Abstract

We have prepared a series of conformationally constrained hexapeptide analogs of substance P which are 500-1500-fold more potent as inhibitors of 125I-labeled Bolton Hunter-conjugated eledoisin binding to rat brain cortex membranes than as inhibitors of 125I-labeled Bolton Hunter-conjugated substance P binding. These analogs stimulate guinea pig ileum contraction (ED50 1-16 nM) and stimulate rat vas deferens contraction (ED50 2-4 microM). However, these peptides are poor stimulators of rat salivation (greater than 40 nmol/100 g body weight). Thus, based on both their receptor potency and pharmacological potency, these peptides are potent and selective tachykinin analogs. These data indicate that a specific carboxyl-terminal conformation is recognized by the 125I-labeled Bolton Hunter-conjugated eledoisin binding site and that this conformation is different from the conformation recognized by the 125I-labeled Bolton Hunter-conjugated substance P binding site. Hexapeptides containing phenylalanine, isoleucine, and valine identical with the carboxyl-terminal sequences of substance P, eledoisin, and neurokinin B, respectively, were nearly equipotent as inhibitors of 125I-labeled Bolton Hunter-conjugated eledoisin binding. The valine analog was only approximately 5-fold less potent than the isoleucine and phenylalanine analogs as an inhibitor of 125I-labeled Bolton Hunter-conjugated substance P binding. Thus, unknown determinants in the amino-terminal sequences of substance P must strongly contribute to the carboxyl-terminal peptide selectivity and conformation. The contraction of guinea pig ileum induced by one of the conformationally constrained analogs is attenuated by pretreatment of the tissue with atropine (2 microM), while that induced by substance P methyl ester, a selective inhibitor of 125I-labeled Bolton Hunter-conjugated substance P binding, is not. Thus, the constrained analog has a higher affinity for the tachykinin receptors in the guinea pig myenteric plexus which are responsible for acetylcholine release than for the tachykinin receptors present on the smooth muscle cells.