Immersion in tricaine methanesulfonate (MS-222) is insufficient for euthanasia in at least one species of fish. The current study investigated the effectiveness of potassium chloride (KCl) to euthanize anesthetized koi (Cyprinus carpio). Twenty-eight healthy koi were anesthetized via immersion in 500 mg/L of buffered MS-222 for 10–12 min, manually removed to room air, and randomly administered 10 mEq/kg KCl (333 mg/ml) via one of three routes—intracardiac injection (IC) (n = 7), intracoelomic injection (ICe) (n = 7), or topical instillment over the gill filaments bilaterally (T) (n = 7)—or received no treatment (control, C) (n = 7). A Doppler ultrasonic flow detector was placed over the heart, and sounds were assessed continuously from immediately prior to treatment until 5 min posttreatment and every 5 min thereafter until Doppler sound cessation, resumption of operculation, or 30 min. Time to Doppler sound cessation or resumption of operculation was recorded. Doppler sound cessation occurred in 7/7 fish in IC (median 0.08, range 0–2.75 min) and 1/7 fish in T (10 min). In T, ICe, and C, 6/7, 7/7, and 7/7 fish, respectively, maintained Doppler sounds to 30 min. All fish in ICe (7/7) and C (7/7) resumed operculation with median (range) times of 22 (7–30) min and 16 (9–29) min from treatment, respectively. Intracardiac KCl at 10 mEq/kg rapidly ceases Doppler sounds and is a successful technique for euthanasia of anesthetized koi. Intracoelomic and topical KCl at 10 mEq/kg were not effective for euthanasia of koi.