Urea-nitrogen salvage occurs when urea, generated in the liver and passed to the bloodstream, diffuses into the gut where it is hydrolyzed by ureolytic microbes into NH3 and CO2. Microbially-liberated urea-N can then be utilized in biosynthesis by gut microbes as well as the host. Although long-understood to be important in ruminants, UNS in non-ruminants is not well described; however, it has recently been implicated in protein conservation in monogastric mammals, including humans, especially under conditions of dietary nitrogen deficiency or increased host nitrogen demand. While these studies suggest an important role for ureolytic microbes and UNS in human health, little is known about the degree to which UNS contributes to host biosynthetic processes, or about beneficial ureolytic microbes in the gut. Therefore, the goal of this study is to exploit the extreme phenotype of the arctic ground squirrel to discover interrelationships between host physiology and the gut microbial community important for host nitrogen metabolism. The hypotheses for this investigation are that 1) reliance upon UNS to meet nitrogen needs varies with dietary nitrogen content and host nitrogen demand and across the annual cycle of hibernation and activity, and that 2) the gut microbial community contains active ureolytic microbes that contribute significantly to host nitrogen homeostasis via UNS. The proposed experimental approach 1) utilizes injections of 13C/15N labeled urea to assess ureolytic activity and UNS in vivo via measurements of 13CO2 in breath (evidence of ureolysis in the gut) and 15N in tissues (evidence of use of microbially-liberated urea-N in host biosynthetic processes), 2) links phylogenic and functional diversity of the gut microbiota using next generation sequencing techniques (metagenomics and metatranscriptomics) and 3) specifically examines the abundance and activity of ureolytic bacteria in the gut using qPCR and qRT-PCR of microbial urease genes, respectively. Through comparison of relative changes in ?15N of tissues of hibernating arctic ground squirrels injected with labeled or unlabeled urea, studies in Specific Aim 1 will demonstrate the degree to which microbially-liberated urea-N is incorporated into host tissues under a long term fast.
In Specific Aim 2, manipulation of dietary protein content will be used to determine the degree to which euthermic animals rely upon UNS for protein conservation under conditions of protein insufficiency. Similar diet manipulations in gestating and lactating squirrels will be conducted to evaluate the influence of dietary protein content and high nitrogen demand on the incorporation of microbially-liberated urea-N in mothers and pups in Specific Aim 3. Our experiments promise to yield an increased knowledge about interrelationships between the gut microbial community and host nitrogen homeostasis, ultimately contributing to our understanding of the relationship between the gut microbial community and human health, in particular under states of compromised nutrition (starvation, low dietary protein) and elevated N demand (gestation and lactation).

Public Health Relevance

Studies suggest the gut microbial community may play an important role in protein conservation under conditions that may lead to undernutrition, such as low nitrogen availability or high nitrogen demand. Our experiments investigate the gut microbial community of the arctic ground squirrel, an animal that naturally pushes the known limits of mammalian physiology, with the goal of understanding the relationship between the gut microbiota and host protein conservation. Our studies will contribute a greater understanding of the beneficial role of gut microbes to host nitrogen metabolism, and hold the potential to lead to development of microbially-based therapeutic interventions that improve nitrogen balance.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Academic Research Enhancement Awards (AREA) (R15)
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Molecular and Cellular Endocrinology Study Section (MCE)
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Maruvada, Padma
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University of Alaska Anchorage
Schools of Arts and Sciences
United States
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