Background an Aim. Cholecystokinin (CCK), peptide YY (PYY) and ghrelin have been proposed to act as satiety hormones. CCK and PYY are stimulated during meal intake by the presence of nutrients in the small intestine, especially fat, whereas ghrelin is inhibited by eating. The sequence of events (fat intake followed by fat hydrolysis and CCK release) suggests that this process is crucial for triggering the effects. The aim of this study was therefore to investigate whether CCK mediated the effect of intraduodenal (ID) fat on ghrelin secretion and PYY release via CCK-1 receptors. Methods: 36 male volunteers were studied in three consecutive, randomized, double blind, cross-over studies. 1) 12 subjects received an ID fat infusion with or without 120 mg orlistat, an irreversible inhibitor of gastrointestinal lipases, in comparison to vehicle. 2) 12 subjects received ID long chain fatty acids (LCF), ID medium chain fatty acids (MCF) or ID vehicle. 3) 12 subjects received ID LCF with and without the CCK-1 receptor antagonist dexloxiglumide (DEXLOX) or ID vehicle plus iv saline (placebo). ID infusions were given for 180 min. The effect of these treatments on ghrelin concentrations and PYY release was quantified. Plasma hormone concentrations were measured in regular intervals by specific RIA systems. Results: 1) ID fat induced a significant inhibition in ghrelin levels (p<0.01) and a significant increase in PYY concentrations (p<0.004). Inhibition of fat hydrolysis by orlistat abolished both effects. 2) LCF significantly inhibited ghrelin levels (p<0.02) and stimulated PYY release (p<0.008), whereas MCF were ineffective compared to controls. 3) DEXLOX administration abolished the effect of LCF on ghrelin and on PYY. ID fat or LCF significantly stimulated plasma CCK (p<0.006 and p<0.004) compared to saline. MCF did not stimulate plasma CCK release. Summary: Fat hydrolysis is essential to induce effects on ghrelin and PYY through the generation of LCF; MCF are ineffective. LCF furthermore stimulated plasma CCK release suggesting that peripheral CCK is the mediator of these actions. The CCK-1 receptor antagonist DEXLOX abolished the effect of ID LCF, both on ghrelin and PYY. Conclusion: Generation of LCF through hydrolysis of fat is a critical step for fat-induced inhibition of ghrelin and stimulation of PYY in humans; the signal is mediated via CCK release and CCK-1 receptors.