1. The effect of pretreatment with pertussis toxin has been studied on responses to muscarinic agonists in guinea-pig atria and smooth muscle in vitro. 2. 48 h after a single intravenous injection of pertussis toxin (3.2-100 micrograms.kg-1), muscarinic receptor-mediated negative inotropic responses in the atria were inhibited in a dose-dependent manner, with complete abolition of responses occurring after administration of 100 micrograms.kg-1. 3. In contrast, there was no effect on atrial positive inotropic responses to isoprenaline. In addition, no effect was observed on contractile responses to carbachol and pilocarpine in the ileum, trachea, oesophageal muscularis mucosae and urinary bladder, either in terms of potency or maximal response, at all dose levels of pertussis toxin studied. 4. It is concluded that muscarinic receptors in the atria, but not smooth muscle, are probably coupled to the inhibitory regulatory protein Ni, which is functionally inactivated by pertussis toxin. The differences in coupling between atrial and smooth muscle muscarinic receptors provide further evidence for muscarinic receptor heterogeneity in these two tissues.