Ghrelin receptor in two species of anuran amphibian, bullfrog (rana catesbeiana), and japanese tree frog (hyla japonica)

We have identified cDNA encoding a functional growth hormone secretagogue-receptor 1a (GHS-R1a, ghrelin receptor) in two species of anuran amphibian, the bullfrog (Rana cates-beiana), and the Japanese tree frog (Hyla japonica). Deduced receptor protein for bullfrog and Japanese tree frog (tree frog) was comprised of 374- and 371-amino acids, respectively.The two receptors shared 86% identity, and are grouped to the clade of the tetrapod homologs by phylogenetic analysis. In functional analyses, ghrelin and GHSR1a agonists increased intracellular Ca²⁺ concentration in GHS-R1a-transfected-HEK293 cell, but ligand selectivity of ghrelin with Ser³ and Thr³ was not observed between the two receptors. Bullfrog GHS-R1a mRNA was mainly expressed in the brain, stomach, and testis. In the brain, the gene expression was detected in the diencephalon and mesencephalon, but not in the pituitary. Tree frog GHS-R1a mRNA was predominantly expressed in the gastrointestinal tract and ovary, but not detected in the pituitary. In bullfrog stomach but not the brain, GHS-R1a mRNA expression increased after 10 days of fasting. For tree frog, GHS-R1a mRNA expression was increased in the brain, stomach and ventral skin by 10 days of fasting, and in the stomach and ventral skin by a dehydration treatment. Intracere-broventricular injection of ghrelin in dehydrated tree frog did not affect water absorption from the ventral skin.These results suggest that ghrelin is involved in energy homeostasis and possibly in osmoregulation in frogs.