealed that strong UT-B staining was present in the stratum basale of deer rumen. Western blotting analysis demonstrated that a 50 kDa UT-B2 protein was significantly more abundant in adult females compared to adult males. This study confirms the presence of UT-B2 urea transporters in deer rumen and suggests that sex-related differences occur, bringing new insight into our understanding of rumen physiology.