BACKGROUND: High protein (HP) diets have been proposed to reduce body weight in humans. The diets are known to alter energy metabolism, which can affect the quality of [18F]FDG PET heart images. In this preclinical study, we therefore explore the impact of a prolonged HP diet on myocardial [18F]FDG uptake. METHODS: C57BL/6J (Black six (Bl6)) and apolipoprotein E-deficient (apoE-/-) mice were fed chow, a HP diet, or a low protein (LP) diet for 12 weeks. At baseline and after treatment, the animals were injected with 33.0 MBq of [18F]FDG and a 30 min PET/CT scan was made. Myocardial volume and [18F]FDG uptake were quantified using PET and the % of body fat was calculated from CT. RESULTS: Myocardial [18F]FDG uptake was similar for all diets at the follow-up scan but an increase between baseline and follow-up scans was noticed in the LP groups. Myocardial volume was significantly smaller in the C57BL HP group compared to the other Bl6 groups. Body weight increased less in the two HP groups compared to the chow and LP groups. Body fat percentage was significantly higher in the LP groups. This effect was stronger in C57BL mice (28.7%) compared to apoE-/- mice (15.1%). CONCLUSIONS: Myocardial uptake of [18F]FDG in mice is not affected by increased protein intake but [18F]FDG uptake increases when the amount of protein is lowered. A lower body weight and percentage of body fat were noticed when applying a HP diet.