1. Iontophoretic application of acetylcholine to neurones in the medulla and pons of the unanaesthetized decerebrate cat excited 35.5% and inhibited 22%. This was not a representative sample of lower brain stem neurones, since most penetrations were made medially. Some cholinoceptive neurones were in the medial reticular formation.2. The time course of the response to different doses has been described.3. Nicotine and nicotinic agents 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP) and choline phenyl ether (TM1) excited neurones excited by acetylcholine but, with one exception, did not inhibit neurones inhibited by acetylcholine.4. Muscarinic agents mimicked both the excitatory and inhibitory responses to acetylcholine. (+/-)-muscarine and bethanechol (beta-methylcholine carbamate) had a prolonged and powerful inhibitory action.5. Oxotremorine [1-(2-oxopyrrolidino)-4-pyrrolidino-butyne-2] had a long-lasting excitatory action on neurones excited by acetylcholine.6. Eserine and neostigmine potentiated the action of acetylcholine but in addition they had a strong excitatory effect on many brain stem neurones, independently of the action of acetylcholine.7. Excitatory and inhibitory responses to acetylcholine could be antagonized by gallamine, hexamethonium and atropine. Dihydro-beta-erythroidine (DHbetaE) antagonized only excitatory responses. (+)-Tubocurarine was a weak antagonist.8. It is concluded that the excitatory responses have both nicotinic and muscarinic properties but the inhibitory responses are only muscarinic. Since the pharmacology of these two types of response is different, it is concluded that the inhibitory action is not due to an indirect effect from excitation of a neighbouring inhibitory neurone.