Leptin acts not only as an anorexigenic hormone but also regulates cell-mediated immunity via leptin receptors (Ob-R) expressed on T- and B-lymphocytes. However, the impact of leptin on NK cells is currently elusive. We evaluated leptin effects on NK cells in relation to the body weight in rats using in vivo and in vitro approaches. Leptin was injected intravenously in male lean and diet-induced obese Lewis and F344 rats. NK cell numbers were analyzed in blood and spleen by FACS and immunohistochemistry and the activity of NK cells was measured by chromium release assay. Ob-R expression was investigated by confocal laser scanning and qRT-PCR. To compare leptin-dependent intracellular signaling under basal, leptin- and tumor cell (MADB106) stimulated conditions, intracellular target proteins of NK cells were evaluated by Western blotting. Number and distribution pattern of splenic NK cells were significantly different in lean and obese animals. Leptin administration resulted in a 4 fold higher stimulation of the NK activity in lean than in obese animals. This was not due to a decreased expression of Ob-R as qRT-PCR revealed significantly higher Ob-Rb mRNA levels in NK cells from obese rats. In contrast, post receptor signaling is differentially abrogated in obese animals with significantly lower activation of post-receptor signaling components (JAK-2p, PKBpT308, AMPK-pT172) following an in vivo leptin challenge. The results for the first time assign leptin a central role as a modulator of NK cell numbers and activity only in lean but not in obese subjects. The differential role of leptin has important implications for the influence of body weight in the response to systemic inflammations and in the immunological defense of cancer.