Pharmacological and chemical properties of astressin, antisauvagine-30 and α-helCRF: Significance for behavioral experiments

Corticotropin releasing factor (CRF) represents an early chemical signal in the stress response and modulates various brain functions through G protein-coupled receptors. Two CRF receptor subtypes, CRF₁ and CRF₂, have been identified. Since the physicochemical properties of CRF receptor antagonists might influence their biological potency, the peptidic antagonists astressin, α-helical CRF_9-41 (α-helCRF) and antisauvagine-30 (aSvg-30) have been analyzed. The rank order of solubility of these compounds in artificial cerebrospinal fluid (aCSF, pH 7.4) was aSvg-30>α-helCRF≫astressin, whereas the rank order of relative lipophilicity as determined with RP-HPLC was α-helCRF>astressin>aSvg-30. The calculated isoelectric points were 4.1 (α-helCRF), 7.4 (astressin) and 10.0 (aSvg-30). According to Schild analysis of the CRF receptor-dependent cAMP production of transfected HEK cells, aSvg-30 exhibited a competitive antagonism and displayed a 340 fold selectivity for mCRF_2β receptor. For astressin, however, the pharmacodynamic profile could not be explained by a simple competitive mechanism as indicated by Schild slopes >1 for rCRF₁ or mCRF_2β receptor. Behavioral experiments demonstrated that after i.c.v. injection, α-helCRF reduced oCRF-induced anxiety-like behavior in the elevated plus-maze, whereas astressin, despite its higher in vitro potency, did not. These findings could be explained by different physicochemical properties of the antagonists employed.