| Budget Amount *help |
¥6,100,000 (Direct Cost: ¥6,100,000)
Fiscal Year 2001: ¥6,100,000 (Direct Cost: ¥6,100,000)
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| Research Abstract |
Small synthetic molecules termed growth hormone secretagogues (GHSs) act through an orphan 0protein coupled receptor, GHS-R, to stimulate growth hormone (GH) release from pituitary. To search for the natural ligand for GHS-R, we constructed a stable CHO cell line expressing rat GHS-R for monitoring intracellular calcium concentration ([Ca^<2+>]_i) changes induced by rat tissue extracts. Using this cell line, we purified a peptide from stomach that potently activated GHS-R. The purified peptide was consisted of 28 amino acids, in which the Ser-3 residue was n-octanoylated. Surprisingly, this n-octanoyl modification at Ser-3 was essential to the activity, since des-acyl-peptide could not induce ([Ca^<2+>]_i) changes. We named the GH-releasing peptide ghrelin (ghre is the Proto-Indo-European root of the word grow).
We cloned mammalian gbrelin cDNAs (human, rat, mouse, canine, porcine, ovine and bovine) and determined their structures. Precursor structures of manimalian ghrelin are highly h
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omologous. In particular, N-terminal 10 amino-acids are identical, which supports the importance of octanoyl modification for ghrelin s activity.
Ghrelin induced GH release in a dose-dependent manner both in vitro and in vivo. ICV injection of ghrelin also stimulated GH release. Ghrein immunoreactive cells in the stomach were found in X/A-like cells, an endocrine cell in stomach. Ghrelin is found in the secretary granules of X/A-like cells whose hormonal product and physiological functions have not been previously clarified. In rat brain, ghrelin immunoreactive neurons were found to be localized in the hypothalamic arcuate nucleus, a region where regulates food intake. In fact, ICV injection of ghrelin strongly stimulated feeding in rats and increased body weight gain. On the other hand, anti-ghrelin IgG robustly suppressed feeding. These facts suggest that ghrelin is a new physiological regulator of nutritional homeostasis.
Recently we purified ghrein peptides from chicken, frog (bullfrog) and fish (eel and trout). Aminoacid sequences of ghrelin peptides in all vertebrates are well conserved. Moreover, the ghrelin peptides are all modified by octanoic acid. These results suggest that ghrelin plays a fundamentally important role in vertebrates. Less
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