Ceftriaxone (CTX) is a third-generation cephalosporin that has proven to be effective in the treatment of infections caused by a wide range of gram-positive and gram-negative microorganisms. This study aimed to determine the plasma and muscle pharmacokinetics of CTX after its administration via the intravenous (IV) and intramuscular (IM) routes to brown trout (Salmo trutta fario) at temperatures of 10 °C–13 °C. In total, 140 healthy brown trout (body weight, 245 ± 38 g) were used. The brown trout received single IV and IM injections of CTX at 25 mg/kg. The IV doses were injected into the caudal vein, whereas the IM doses were injected into the right epaxial muscles. The plasma and muscle tissue concentrations of CTX were measured using high-performance liquid chromatography. Pharmacokinetic parameters were calculated using noncompartmental methods. Following the IV administration of CTX, the elimination half-life (t1/2ʎz), volume of distribution at steady state, total body clearance, and area under the concentration–time curve (AUC0–72) in plasma were 5.83 h, 0.09 L/kg, 0.02 L/h/kg, and 1079.46 h*μg/mL, respectively. After the IM administration of CTX, plasma t1/2ʎz, peak plasma concentration (Cmax), time to reach Cmax, and bioavailability were 22.78 h, 87.92 μg/mL, 0.5 h, and 27.19%, respectively. The AUCMuscle/AUCPlasma ratio following the IV administration was 0.02 and that following the IM administration was 0.04. CTX exhibited low bioavailability and prolonged t1/2ʎz after the IM administration. The prolonged t1/2ʎz of CTX could thus be beneficial in brown trout. Nevertheless, future studies that aim to determine the clinical efficacy and pharmacokinetics after repeated administration of CTX are warranted.