Inhibition of autophagy in the amygdala ameliorates anxiety-like behaviors induced by morphine-protracted withdrawal in male mice

Morphine withdrawal triggers a range of negative affective states, wherein anxiety is typically common, significantly contributing to the morphine relapse. To date, the exact mechanism underlying morphine withdrawal–induced anxiety has remained unclear. Previous studies have proposed that autophagy is involved in the pathogenesis of morphine addiction and anxiety; however, the possible relationship between autophagy and morphine withdrawal–induced anxiety has not been explored before. In this study, we aimed to reveal the potential role of autophagy in anxiety-like behaviors elicited by protracted morphine withdrawal, and which brain region is involved. We established the model mice of anxiety by chronic intermittent escalating-dose morphine administration for 7 days and then withdrawing for 4 days. Anxious behaviors were detected using the open-field test and the elevated plus maze test. Western blot was performed to measure the change of autophagy-associated proteins (ATG5, Beclin-1, and LC3) in different brain regions. Our results showed that intraperitoneal injection of an autophagy inhibitor 3-methyladenine attenuated protracted morphine withdrawal–induced anxiety-like behaviors in male mice. Moreover, protracted morphine withdrawal predominantly promoted autophagy in the amygdala, rather than other related brain regions, suggesting the crucial involvement of the amygdala in autophagy-mediated anxiety after morphine withdrawal. We further validated that 3-methyladenine can effectively reduce autophagy-associated protein levels in the relevant brain region. These findings indicated that protracted morphine withdrawal–elicited autophagy in the amygdala contributes to the anxiety-like behaviors and may have implications for the future treatment of this disorder.