The adipocyte hormone leptin activates signal transducer and activator of transcription 3 (STAT3) in the hypothalamus, mediating increased satiety and increased energy expenditure. To date, leptin-mediated activation of the STAT pathway in vivo has not been established in tissues other than hypothalamus. We now describe leptin receptor expression and in vivo signaling in discrete regions of the mouse gastrointestinal tract. Expression of the functional isoform leptin receptor (OB-Rb) is restricted to the jejunum and is readily detected by RT-PCR in isolated enterocytes from this site. Intravenous injection of leptin rapidly induced nuclear STAT5 DNA binding activity in jejunum of +/+ and obese (ob/ob) mice but had no effect in the diabetic (db/db) mouse that lacks the OB-Rb isoform. In addition, an induction of the immediate-early gene c-fos is observed in jejunum in vivo. Leptin-mediated induction of a number of immediate-early genes and activation of STAT3 and STAT5 in a human model of small intestine epithelium, CACO-2 cells, corroborate this effect. Furthermore, intravenous leptin administration caused a significant 2-fold reduction in the apolipoprotein AIV transcript levels in jejunum 90 min after a fat load. Our results suggest that the epithelium of jejunum is a direct target of leptin action, and this activity is dependent on the presence of OB-Rb. Lack of leptin or resistance to leptin action in this site may contribute to obesity and its related syndromes by directly affecting lipid handling.