The endocannabinoid system (ECS) represents one of the major determinants of metabolic disorders. We investigated potential changes in the endogenous levels of anandamide (AEA), 2-arachidonoylglycerol (2-AG), N-oleoylethanolamine (OEA) and N-palmitoylethanolamine (PEA) in some peripheral organs and tissues of obese Zucker(fa/fa) and lean Zucker(fa/+) rats by qPCR, liquid chromatography mass spectrometry, western blot and enzymatic activity assays. At 10-12 weeks of age AEA levels were significantly lower in BAT, small intestine and heart and higher in soleus of Zucker(fa/fa) rats. In this tissue, also the expression of CB1 receptors was higher. By contrast in Zucker(fa/fa) rats, 2-AG levels were changed (and lower) solely in the small and large intestine. Finally, in Zucker(fa/fa), PEA levels were unchanged, whereas OEA was slightly lower in BAT, and higher in the large intestine. Interestingly, these differences were accompanied by differential alterations of the genes regulating ECS tone. In conclusion, the levels of endocannabinoids are altered during obesity in a way partly correlating with changes of the genes related to their metabolism and activity.