Study 3: Metabolic Flexibility. In order to assess the influence of physiological changes in insulin level on metabolic switching of glucose and FFA fuels, animals were studied in either the Basal or ClampL states (defined previously). In this study a number of whole body and tissue specific glucose and FFA fluxes were evaluated: an estimate of R d, (R^d), rate of plasma FFA appearance (R fa), R g′, glycogen synthesis rate (R gly), tissue FFA utilization index (R f*), and rate of FFA incorporation into storage (R fs). The tracer experiment began following collection of a basal blood sample (Basal study) or attainment of clamp steady state (Clamp study). A bolus (~400 μL) containing D-(U-13C) glucose (13C-glucose, ~6 mg, Cambridge Isotope Laboratories, Inc., Andover, MA) and 2-deoxy-D-[U-14C] glucose (14C-2DG, ~5 × 107 dpm, Amersham, Solna, Sweden) was injected intravenously via a dedicated jugular catheter and arterial blood samples (75 μL) were collected at 2, 5, 10, 15, 20, and 25 min. Then at 30 min an albumin-palmitate-fatty acid tracer complex was infused via another dedicated jugular catheter at 230 μL/min
