ObjectiveTo evaluate the pharmacokinetics of amitriptyline and its active metabolite nortriptyline after intravenous (IV) and oral amitriptyline administration in healthy dogs. Study designProspective randomized experiment. AnimalsFive healthy Greyhound dogs (three males and two females) aged 2–4 years and weighing 32.5–39.7 kg. MethodsAfter jugular vein catheterization, dogs were administered a single oral or IV dose of amitriptyline (4 mg kg−1). Blood samples were collected at predetermined time points from baseline (0 hours) to 32 hours after administration and plasma concentrations of amitriptyline and nortriptyline were measured by liquid chromatography triple quadrupole mass spectrometry. Non-compartmental pharmacokinetic analyses were performed. ResultsOrally administered amitriptyline was well tolerated, but adverse effects were noted after IV administration. The mean maximum plasma concentration (CMAX) of amitriptyline was 27.4 ng mL−1 at 1 hour and its mean terminal half-life was 4.33 hours following oral amitriptyline. Bioavailability of oral amitriptyline was 6%. The mean CMAX of nortriptyline was 14.4 ng mL−1 at 2.05 hours and its mean terminal half-life was 6.20 hours following oral amitriptyline. Conclusions and clinical relevanceAmitriptyline at 4 mg kg−1 administered orally produced low amitriptyline and nortriptyline plasma concentrations. This brings into question whether the currently recommended oral dose of amitriptyline (1–4 mg kg−1) is appropriate in dogs.