Hexarelin modulates the expression of growth hormone secretagogue receptor type 1a mRNA at hypothalamic and pituitary sites.

Ghrelin and the synthetic growth hormone secretagogues (GHSs) activate a G-protein-coupled receptor (GHS-R) originally cloned from the pituitary, but which is also expressed in the hypothalamus, in other areas of the brain and in numerous peripheral tissues. Several studies have shown that growth hormone (GH)-releasing hormone (GHRH) is necessary for GHSs to exert maximal GH release in vivo. The exact mechanism of this synergism is not clear. Previous data suggest that GHSs can affect pituitary GHS-R mRNA expression; however, it is unknown whether this effect is age dependent and whether hypothalamic GHS-Rs are also affected. In this study, we tested whether (a) the synthetic GHS hexarelin regulates mRNA expression of its own receptor at the pituitary and/or hypothalamus and whether this effect is age dependent, and (b) whether short-term treatment with GHRH or, conversely, passive immunization against GHRH affects pituitary GHS-R1a mRNA expression in infant (10 days old) and young adult rats. GHS-R1a mRNA expression was measured with competitive reverse transcriptase-polymerase chain reaction. Hexarelin treatment significantly increased pituitary and hypothalamic GHS-R1a mRNA levels in normal infant rats, but not in normal young adult rats. In addition, hexarelin administration also stimulated pituitary GHS-R1a mRNA in infant as well as in young adult rats passively immunized against GHRH. GHRH treatment significantly enhanced pituitary GHS-R1a mRNA expression in GHRH-deprived young adult rats, though it did not affect the basal levels of GHS-R1a mRNA in normal infant and adult rats. These data further support the hypothesis that GHRH can affect GHS-R1a expression and that hexarelin upregulates the expression of its own receptor at the pituitary as well as the hypothalamus in an age-dependent fashion.

Ontogeny and tissue-specific regulation of ghrelin mRNA expression suggest that ghrelin is primarily involved in the control of extraendocrine functions in the rat.

Ghrelin is a 28-amino-acid gastric peptide that potently stimulates growth hormone (GH) secretion in vivo and in vitro. Ghrelin-expressing cells have been found in the oxyntic region of the stomach and in the arcuate nucleus of the hypothalamus. The aim of this work was to investigate the regional distribution and developmental changes in ghrelin mRNA levels in the pituitary, hypothalamus and gastrointestinal (GI) tract of the rat using a semiquantitative RT-PCR assay. We also describe the effects of ghrelin immunoneutralization in late gestation and those resulting from induction of an isolated GH deficiency in adult rats. Ghrelin mRNA was already expressed in the fetus by embryonic day 12 (E12), by E17 most of ghrelin mRNA was in the trunk. At E17, in situ hybridization did not reveal a clear expression of ghrelin mRNA in fetal stomach but showed high ghrelin mRNA levels in the placenta. In the pituitary gland, levels of ghrelin mRNA were high after birth but declined significantly with puberty, whereas in the hypothalamus they were barely detectable at birth and remained very low at all subsequent time points tested. In the GI tract, ghrelin mRNA levels were high from birth to 270 days of life. Immunoneutralization of ghrelin at E16 had no effect on survival or development. Rats showed normal somatotropic function, ghrelin expression and onset of puberty. In young adult rats, passive immunization against GHRH did not affect ghrelin mRNA levels in the pituitary, hypothalamus and stomach. Only a 72-hour fasting period induced a significant increase in ghrelin mRNA levels in the stomach, but not in the pituitary and hypothalamus. These results strongly indicate that ghrelin is an important GI hormone expressed early in life and primarily sensitive to nutritional status.

Short ghrelin peptides neither displace ghrelin binding in vitro nor stimulate GH release in vivo.

Ghrelin is an acylated peptide recently isolated from rat stomach that potently stimulates GH release in vitro and in vivo in rat and man. Ghrelin specifically activates the receptor for the growth hormone secretagogues (GHS-Rla), and it has been proposed as the endogenous ligand mimicked by these synthetic compounds. Very recently, it was shown in cells transfected with the GHS-Rla that short acylated peptides encompassing the first 4-5 residues of ghrelin were capable of increasing intracellular calcium almost as efficiently as the full-length ghrelin. In the present study, we demonstrate that truncated analogs of ghrelin are ineffective in stimulating GH release in neonatal rats and do not displace radiolabelled ghrelin from binding sites in membranes from human hypothalamus and pituitary. In conclusion, our data demonstrate that the ability of short ghrelins to stimulate the GHS-Rla in transfected cells is not predictive of their capability to stimulate GH secretion in vivo.