Due to environmental and medicinal exposures to lithium, as well as its uptake and accumulation in various plant species as human food source, concerns about lithium toxicity and negative impact on different organs especially heart have been raised. The toxicity mechanism of lithium is still unclear, but it has been suggested that some its harmful effects may be related to mitochondrial dysfunction and oxidative stress. Previous studies have demonstrated that plant-derived natural compounds can ameliorate mitochondrial dysfunction induced by various chemicals. In the current study, we examined the effects of vanillic acid as a plant-derived natural compound on lithium-stimulated mitochondrial dysfunction in rat heart-isolated mitochondria and its potential mechanisms of attenuating damages to improve function of mitochondria during 60 min. Mitochondrial injury in rat heart-isolated mitochondria was induced by lithium (125 µM, according to previous studies) and portative effect of vanillic acid (10, 50, and 100 µM) was assessed using mitochondrial toxicity parameters such as the functional state of mitochondria, reactive oxygen species (ROS) formation, mitochondrial membrane potential (MMP) collapse, mitochondrial swelling, and malondialdehyde (MDA) levels. Our results confirmed that vanillic acid (10, 50, and 100 µM) significantly mitigated mitochondrial dysfunction triggered by lithium, evidenced by the decline in formation of ROS and MDA, improvement the mitochondrial membrane potential, inhibition of mitochondrial swelling, and the increase of the functional state of mitochondria. Our findings suggested that vanillic acid mitigated mitochondrial dysfunction via maintenance of mitochondrial function, inhibition of oxidative stress and mitochondrial swelling, it could be developed as mitochondrial protective agents in the prevention of cardiotoxicity induced by lithium.