Time course of hepatic gluconeogenesis during hindlimb suspension unloading

The goal of this work was to determine the time-dependent changes in fractional hepatic gluconeogenesis(GNG)duringconditionsofhindlimbsuspensionunloading(HSU),a‘groundbased’ method for inducing muscular atrophy to simulate space flight. We hypothesized that GNG would increase in HSU conditions as a result of metabolic shifts in the liver and skeletal muscle. A significant and progressive atrophy was observed in the soleus (30, 47 and 55%) and gastrocnemius muscles (0, 15 and 17%) after 3, 7 and 14 days of HSU, respectively. Fractional hepatic GNG was determined following the incorporation of deuterium from deuterated water ( 2 H2O) into C–H bonds of newly synthesized glucose after an 8 h fast. Enrichment of plasma glucose with 2 H was measured using the classic method of Landau et al. (the ‘hexamethylenetetramine (HMT) method’), based on specific 2 H labelling of glucose carbons, and the novel method of Chacko et al. (‘average method’), based on the assumption of equal 2 H enrichment on all glucose carbons (except C2). After 3 days of HSU, fractional GNG was significantlyelevatedintheHSUgroup,asdeterminedbyeithermethod(∼13%,P <0.05).After 7 and 14 days of HSU, gluconeogenesis was not significantly different. Both analytical methods yieldedsimilartime-dependenttrendsingluconeogenicrates,butGNGvaluesdeterminedusing the average method were consistently lower (∼30%) than those found by the HMT method. To compare and validate the average method against the HMT method further, we starved animals for 13 h to allow for hepatic GNG to contribute 100% to endogenous glucose production. The HMT method yielded 100% GNG, while the average method yielded GNG of ∼70%. As both methodsusedthesamevaluesofprecursorenrichment,wepostulatedthattheunderestimation of gluconeogenic rate was as a result of differences in the measurements of product enrichment ( 2 H labelling of plasma glucose). This could be explained by the following factors: (i) loss of deuterium via exchange between acetate and glucose; (ii) interference caused by fragment m/z 169, representing multiple isobaric species; and (iii)interference from other sugars at m/z 169. In conclusion, HSU caused a time-dependent increase in hepatic gluconeogenesis, irrespective of the analytical methods used.