We describe the effect of eight different imidazoline/guanidinium compounds on the settlement and metamorphosis of larvae of the barnacle Balanus improvisus. These agents were chosen on the basis of their similar pharmacological classification in vertebrates and their chemical similarity to medetomidine and clonidine, previously described as highly potent settlement inhibitors (nanomolar range). Seven of the tested compounds were found to inhibit settlement in a dose-dependent manner in concentrations ranging from 100 nM to 10 µM without any significant lethal effects. In vertebrate systems these substances have overlapping functions and interact with both α-adrenoceptors as well as imidazoline binding sites. Antagonizing experiments using the highly specific α2-antagonist methoxy-idazoxan or agmatine (the putative endogenous ligand at imidazoline receptors) were performed to discriminate between putative pharmacological mechanisms involved in the inhibition of cyprid settlement. Agmatine was not able to reverse the effect of any of the tested compounds. However, methoxy-idazoxan almost completely abolished the settlement inhibition mediated by guanabenz (α2-agonist, I2 ligand), moxonidine (α2-agonist, I1 ligand) and tetrahydrozoline (α-agonist, I2 ligand). The actions of cirazoline (α1-agonist, I2 ligand) BU 224 (I2 ligand) and metrazoline (I2 ligand) were not reversed by treatment with methoxy-idazoxan. These results suggest that the settlement inhibition evoked by the I2 ligands and α2-agonists used in this study of the neurologically simple but well-organized barnacle larva is mediated through different physiological targets important in the overall settlement process. J. Exp. Zool. 303A:551–562, 2005. © 2005 Wiley-Liss, Inc.