A series of tetraamines related to methoctramine (1) was synthesized and evaluated for its blocking activity on M2 and M3 muscarinic receptors of guinea pig left atria and ileum, respectively. Thus, tetraamines 2-7 were synthesized to evaluate the effect on affinity of replacing the 2-methoxybenzyl moiety of methoctramine by a phenethyl-type substituent. Furthermore, tetraamines 8 and 9 were investigated to analyze the effect on affinity of the chain length separating the inner nitrogens and the inner from outer nitrogens while keeping the total distance between the two outer nitrogens equal to that of methoctramine. It turned out that all the tetraamines investigated, although showing a significant affinity, were less active than methoctramine at M2 muscarinic receptors. The underlying drug-receptor interaction mechanisms are discussed.