In isolated rat iris sphincter muscle, there was no attempt to measure mechanical tension changes, because of the small size of the preparation. In this study, responses of the isolated rat iris sphincter to some agents and electrical stimulation were examined. Acetylcholine and electrical stimulation produced powerful contractions of the iris sphincter. These contractile responses were suppressed by atropine and enhanced by physostigmine. Norepinephrine (10 .mu.M) induced a weak contraction of the sphincter muscle and 1 mM isoproterenol induced a very weak relaxation. These responses were antagonized by phentolamine and propranolol, respectively. In the presence of 0.1 .mu.M atropine, electrical stimulation produced a weak .alpha.-adrenergic contraction and a very weak .beta.-adrenergic relaxation. Electrically induced responses were abolished by tetrodotoxin. In the rat iris sphincter, powerful contraction is due to the activation of muscarinic receptors, and there are weak .alpha.-adrenergic contraction and weak .beta.-adrenergic relaxation. In rats, muscarinic contraction of the sphincter muscle is important in the regulation of pupil diameter.