Chemical characterization of leptin-activated neurons in the rat brain

Leptin has profound effects on food intake, body weight, and neuroendocrine status. The lack of leptin results in hormonal and metabolic alterations and a dramatic increase in body weight. Leptin acts in the brain, especially in the hypothalamus; however, the central nervous system sites that respond to leptin have not been examined comprehensively. In this study, we explored systematically the distribution of leptin-activated neurons throughout the rat brain. Furthermore, we investigated the chemical identity of subsets of these leptin-activated cells. Fos-like immunoreactivity (Fos-IR) was investigated in the rat brain after two different doses of leptin (1.0 mg/kg and 5.0 mg/kg) at 2 hours and 6 hours after injections. The induction of Fos-IR was observed in hypothalamic nuclei, including the paraventricular nucleus (PVH), the retrochiasmatic area (RCA), the ventromedial nucleus (VMH), the dorsomedial nucleus (DMH), the arcuate nucleus (Arc), and the ventral premammillary nucleus (PMV). In addition, leptin-induced Fos-IR was found in several nuclei of the brainstem, including the superior lateral and external lateral subdivisions of the parabrachial nucleus (slPB and elPB, respectively), the supragenual nucleus, and the nucleus of the solitary tract (NTS). By using double-labeling immunohistochemistry or immunohistochemistry coupled with in situ hybridization, leptin-activated neurons were found that contained cocaine- and amphetamine-regulated transcript mRNA in several hypothalamic nuclei, including the RCA, Arc, DMH, and PMV. In the Arc and DMH, leptin-induced Fos-IR was observed in neurons that expressed neurotensin mRNA. Dynorphin neurons in the VMH and in the Arc also expressed Fos-IR. In the brainstem, we found that cholecystokinin neurons in the slPB and glucagon-like peptide-1 neurons in the NTS were activated by leptin. We also investigated the coexpression of Fos-IR and the long form of the leptin receptor (OBRb) mRNA. We found double-labeled neurons surrounding the median eminence and in the RCA, Arc, VMH, DMH, and PMV. However, in brainstem sites, very little OBRb mRNA was found; thus, there were very few double-labeled cells. These results suggest that leptin stimulates brain pathways containing neuropeptides that are involved in the regulation of energy balance, autonomic homeostasis, and neuroendocrine status.