Oral Fentanyl Consumption and Withdrawal Impairs Fear Extinction Learning and Enhances Basolateral Amygdala Principal Neuron Excitatory-Inhibitory Balance in Male and Female Mice

Abstract

The number of opioid overdose deaths has increased over the past several years, mainly driven by an increase in the availability of highly potent synthetic opioids, like fentanyl, in the illicit drug supply. While many previous studies on fentanyl and other opioids have focused on intravenous administration, other routes of administration remain relatively understudied. Here, we used a drinking in the dark (DiD) paradigm to model oral fentanyl self-administration using increasing fentanyl concentrations in male and female mice over 5 weeks. Fentanyl consumption peaked in both female and male mice at the 30 µg/mL dose, with female mice consuming significantly more fentanyl than male mice. Mice consumed enough fentanyl to precipitate withdrawal with naloxone, with males having more severe withdrawal symptoms. Fentanyl consumption disrupted normal sleep rhythms in both male and female mice. We also performed behavioral assays to measure approach/avoidance behavior and reward-seeking during fentanyl abstinence. Female mice showed more avoidance behaviors in the open field assay, whereas male mice showed evidence of these behaviors in the light/dark box assay. Female mice also showed increased reward-seeking in the sucrose preference test. Fentanyl-consuming mice of both sexes showed impaired cued fear extinction learning following fear conditioning. Our experiments show that long-term oral fentanyl consumption disrupts sleep rhythms, promotes anxiety-like behavior, increases motivation for reward, and disrupts fear extinction learning. This model could be useful to further study fentanyl withdrawal syndrome and anxiety-like phenotypes associated with protracted fentanyl withdrawal.

Escalation of intravenous fentanyl self-administration and assessment of withdrawal behavior in male and female mice

“…One prior study demonstrated that mice maintained stable levels of oral fentanyl consumption and showed a significant preference for the active lever associated with fentanyl in an operant chamber (63). Another study utilized a drinking in the dark paradigm to model oral fentanyl self-administration in mice, finding that mice consumed increasing amounts of fentanyl as the dose increased (37). Leonardo et al (33) successfully established fentanyl IVSA in mice, showing rapid acquisition and a dose-responsive curve.…”

“…Thus, it is critical to establish models of FUD in mice, including IVSA models. A previous study showed that mice are capable of initiating IVSA of fentanyl (33), and other mouse models have demonstrated withdrawal symptoms following oral or vapor self-administration (36)(37)(38).…”