Abstract Background and Aims Mitochondrial homeostasis is controlled by biogenesis, dynamics, and mitophagy. Mitochondrial dysfunction plays a central role in cardiovascular and renal disease and omega-3 fatty acids (FA) are beneficial for cardiovascular disease. We aimed to investigate whether omega-3 FA regulates the expression of mediators of mitochondrial biogenesis, dynamics, and mitophagy in kidney and heart of adenine-induced uremic rats. Method Sprague-Dawley rats were divided into three groups: normal control, adenine control, and omega-3 FA. Uremia was induced by feeding diets containing 0.75% adenine and 2.5% protein for the first 3 weeks. During next 4 weeks, they were received 2.5% protein with or without omega-3 FA (300 mg/kg/day). The renal and cardiac expression of PGC-1α, SIRT1/3, Nrf2, DRP-1, OPA1, Mfn1/2, PINK1, BNIP3 and NIX were examined by western blot analysis. The qPCR was used to determine mitochondrial DNA (mtDNA). Results Compared to normal, serum creatinine and heart weight/body weight in adenine control was increased and improved in omega-3 FA group. Compared with normal, PGC-1α, SIRT1/3 and Nrf2 were down-regulated in kidney and heart of adenine control. PGC-1α expression of kidney and heart was recovered in omega-3 FA group. DRP-1 of kidney was up-regulated but DRP-1 of heart was down-regulated in adenine control. DRP-1 of heart was recovered in omega-3 FA group. PINK1, BNIP3 and NIX were down-regulated in heart of adenine control and recovered in omega-3 FA group. PINK1 was down-regulated but BNIP3 and NIX were up-regulated in kidney of adenine control and those were mitigated in omega-3 FA group. MtDNA was decreased in kidney and heart of adenine control group but mtDNA of heart was recovered in omega-3 FA group. Conclusion DRP-1 related with mitochondrial fission may oppositely work in uremic kidney and heart. Omega-3 FA is beneficial for mitochondrial homeostasis by activating mitochondrial biogenesis and PINK1 dependent mitophagy in kidney and heart of uremic rats.