I-125-antisauvagine-30: a novel and specific high-affinity radioligand forthe characterization of corticotropin-releasing factor type 2 receptors

Corticotropin-releasing factor (CRF) receptors type 1 (CRF1) and type 2 (CR F2) differ from each other in their pharmacological properties. The human a nd ovine CRF versions bind to CRF1 receptors with significantly higher affi nity than to CRF2 receptors. Recently antisauvagine-30, an N-terminally tru ncated version of the CRF analog sauvagine, was characterized as a specific antagonist to mouse CRF2B. We have synthesized the radiolabeled version I- 125-antisauvagine-30 and tested it for its affinity at human CRF1 (hCRF(1)) , hCRF(2A), Xenopus CRF1 (xCRF(1)) and xCRF(2) receptors. In control bindin g studies I-125-labeled hCRF, sauvagine and astressin were also bound to th ese receptors. I-125-antisauvagine-30 exclusively bound to hCRF(2A) and xCR F(2) but not to hCRF(1) and xCRF(1) receptors. I-125-antisauvagine-30 bindi ng to hCRF(2A), and xCRF(2) receptors was saturable and of high affinity (h CRF(2A): K-d=125 pM; xCRF(2): K-d=1.1 nM). In displacement binding experime nts using I-125-antisauvagine-30 as radioligand several CRF analogs bound t o hCRF(2A) and xCRF(2) receptors with similar rank orders as reported with other CRF radioligands. Finally, preliminary studies using I-125-antisauvag ine-30 binding to membrane homogenates prepared from different rat brain st ructures showed that the peptide bound specifically to brain areas expressi ng CRF2 receptors. These data demonstrate that I-125-antisauvagine-30 is th e first high-affinity ligand to specifically label CRF2 receptors. (C) 2000 Elsevier Science Ltd. All rights reserved.