Neuropeptides, including neuropeptide Y and melanocortins, mediate lipolysis in murine adipocytes.

OBJECTIVE: To determine whether key appetite-regulating neuropeptides such as melanin-concentrating hormone (MCH), neuropeptide Y (NPY), and alpha-melanocyte-stimulating hormone (alpha-MSH), which are known to mediate energy balance through centrally mediated pathways, also have direct acute effects on the lipolytic activity of murine adipocytes. RESEARCH METHODS AND PROCEDURES: Fully differentiated 3T3-L1 adipocytes serum starved overnight in Dulbecco's modified Eagle medium containing 2% bovine serum albumin or freshly isolated mouse adipocytes were incubated for up to 2 hours in the absence and presence of 100 nM each of NPY, MCH, alpha-MSH, the melanocortin receptor agonist MTII, or isoproterenol as a control. Free fatty acids secreted into the incubation medium were measured using a commercially available nonesterified fatty acid C test kit. RESULTS: Treatment of 3T3-L1 cells with 100 nM NPY decreased basal free fatty acid secretion (basal, 0.006 +/- 0.001 vs. NPY, 0.001 +/- 0.0003 nM at 90 minutes; p < 0.05), whereas both alpha-MSH and MTII stimulated up to a 7-fold increase in free fatty acid release (MTII, 0.238 +/- 0.004 vs. basal, 0.024 +/- 0.002 nM at 2 hours; p < 0.05; and alpha-MSH, 0.22 +/- 0.005 vs. basal, 0.04 +/- 0.003 nM at 2 hours; p < 0.05). Treatment with 100 nM MCH had no effect on basal free fatty acid release or on alpha-MSH-induced lipolysis during concurrent treatment. Conversely, concurrent treatment with 100 nM NPY dramatically inhibited (by approximately 90%) alpha-MSH-induced lipolysis. Similar treatment of freshly isolated mouse adipocytes showed virtually identical results. DISCUSSION: In addition to their centrally mediated actions, appetite-regulating neuropeptides modulate adipose tissue mass through direct peripheral effects. Systemic administration of pharmacological agents altering the effects of these neuropeptides may form the basis of future obesity therapies. Thus, some of these agents will likely have direct effects on adipocytes that may serve to alter their therapeutic effectiveness.

Melanin-concentrating hormone activates signaling pathways in 3T3-L1 adipocytes.

Energy homeostasis is regulated by peripheral signals, such as leptin, and by several orexigenic and anorectic neuropeptides. Recently, we reported that the orexigenic neuropeptide melanin-concentrating hormone (MCH) stimulates leptin production by rat adipocytes and that the MCH receptor (MCH-R1) is present on these cells. Here, we show that MCH-R1 is present on murine 3T3-L1 adipocytes. Treatment of 3T3-L1 adipocytes with 1 micromolar MCH for up to 2 h acutely downregulated MCH-R1, indicating a mechanism of ligand-induced receptor downregulation. Potential signaling pathways mediating MCH-R1 action in adipocytes were investigated. Treatment of 3T3-L1 adipocytes with 1 micromolar MCH rapidly induced a threefold and a fivefold increase in p44/42 MAPK and pp70 S6 kinase activities, respectively. In addition, 3T3-L1 adipocytes transiently transfected with a murine leptin-luciferase promoter construct showed a fourfold and a sixfold increase in leptin promoter-reporter gene expression at 1 h and 4 h, respectively, in response to MCH. Activity decreased to basal levels at 8 h. Furthermore, MCH-stimulated leptin promoter-driven luciferase activity was diminished in the presence of the MAP/ERK kinase inhibitor PD-98059 and in the presence of rapamycin, an inhibitor of pp70 S6 kinase activation. These results provide further evidence for a functional MCH signaling pathway in adipocytes.