PBEF/Visfatin is an adipokine highly enriched in adipose tissue which is increased in obesity and has been shown to be involved in numerous physiological processes. The adrenal gland plays a pivotal role in the integration of metabolic activity and energy balance, through production of steroids, mainly glucocorticoids. Steroidogenesis is a tightly controlled and essential process, which is mediated by steroidogenic acute regulatory protein (StAR), a protein that transports cholesterol across the mitochondrial membrane and is also the rate limiting step of this process. We have previously shown that adipokines are potent regulators of both StAR expression/activity and steroidogenesis via activation of multiple signalling pathways in the pluripotent human adrenal cell line H295R. We therefore investigated the role of PBEF/visfatin on StAR activation in these cells. H295R cells were cultured in EMEM media before being treated with a range of recombinant visfatin concentrations. Using qRT-PCR and Western blot analysis, we observed significant time- and dose-dependent up-regulation of StAR mRNA and protein expression after 4 h. Incubation with visfatin induced phosphorylation of MAPK and PI3/AkT pathways. We found that direct inhibition of MAPK/PI3 (AkT) pathways, using specific inhibitors for ERK1/2, p38 MAPK and PI3/AkT, resulted in significant down-regulation of visfatin-induced StAR protein expression. Furthermore, using sandwich ELISA, visfatin was found to increase steroid release into cell culture media. This is the first study describing a role for PBEF/visfatin in the regulation of StAR expression/activity and steroid production. We also comprehensively described the signalling pathways involved in regulating visfatin-induced complexity of hormone biosynthesis in these cells. These data confirm the importance of adipokines in the process of steroidogenesis and suggest that adipokines may play a key regulatory role in integration of metabolic activity and energy balance via the adrenal gland.