Abstract Background Dietary n− 3 polyunsaturated fatty acids (PUFAs) have a role in preventing cardiovascular and hepatic diseases. However, their effects might differ significantly depending on individual dietary patterns. The aim of the present study was to evaluate the effects of dietary supplementation with ω-3 fatty acids (FA), administered in different schedules, on hepatic and aortic histological structure, lipid profile, and body weight (BW) in male Wistar rats under standard (SD), high-fat diet (HFD) and mixed feeding conditions. Methods PUFA treatment consisted of the administration of 50 mg/kg fish oil (FO) daily by oral gavage. HFD was obtained by adding a suspension of 4% cholesterol, thiouracil and cholic acid to the animals’ drinking water. The rats were maintained on the diets for 6 weeks, and different schedules of PUFA administration were used. At 14, 28, and 42 days, the morphology of liver and aortic samples and the levels of total cholesterol (TC), high-density lipoprotein cholesterol (HDL), low-density lipoprotein cholesterol (LDL), and triglycerides (TG) were assessed. Results The HFD groups exhibited significant hyperlipidemia and aortic inflammation, with progression to atherogenesis after 6 weeks. Administration of PUFAs slightly attenuated the aortic changes in these groups and reduced the liver’s tendency to steatosis. FO-induced metabolic improvement was more evident in SD than in HFD rats. For instance, after the first 2 weeks, SD animals that received PUFAs had significantly increased HDL levels vs. controls (62.375 ± 4.10 vs. 52.625 ± 8.38 mg/dL, P < 0.05), but HFD rats did not, and decreased TG levels were observed exclusively in the SD rats (57.6 ± 4.09 vs. 66 ± 4.69 mg/dL, P < 0.05). After 6 weeks of n− 3 PUFA administration, LDL was significantly lower in the SD rats than in controls (13.67 ± 4.13 vs. 30.83 ± 2.86 mg/dL, P < 0.001), but the decrease in the HFD rats, although significant (49.17 ± 5.85 mg/dL vs. 57.17 ± 4.96 g/dL, P < 0.05), was not as marked. In the mixed-diet groups, administration of 50 mg/kg/day FO for 14 days under SD conditions following 4 weeks of HFD slightly decreased TG (86.625 ± 11.67 vs. 73 ± 4.52 mg/dL, P < 0.05) and increased HDL (45.875 ± 5.28 vs. 56 ± 3.16 mg/dL). However, in these animals, n-3 PUFA administration had no effect on LDL or TC. Administration of half of the above dose failed to improve any biochemical parameters. FO protected against excessive weight gain mainly under SD conditions. Conclusions The results show that FO confers more protection against cardiovascular risk factors (increased LDL and TG, decreased HDL) and liver lipid accumulation when given to rats consuming regular diets than when given to rats consuming a high-fat diet. This argues that priority should be given to consumption of a healthy diet rather than to the use of supplements. The effectiveness of n-3 PUFAs might be reduced in the case of hyperlipidic intake or after consumption of a high-fat diet.