Naunyn-Schmiedeberg's Arch Pharmacol (1989) 340:107--110 Naunyn-Schmiedeberg's Archivesof Pharmacology © Springer-Verlag1989 Contractile activity of the N-acylated C-terminal part of substance P7- 11 in guinea pig trachea Effect of epithelium removal Eric Tschirhart 1., Pascal Schmitt 2, Claude Bertrand 1, Michel Mayer 2, Sylviane Magneney 2, Yves Landry 1, and Robert Michelot 2 1 Laboratoire de Neuroimmunopharmacologie, Universit6 Louis Pasteur, Strasbourg I, BP 24, F-67401 Illkirch Cedex, France 2 Institut de Chimie des Substances Naturelles, CNRS, F-91198 Gif Sur Yvette, France Summary. Substance P, neurokinin A, neurokinin B, and N-acylated pentapeptide X-Phe-Phe-Gly-Leu-Met-NH2 analogs of substance P7-11 were tested for their spas- mogenic activities in intact or in epithelium-denuded tra- cheal strips from guinea pig. Epithelium removal enhanced the efficacies and potencies relative to substance P of all the peptides tested in guinea pig trachea. In epithelium- containing preparations, the presence of a cyclic substituent (o-hydroxyphenyl-acetyl, p-hydroxyphenyl-acetyl, pyro- glutamyl) in Phe 7 greatly enhanced the potency and efficacy compared to substance P. These substitutions were twice as active as neurokinin A itself. The presence of an aliphatic chain (non-protected and t-butyloxycarbonyl-protected aminopropyl and aminocaproyl) in Phe 7 also improved the potency and the efficacy of the synthetic peptides. The ali- phatic substituents could favour an increase in local concen- tration of the peptides in the vicinity of the receptor(s) allowing a more effective ligand-receptor interaction. Thus, lipophilicity could be determinant in the potency of the peptides in intact guinea pig trachea. In epithelium-denuded tracheal strips from guinea pig, all the synthetic peptides were more effective than substance P but less active than neurokinin A which probably reflects the presence of the NK2 receptor subtype, which may be predominant in this type of epithelium-denuded preparation. Our results suggest that hydrophobicity plays a strong role in the interaction of the peptides, namely substance P and its analogues with the membrane and possibly the receptors themselves. Key words: Guinea-pig trachea - Substance P7-11 -- Air- way epithelium - Tachykinin receptor - Lipophilicity Introduction Substance P (SP), which is a potent bronchoconstrictor in the guinea pig both in vivo (Andersson and Persson 1976) and in vitro (Nilsson et al. 1977) belongs to the tachykinin family and shares with these peptides a common C-terminal sequence, Phe-X-Gly-Leu-Met-NH2, where X is Phe for SP * Present address: Merrell Dow Research Institute, Strasbourg Research Center, 16, Rue d'Ankara, F-67084 Strasbourg Cedex, France Send offprint requests to E. Tschirhart at the above address and Val, Tyr, or Ile for the other tachykinins. This C-terminal sequence is mostly responsible for their spasmogenic ac- tivity. This fragment is a full agonist on smooth muscle preparations but has a low potency. Greater potency is usually observed with larger fragments such as the hepta or octa C-terminal sequences (Bergman et al. 1974; Bury and Mashford 1976; Regoli et al. 1984). Recent reports have established that modifications starting from the C-terminal hexapeptide sequence may lead to active agonists (Poulos et al. 1986; Laufer et al. 1986) and these data seem to rule out the possibility of shorter fragments having valuable agonist or antagonist activity. In 1976, Niedrich and coworkers re- ported that N-acylation of pentapeptide terminal sequences of tachykinins could modify their potencies and more re- cently others have reported that the N-acylated pentapeptide SP7-11 could be more active than SPj_ 11 itself on smooth muscle (Bienert et al. 1983). This increase in activity was also recently correlated to an increase in lipophilicity of the peptides (Niedrich et al. 1981). In this study, the potency and activity of N-acylated pentapeptides in guinea pig trachea have been examined in order to evaluate to what extent slight modifications at the N-terminal part of this pentapeptide might modify the activity and the potency of the fragment. The guinea pig trachea was chosen since this preparation has been shown to be quite sensitive to short fragments of SP (Mizrahi et al. 1982). The receptors involved in the contractile activity of the neurokinins in the guinea pig trachea could be of the NK2 type as suggested by Regoli and coworkers (1987). Recent studies have suggested that the airway epithelium could modulate substance P-induced contraction in guinea pig upper airways by the release of an inhibitory factor (Tschirhart and Landry 1986) or via a possible direct metabolization of the peptides in the airway epithelium (Devillier et al. 1988). Therefore, the possible modulatory role of the airway epithelium was taken into account in the assessment of the bronchoconstrictor activity of the naturally occurring peptides and of the synthetic pentapeptides. Materials and methods Organ bath studies. Male albino guinea pigs (Elevage St. Antoine, Pleudaniel, France), weighing 350 to 400 g were anaesthetized with 45 mg/kg sodium pentobarbitone. Tra- cheas were rapidly dissected out and immersed in physiologi-