Administration of ghrelin, an endogenous ligand for the growth hormone secretagogue receptor 1a (GHSR 1a), induces potent stimulating effects on GH secretion and food intake. However, more than seven years after its discovery, the role of endogenous ghrelin remains elusive. Recently a second peptide, obestatin, also generated from proteolytic cleavage of preproghrelin has been identified. This peptide inhibits food intake and gastrointestinal motility but does not modify in vitro GH release from pituitary cells. In this study we have reinvestigated obestatin functions by measuring plasma ghrelin and obestatin levels in a period of spontaneous feeding in ad libitum fed and 24h-fasted mice. While fasting resulted in elevated ghrelin levels, obestatin levels were significantly reduced. Exogenous obestatin per se did not modify food intake in fasted and fed mice. However, it inhibited ghrelin orexigenic effect that were evident in fed mice only. The effects of obestatin on GH secretion were monitored in superfused pituitary explants and in freely moving rats. Obestatin was only effective in vivo to inhibit ghrelin stimulation of GH levels. Finally, the relationship between octanoylated ghrelin, obestatin and GH secretions was evaluated by iterative blood sampling every 20 minutes during 6 hours in freely moving adult male rats. The half-life of exogenous obestatin (10 microg iv) in plasma was about 22 minutes. Plasma obestatin levels exhibited an ultradian pulsatility with a frequency slightly lower than octanoylated ghrelin and GH. Ghrelin and obestatin levels were not strictly correlated. In conclusion these results show that obestatin, like ghrelin, is secreted in a pulsatile manner and that in some conditions; obestatin can modulate exogenous ghrelin action. It remains to be determined whether obestatin modulates endogenous ghrelin actions.