The purpose of this study was to examine whether n-3 PUFA intake affects n-3 and n-6 FA levels in plasma and red blood cells as well as omega-3 index in old male and female spontaneously hypertensive (SHR) and healthy rats. Plasma linoleic acid and eicosapentaenoic acid increased due to n-3 PUFA intake in SHR and healthy rats. Comparing to healthy rats the levels of PUFA in red blood cells of SHR were lower in males and higher in females with exception of arachidonic acid, which was high in males and low in females. Feeding of rats with n-3 PUFA resulted in increase of red blood cells levels of eicosapentaenoic acid and/or docosahexaenoic acid in a sex- and strain-dependent manner. Moreover, n-3 PUFA intake decreased arachidonic acid in healthy female rats but increased it in SHR and did not affect it in males. Omega-3 index was lower in SHR comparing to healthy rats and it increased due to the consumption of n-3 PUFA. Results point out sex- and strain-related differences in red blood cells levels of n-3 and n-6 PUFA in basal conditions as well as in response to n-3 PUFA intake.