Plasma glucose and insulin responses to a muscarinic agonist (bethanechol chloride) and a muscarinic antagonist (atropine) were evaluated in obese C57BL/6J ob/ob mice and in lean C57BL/6J + /? mice. In lean +/? mice, plasma glucose decreased in response to 1 and 2 micrograms/g bethanechol chloride, whereas insulin increased significantly. In ob/ob mice, insulin increased remarkably in response to bethanechol administration (saline, 632 +/- 80 microU/ml; 2 micrograms/g bethanechol chloride, 1794 +/- 97 microU/ml; n = 10), but surprisingly, plasma glucose also rose significantly (saline, 230 +/- 14 mg/dl; 2 micrograms/g bethanechol chloride, 363 +/- 18 mg/dl, n = 10). This exaggerated hyperglycemia in ob/ob mice was not associated with significant changes in plasma glucagon. Furthermore, administration of propranolol hydrochloride did not diminish bethanechol chloride-induced hyperglycemia in ob/ob mice. Administration of atropine (2.5, 5, and 10 mg/kg body wt) induced a significant decrease in plasma insulin without changes in plasma glucose in ob/ob mice, whereas neither plasma insulin nor plasma glucose changed in lean mice. Finally, conversion of [14C]alanine to glucose was increased in ob/ob mice after bethanechol chloride administration, indicating that muscarinic stimulation increases gluconeogenesis in an animal model of type II (non-insulin-dependent) diabetes.