Anesthesia and Intubation of Preadolescent Mouse Pups for Cardiothoracic Surgery

Murine surgical models play an important role in preclinical research. Mechanistic insights into myocardial regeneration after cardiac injury may be gained from cardiothoracic surgery models in 0-14-day-old mice, the cardiomyocytes of which, unlike those of adults, retain proliferative capacity. Mouse pups up to 7 days old are effectively immobilized by hypothermia and do not require intubation for cardiothoracic surgery. Preadolescent (8-14-day-old) mouse pups, however, do require intubation, but this is challenging and there is little information regarding anesthesia to facilitate intubation. Here, we present dosage regimens of ketamine/xylazine/atropine in 10-day-old C57BL/6J mouse pups that allow endotracheal intubation, while minimizing animal mortality. Empirical titration of ketamine/xylazine/atropine dosage regimens to body weight indicated that the response to anesthesia of mouse pups of different weights was non-linear, whereby doses of 20/4/0.12 mg/kg, 30/4/0.12 mg/kg, and 50/6/0.18 mg/kg facilitated intubation of pups weighing between 3.15-4.49 g (n = 22), 4.50-5.49 g (n = 20), and 5.50-8.10 g (n = 20), respectively. Lower-body-weight pups required more intubation attempts than heavier pups (p < 0.001). Survival post-intubation correlated with body weight (59%, 70%, and 80% for low-, mid-, and high-weight groups, respectively, R² = 0.995). For myocardial infarction surgery after intubation, a surgical plane of anesthesia was induced with 4.5% isoflurane in 100% oxygen and maintained with 2% isoflurane in 100% oxygen. Survival post-surgery was similar for the three weight groups at 92%, 86%, and 88% (p = 0.91). Together with refinements in animal handling practices for intubation and surgery, and minimizing cannibalization by the dam post-surgery, overall survival for the entire procedure (intubation plus surgery) correlated with body weight (55%, 60%, and 70% for low-, mid-, and high-weight groups, respectively, R² = 0.978). Given the difficulty encountered with intubation of 10-day old pups and the associated high mortality, we recommend cardiothoracic surgery in 10-day-old pups be restricted to pups weighing at least 5.5 g.