Ghrelin is a powerful orexigenic peptide predominantly secreted by the stomach. Blood concentration of ghrelin increases before meals and fall postprandial. Its regulation appears to be influenced by the type of macronutrient ingested, the vagus nerve stimulation and by other post-meal stimulated hormonal factors. However, the direct role of nutrients (amino acids or lipids), neuronal (vagal neurotransmitter acetylcholine) and satiety-inducing factor such as CCK are not known. To study this we applied amino acids, lipids, acetylcholine and CCK via vascular perfusion to the isolated stomachs and found that amino acids significantly reduced ghrelin release from the isolated stomach by approximately ∼30% vs. the control while lipids (10% intralipid) had no affect. Acetylcholine (1 μM) increased ghrelin release from the stomach by ∼37% whereas insulin (10 nM) decreased it by ∼30% vs. the control. Interestingly, CCK (100 nM) potently increased ghrelin release by ∼200% vs. the control. Therefore it appears that ghrelin secretion from the stomach is under direct influence of amino acids, neurotransmitter acetylcholine and hormones such as insulin and CCK.