Fentanyl‐Induced Sleep Disruption in C57BL/6J (B6) Mice is not Secondary to Increased Motor Activity or Hypothermia

Abstract

Studies of mice have enhanced the understanding of opioid‐induced respiratory depression (10.1152/jn.00017.2021) and opioid abuse (10.1016/j.neuropharm.2013.08.028). The potential that mechanistic insights gained from studies of mice may be relevant for humans is consistent with the close genetic homology between these two species. Features of mouse biology, however, may limit the extent to which studies of opioid effects can be generalized. Fentanyl administered to mice can increase motor activity (10.1152/jn.00711.2020), cause hypothermia (10.1124/jpet.102.037655), alter cortical electroencephalogram (EEG) power, and disrupt the temporal organization of sleep (10.1152/jn.00266.2021). Our recent finding that fentanyl causes dose‐dependent sleep disruption in B6 mice (10.1096/fasebj.2021.35.S1.01706) led us to test the hypothesis that fentanyl‐induced increases in wakefulness are not caused by changes in motor activity or body temperature. The FASEB Statement of Principles for the use of Animals in Research and Education was followed. Adult males (n=8) were housed in a temperature‐ and humidity‐controlled room with a 12:12h light:dark cycle. Mice were implanted with electrodes and subcutaneous telemeters (DSI HD‐X02) to record EEG, electromyogram, motor activity, and body temperature. Seven to 10 days after recovery from surgery each mouse received a series of subcutaneous injections (0.3 mL) of saline (vehicle) and increasing half‐log doses of fentanyl (0.001 to 3 mg/kg). All injections were made within 60 min of light onset and mice were recorded for 4 h. Injections in the same mouse were separated by 3 or 4 days. Repeated measures mixed model ANCOVAs were performed to evaluate percent time awake between fentanyl dose while accounting for the effects of motor activity and body temperature. Each ANCOVA model was evaluated to determine if the motor activity and body temperature slopes at each level of fentanyl were equal to zero by including the fentanyl main effect and the fentanyl by motor activity (p = 0.11) or fentanyl by body temperature (p = 0.19) interaction. This established the lack of dependence between fentanyl dose and motor activity or body temperature when explaining differences in percent of time in wakefulness. Subsequent common slope models were performed including the main effects for fentanyl with body temperature and fentanyl with motor activity to confirm no effects of either covariate on percent time awake. Motor activity (p = 0.611) and body temperature (p = 0.614) were not found to be significant predictors of increased wakefulness. This finding supports the interpretation that fentanyl causes sleep disruption that is not secondary to enhanced motor activity or hypothermia.