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The prevalence of obesity is still increasing, despite obesity treatment strategies that aim at reducing energy intake. In addition to this, exercise programmes designed to increase energy expenditure have only a low efficiency and have generated mixed results. Therefore, strategies based on increasing energy expenditure via nonexercise means are currently under investigation. One novel strategy is the modulation of adaptive thermogenesis.Among others, adaptive thermogenesis can be modulated by changing dietary composition, treatment with hormone mimetics as well as by cold exposure. In humans, a large part of the adaptive thermogenic response is, in addition to a putative role of brown adipose tissue, determined by the skeletal muscle mass via the process of mitochondrial uncoupling. Here, we describe the molecular processes involved in mitochondrial uncoupling, state-of-the-art techniques to measure mitochondrial uncoupling in vitro and in vivo, as well as the current strategies to mitochondrial uncoupling.Data generated in rodents and humans implicate that increasing adaptive thermogenesis by increasing skeletal muscle mitochondrial uncoupling indeed elevates total energy expenditure and thus may provide a promising target for the treatment of obesity.