The aim of this study is to explore the mechanisms by which electroacupuncture (EA) enhances the hypoglycemic effect of exogenous insulin in a streptozotocin- (STZ-) diabetic rats. Animals in the EA group were anesthetized and subjected to the insulin challenge test (ICT) and EA for 60 minutes. In the control group, rats were subjected to the same treatment with the exception of EA stimulation. Blood samples were drawn to measure changes in plasma glucose, free fatty acids (FFA), and insulin levels. Western blot was used to assay proteins involved in insulin signaling. Furthermore, atropine, hemicholinium-3 (HC-3), and Eserine were used to explore the relationship between EA and cholinergic nerve activation during ICT. EA augmented the blood glucose-lowering effects of EA by activating the cholinergic nerves in STZ rats that had been exposed to exogenous insulin. This phenomenon may be related to enhancement of insulin signaling rather than to changes in FFA concentration.