4-Aminopyridine and 3,4-diaminopyridine produced concentration-dependent contraction on guinea-pig isolated ileum incubated in Tyrode solution. The EC30 values were 1.14 x 10(-4) and 1.39 x 10(-4) M, respectively. 2. Calcium channel blockers such as verapamil, diltiazem, nifedipine, flunarizine, and lanthanum chloride antagonized the contracting effect induced by 4-aminopyridine and 3,4-diaminopyridine in guinea-pig isolated ileum. 3. Diazoxide and atropine sulphate behaved similarly as antagonists of the contracting effect induced by 4-aminopyridine and 3,4-diaminopyridine in guinea-pig isolated ileum. 4. It is concluded that the aminopyridines exert their effects through the release of acetylcholine from parasympathetic nerve terminals.