Intrahepatic Injection of Sodium Pentobarbital as an Alternative to Intraperitoneal Injection for the Euthanasia of Rats (Rattus norvegicus).

The most commonly accepted method of rat euthanasia in North America is intraperitoneal injection of sodium pentobarbital (PB). However, misinjection can occur, and intraperitoneal PB may cause pain and distress. The objective of this study was to test an alternative method of euthanasia: intrahepatic injection of PB. A pilot study was conducted to develop a method of intrahepatic injections (evaluated using CT scans and test injections), followed by a full study comparing intraperitoneal (n = 14) and intrahepatic PB injections (n = 66) in adult rats. Full study outcomes were: 1) time from injection to loss of right- ing reflex (LORR), 2) time from injection to cessation of heartbeat (CHB), 3) number of failed euthanasia attempts, and 4) confirmation of successful intrahepatic injection or misinjection via necropsy. All injections were performed by a veterinary student. CT revealed that intrahepatic injections were feasible. Times (median [range]) to LORR and CHB were faster after successful intrahepatic injections (LORR, 3 s [1 to 5 s]; CHB, 8 s [2 to 242 s]) than after intraperitoneal injections (LORR, 89.5 s [73 to 110 s], CHB: 284.5 s [237 to 423 s]). The misinjection rate was higher with intrahepatic injections (59%) than with intraperitoneal injections (29%), but intrahepatic misinjection still resulted in fast and successful euthanasia (LORR, 29 s [1 to 96 s]; CHB, 216 s [12 to 330 s]), with the injectate distributed between the intraperitoneal and intrahepatic locations. The number of failed euthanasia attempts with intrahepatic injections was low (n = 2). Intrahepatic injections show potential as an alternative to intraperitoneal injections for rat euthanasia.

Pharmacologic parameters of MS222 and physiologic changes in frogs (Xenopus laevis) after immersion at anesthetic doses.

We evaluated the anesthetic efficacy of MS222 (dose, 1 or 2 g/L; pH 7) administered as an immersion bath (duration, 20 min) for nonbreeding female Xenopus leavis frogs (n = 33; average body weight, 103 ± 16 g). The acid acetic test, the withdrawal reflex, righting behavior, heart rate, respiratory frequency, and blood oxygen saturation were used to evaluate the level of anesthesia. Acetic acid and withdrawal reflex responses were present at 30 and 60 min following immersion for the 1- and 2-g/L doses, respectively. MS222 had no effect on heart rate or oxygen saturation, but caused pronounced respiratory depression, as expected. Microscopic observations of selected tissues (heart, lung, liver, kidneys, and skin) showed no evidence of lesions at 24 h after immersion. In addition, we calculated the pharmacokinetics of MS222 in plasma and analyzed the drug by HPLC-tandem mass spectrometry. The calculated half-life of MS222 is 3.2 h. We conclude that MS222 administered at 1 or 2 g/mL via immersion bath for 20 min is an effective anesthetic that can be used for surgical procedures of less than 30 or 60 min, respectively, in Xenopus leavis.

Electroencephalographic and physiologic changes after tricaine methanesulfonate immersion of African clawed frogs (Xenopus laevis).

The objective of this study was to determine electroencephalographic and complementary physiologic changes in Xenopus leavis frogs after bath immersion in MS222. We also evaluated the addition of sodium pentobarbital injected intracoelomi- cally 2 h after MS222 immersion to achieve euthanasia. Frogs (n = 9) weighing 105.5 ± 8.4 g (mean ± 1 SD) were immersed in MS222 at either 1 or 3 g/L until anesthesia was achieved; a conductive stainless steel screw then was implanted in the skull on top of the outer pial surface of the brain. Frogs were immersed again in MS222 at the same concentration as previously, and electroencephalograms, heart rate, oxygen saturation, and respiratory movements were recorded. Amplitude and mean frequency of the electroencephalographic signal were evaluated at 15-min intervals until a flat-line signal was achieved. At 2 h after induction, frogs were injected intracoelomically with sodium pentobarbital (0.5 mL; 240 mg/mL) to accelerate euthanasia. Immersion of frogs in 1 or 3 g/L of MS222 depressed cerebral activity within 30 min without a significant effect on cardiac function. Intracoelomic injection of sodium pentobarbital at 2 h after MS222 administration rapidly (3.2 ± 1.7 min) induced cardiac arrest. In conclusion, immersion in MS222 can be used for the collection of organs from X. laevis frogs, but the addition of pentobarbital is required to achieve euthanasia.