OBJECTIVE: To identify any systemic effects of topical and subconjunctival administration of atropine sulfate in the horse. Animals studied Six mature grade horses were treated hourly in one eye with topical ophthalmic atropine drops for 24 h. Five horses were treated subconjunctivally in one eye with 3 mg of atropine sulfate. Procedures Pupillary light reflexes, pupil size, electrocardiographic parameters, girth measurements, intestinal motility, and clinical signs of abdominal pain were monitored. RESULTS: Alteration in auscultated gut motility and clinical signs of abdominal pain were the most sensitive indicators of the systemic manifestations of the topically applied atropine. Gut motility was absent in all horses for periods of 2-18 h in all four abdominal quadrants in horses given topically administered atropine. Signs of abdominal pain were observed in four of six horses that received topical atropine. In the subconjunctival test study, gut motility was absent in three horses for periods of 3-7 h. Uniocular subconjunctival injection of 3 mg atropine sulfate produced signs of abdominal pain in one of six horses. Conclusion The ophthalmic administration of atropine can affect gut motility and induce signs of colic in selected horses.