IDENTIFICATION OF A PITUITARY GROWTH HORMONE-RELEASING PEPTIDE (GHRP)RECEPTOR SUBTYPE BY PHOTOAFFINITY-LABELING

Hexarelin, an analogue of GHRP-6, in which D-Tryptophan has been repla ced by its 2-methyl derivative, is known to release growth hormone (GH ) in vivo and in vitro by direct action on receptors present in anteri or pituitary cells. Measurement of second messengers (c-AMP, Ca++, IP3 ) upon somatotrophs stimulation, suggests the existence of more than o ne GHRP receptor subtype. In order to document such an hypothesis, we have used a new photoactivatable derivative of Hexarelin, Tyr-Bpa-Ala- Hexarelin. This derivative was shown to be fully active in the release of GH in vivo with neonate rats. Using this photoactivatable ligand, we have specifically labeled a protein with an apparent M-r of 57 000 in human, bovine and porcine anterior pituitary membranes. Hexarelin a nd the spiroindoline sulfonamide MK-0677 displaced the M-r-57 000 phot olabeled band with an apparent ED50 of 6x10(-7) M and 2x10(-5) M respe ctively. Taking into account the high efficiency (>60%) of covalent in corporation of the Bpa residue, this photoactivatable Hexarelin deriva tive has allowed the identification of a pituitary GHRP receptor subty pe, which is apparently distinct from the recently cloned GH secretago gue receptor.