Dysregulation of fatty acid oxidation plays a pivotal role in the pathophysiology of obesity, insulin resistance, and in impaired thermoregulation. We generated a gene-trap knockout mouse (Hadh-/- mouse; gene name: Hadh, protein name: SCHAD) to disrupt one of the last steps in mitochondrial beta-oxidation. Deletion of Hadh results in a reduced body weight due to a loss of acylcarnitines via the urine and a compensatory elevated beta-oxidation (Schulz et al, Endrocrinology 2011). To study the link of beta-oxidation and thermogenesis in vivo we carried out cold exposure analysis with and without exogenous energy supply. To study cold tolerance of Hadh+/+- and Hadh-/- mice, animals were kept single housed in a climate cabinet for 3 hours at 4°C to measure their rectal temperature every 30 minutes. Moreover, we isolated brown adipocytes of Hadh+/+- and Hadh-/- mice, immortalized the adipocytes with a SV40 virus and performed in vitro studies with differentiated adipocytes. Hadh-/- mice exhibited significantly higher body temperature than their wild-type littermates under control conditions. In contrast, acute cold exposure led to a significant reduction of body temperature of Hadh-/- mice due to their inability to use fat as an energy source under these conditions. After cold exposure levels of phosphorylated hormone-senstive lipase (pHSL) were also lower in brown adipose tissue (BAT) of Hadh-/- mice than in BAT of Hadh+/+ mice. Moreover, preliminary data of immortalised brown adipocytes of Hadh-/- mice displayed reduced isoproterenol-induced lipolysis and exhibited lower amounts of pHSL. Hadh is required for maintaining thermogenesis by influencing lipolysis in brown adipocytes.