Leptin has potent lipid-lowering effects in peripheral tissues and plasma that are proposed to be important for the prevention of cellular lipotoxicity and insulin resistance. The current study addressed in vivo the effects of acute leptin delivery on liver triglyceride (TG) metabolism, the consequence of hepatic leptin action on whole-body TG homeostasis, and the mechanisms of leptin action. A 120-min iv leptin infusion (plasma leptin, approximately 14 ng/ml) decreased liver TG levels (53 +/- 3%; P = 0.001), but not skeletal muscle TG levels, and increased liver phosphatidylinositol 3-kinase activity (341 +/- 95%; P = 0.01) in lean rats. Leptin had no effect on liver TG levels or phosphatidylinositol 3-kinase activity in diet-induced obese rats. In lean animals, leptin decreased the plasma TG concentration (20 +/- 7%; P = 0.017), the rate of TG accumulation in plasma after tyloxapol administration (26 +/- 6%; P = 0.003), and TG secretion from isolated liver (51 +/- 8%; P = 0.004). To determine possible metabolic fates of depleted hepatic TG, we assessed leptin effects on liver oxidative metabolism. Leptin increased hepatic acetyl-coenzyme A carboxylase phosphorylation (85 +/- 13%; P = 0.006), fatty acid oxidation (49 +/- 7%; P = 0.001) and ketogenesis (69 +/- 15%; P = 0.004). Finally, intracerebroventricular delivery of leptin for 120 min had no effect on liver TG levels, but did increase signal transducer and activator of transcription 3 phosphorylation (162 +/- 40%; P = 0.02). These data present in vivo evidence for a role for leptin in the acute regulation of hepatic TG metabolism, and whole body TG homeostasis. A likely contributing mechanism for these effects is leptin-induced partitioning of TG into oxidative pathways.