Characterizing the Real‐Time Endogenous Methane Release in a Rat for 26‐days: Effect of Diets, Fasting, and Ethanol Consumption Across Different Time Scales

Abstract

IntroductionMethane is a byproduct of methanogenic archaea that oxidize H2 gas produced by other microflora in the gut. This methane gas is readily absorbed into the into systemic circulation and excreted in the breath down its concentration gradient (similar to CO2) or may be acutely expelled as flatus. While ruminant animals can generate over 100 liters of methane per day, technological limitations have precluded detailed measurements of methane production and release in humans and other non‐ruminants.MethodsWe paired a Promethion metabolic measurement system with a high‐precision (<1ppb) methane analyzer capable of real‐time methane measurements in an individual mouse or rat. We then subjected a rat to multiple diet switches, 48h of fasting, and ethanol treatments and continuously measured food/water intake, metabolic rate, and methane release over 26 days.ResultsVCO2 responses were unaffected by diet changes and only decreased slightly during the fasting treatment. Methane release was diminished by some diet treatments and could be restored by others. Ethanol had no observable effects. We discovered that methane production generally followed a circadian cycle whereby total rates were ~50% higher and large flatus events were more frequent during the scotophase. Methane release during flatus varied by >1,000‐fold ranging from as little as 0.1 up to 60μl/min. Integrating the continuous data stream (at 1 Hz) we confirmed that the vast majority of methane release occurred in the breath and not in the flatus as is often repeated in the literature. Finally, we provide evidence that the constitutive rate of methane release in the breath is positively correlated with rates of CO2 release, the latter of which is indicative of overall activity levels of the rat.ConclusionGiven the growing number of molecular‐based studies investigating how gut microbiota can alter physiological function at higher levels of biological organization the approach of using real‐time methane measurements promises to provide a powerful new tool to establish causal linkages between microbiome dynamics and whole‐animal responses.