The changes in mitochondria bioenergetics play a critical role in neuronal function with age. Mitochondria from synaptosomes in the cerebral cortex of young (6 months), n=6, middle-aged (13 months), n=6 and old (26months), n=6, mice were isolated and transferred to mitochondria DNA-less LL/2-m21 cell line (rho-zero) to generate young, middle-aged and old cybrids. The cybrids show a significant reduction in mitochondria membrane potential (MMP) and ATP levels with age. Melatonin has been reported to be a potent therapeutic for age related cell dysfunction. Therefore we investigated how melatonin can influence mitochondrial function during ageing. Melatonin treatment in young mitochondria bearing cybrids, S-Y-24 lowered MMP from 130496.50±5118.75 to 84062±2633.43 (P<0.05), n=6 and increased MMP in middle-aged mitochondrial bearing cybrid S-M-29 from 10399.43 ±177.11 to 12862.79±186.51 (P<0.05), n=6. In the old cybrids, melatonin treatment raised MMP from 24151.46±1025.93 to 29824.68±563.96, P<0.05, n=4 in galactose medium. In the young cybrids, there was no significant difference in ATP levels between the melatonin treated cybrids – 4169.23±83.37 and controls – 4108.24±114.63, P>0.05, n=6. The young mitochondria bearing cybrids were able to produce ATP optimally in the galactose medium showing they have functional mitochondria. ATP level was significantly increased in the middle-aged mitochondria cybrid, S-M-29 from 3080.28±55.63 to 8542.85±130.38, P<0.05, n=6 and in old mitochondria bearing cybrids, S-O-48 from 2456.65±46.13 to 12415.50±265.16, P<0.05,n=6 with melatonin treatment. The unpaired t-test was used to analyze all the data presented in this study and the level of significance placed at P<0.05. These results establish a strong relationship between melatonin and mitochondria bioenergetics. Melatonin may be a potential therapeutic mitochondria target in neuronal dysfunction observed in ageing.