A physiologically based model proposed by our group has been developed to assess glucose transport and phosphorylation in skeletal muscle. In this study, we investigated whether our model has the ability to detect a glucose-induced increase in glucose transport in skeletal muscle. In particular, we used small-animal positron emission tomography (PET) data obtained from [18F]6-fluoro-6-deoxy-D-glucose ([18F]6FDG). A 2 h PET scan was acquired following a bolus injection of [18F]6FDG in rats currently under euglycemic or hyperglycemic conditions, while somatostatin was infused during both conditions in order to prevent a rise in the endogenous plasma insulin concentration. We were thus able to assess the effect of hyperglycemia per se. For a comparison of radiopharmaceuticals, additional rats were studied under the same conditions, using [18F]2-fluoro-2-deoxy-D-glucose ([18F]2FDG). When [18F]6FDG was used, the time-activity curves (TACs) for skeletal muscle had distinctly different shapes during euglycemic and hyperglycemic conditions. This was not the case with [18F]2FDG. For both [18F]6FDG and [18F]2FDG, the model detects increases in both interstitial and intracellular glucose concentrations, increases in the maximal velocity of glucose transport and increases in the rate of glucose transport, all in response to hyperglycemia. In contrast, there was no increase in the maximum velocity of glucose phosphorylation or in the glucose phosphorylation rate. Our model-based analyses of the PET data, obtained with either [18F]6FDG or [18F]2FDG, detect physiological changes consistent with established behavior. Moreover, based on differences in the TAC shapes, [18F]6FDG appears to be superior to [18F]2FDG for evaluating the effect of hyperglycemia on glucose metabolism in skeletal muscle.