The adipokine leptin is essential for maintaining energy homeostasis, and signals via receptors (LepR) in the brain to reduce food intake and increase energy expenditure. While the feeding effects of leptin are well known, the pathways that mediate leptin’s effects on energy expenditure are less understood (1,2). Here we show that LepR expression on neurones containing prolactin-releasing peptide (PrRP) is essential for normal energy expenditure in the mouse, and specifically for the effects of leptin on thermogenesis.Immunohistochemistry revealed that PrRP colocalises strongly with LepR-eGFP in the dorsomedial nucleus of the hypothalamus (DMH) in fluorescent reporter animals. Furthermore, following intraperitoneal (ip) leptin injection, DMH PrRP neurones exhibited a robust increase in STAT3 phosphorylation, indicating a direct neuronal response to leptin. Also, using whole-cell patch clamping of PrRP-eGFP brain slices, DMH PrRP neurones were shown to be inhibited by 100nM leptin (11/18 cells showed a decrease in membrane potential and firing rate).To study the physiological effects of leptin, a PrRP-cre knock-in mouse was crossed with a floxed LepR mouse (3) (PrRP-cre::LepRflox/flox), to delete LepR solely in PrRP neurones. These crossed animals exhibited obesity, with no difference in baseline feeding, but had decreased oxygen consumption (3069±31ml/kg/hr vs 2898±56ml/kg/hr, LepRflox/flox vs PrRP-cre::LepRflox/flox, n=5/4, p<0.05; values are means±SEM, compared by ANOVA). The mice also had lower core-body temperature as measured by a temperature transmitter implanted under 2% isoflurane anaesthesia (36.83±0.08°C vs 36.57±0.10°C, LepRflox/flox vs PrRP-cre::LepRflox/flox, n=6, p<0.05). Furthermore, ip injection of leptin caused a sustained increase in body temperature of ~0.5°C for the 3 hours post-injection in LepRflox/flox mice (n=6, p<0.001), but had no effect in PrRP-cre::LepRflox/flox animals.Finally, under 2% isoflurane, an adeno-associated virus (AAV) containing a cre-dependent anterograde tracer was injected into the DMH of PrRP-cre mice. Infected PrRP neurones showed local projections within the DMH and substantial projections to the paraventricular nucleus of the hypothalamus (PVH). Additionally, a retrograde tracer injected into the PVH colocalised with PrRP-eGFP neurones in the DMH. In summary, leptin acts directly through Lepr on DMH PrRP neurones to control energy expenditure and thermogenesis. Furthermore, DMH PrRP neurones project predominantly to the PVH, a major site of integration and processing of autonomic functions.