Abstract A combination of microdialysis (MD) and 2-[ 18F ]fluoro-2-deoxy-D-glucose (FDG) was used to assess FDG uptake, phosphorylation and the glucose metabolic index (Rg′) in certain tissues of fed and fasting anesthetized Sprague-Dawley rats which received an i.v. bolus injection of insulin or saline during the course of the study. The relative recovery for FDG for the MD probes was also measured as a function of flow rate and temperature. The elimination half-life (T 1/2 FDG) of FDG from the plasma and the extracellular fluid of muscle and liver was studied with MD. The phosphorylation of FDG in muscle, liver, subcutaneous fat and mesenteric fat from homogenates of these tissues was analyzed by a radioHPLC-method and the Rg′ was calculated. The results show that the nutritional status does not affect the T 1/2 FDG, the total uptake of FDG 6-phosphate or the Rg′ values in the studied tissues at ambient glucose. Insulin stimulation decreased T 1/2 FDG, and increased the total FDG 6-P accumulation and Rg′ in the muscle of fed and fasted rats. In adipose tissues the insulin stimulation enhanced the phosphorylation but in muscle the proportion of FDG 6-P remained unchanged. Rg′ in adipose tissue was higher after insulin administration in fed rats than without insulin but with fasted rats there were no differences in Rg′ values with or without insulin, although the proportion of FDG 6-P did increase. The Rg′ values for the livers were unaffected by any of the manipulations, but fasted rats accumulated proportionately more FDG 6-P after insulin administration than did fed rats. These results indicate that the combination of MD and FDG is a valuable and reliable tool when studying glucose metabolism in physiological and pathological models in vivo.