Neuropeptide B (NPB) regulates appetite, reduces obesity and promotes energy expenditure by activating NPBW1 and NPBW2 receptors. Humans express both types of receptors, while rodents NPBW1, only. Studies using genetically engineered mice depleting NPBW1 and neuropeptide B-deficient mice developed mild adult onset obesity. These data suggested that NPB may be involved in controlling adipose tissue functions. Nevertheless, the role of NPB in controlling of preadipocytes biology and adipogenesis is unknown. Thus, we evaluated the effects of NPB on brown and white preadipocytes proliferation and differentiation. The role of NPB in adipogenesis was evaluated in vitro using isolated rat white and brown preadipocytes. All procedures concerning welfare of animals were accorded with the ethical standards by the National Ethics Commission for Investigations on Animals in Poland. Cells were isolated and cultured in DMEM/F12 with 10% FBS . Proliferation was measured using BrdU ELISA kit (1, 10 and 100 nM NPB and control group; 24 h). Experiments assessing the role of NPB in differentiation were performed using differentiating medium containing T3, insulin and/or dexamethasone with or without NPB. Expression of adipogenic genes were analysed in 1, 3 and 6th days using real-time PCR. Intracellular lipid accumulation was analysed using ORO staining and microphotographic documentation. Glycerol released into the medium was measured using Free Glycerol Reagent. Analysis of variance followed by the Bonferroni post hoc test was used to determine statistical significance. Values are means ± S.E.M. We found Npb and Npw1 mRNA expression in rat white and brown preadipocytes. Our findings indicates that 10 and 100 nM NPB stimulates brown (0,388 ± 0,040 and 0,410 ± 0,024 vs. control 0,270 ± 0,021 OD450nm; n=8, p<0,05), but not white, preadipocytes proliferation. Next, we found that NPB increased triacylglycerol accumulation induced by differentiation medium in rat brown (0,176 ± 0,013 OD520nm vs. 0,191 ± 0,011 and 0,192 ± 0,017 for 10 and 100 nM NPB treated) and white (0,200 ± 0,005 OD520nm vs. 0,235 ± 0,008 and 0,234 ± 0,072 for 10 and 100 nM NPB treated) adipocytes evaluated at 1th day (n=6, p<0,05). Moreover, NPB increases glycerol release in brown adipocytes (0,2309 ± 0,005 and 0,2249 ± 0,006 nmol/l for 10 and 100 nM NPB vs. control 0,2016 ± 0,006 at 1th ; 0,2275 ± 0,005 nmol/l vs. 0,2673 ± 0,008 NPB 100 nM treated at 3th day of differentiation; n=7, p<0,05). NPB at 100 nM increased Ucp-1 and Prdm16 mRNA expression in brown adipocytes at 1, 3 and 6th day of differentiation (n=5, p<0,01). Expression of Pparγ and C/ebpβ were increased by 100 nM of NPB in white adipocytes differentiated for 1 day (n=5, p,0,05). In summary, we found that NPB stimulates proliferation and differentiation of rat white and/or brown preadipocytes into mature adipocytes. These results suggest that NPB may contribute to energy homeostasis regulation by controlling brown and white adipogenesis.