Central amygdalar PKCδ neurons mediate fentanyl withdrawal.

Aversion to opioid withdrawal is a significant barrier to achieving lasting opioid abstinence. The central amygdala (CeA), a key brain region for pain, threat-detection, autonomic engagement, and valence assignment, is active during opioid withdrawal. However, the role of molecularly distinct CeA neural populations in withdrawal remains underexplored. Here, we investigated the activity dynamics, brain-wide connectivity, and functional contribution of Protein Kinase C-delta (PKCδ)-expressing neurons in the CeA lateral capsule (CeLCPKCδ) during fentanyl withdrawal in mice. Mapping activity-dependent gene expression in CeLCPKCδ neurons revealed a highly withdrawal-active subregion in the anterior half of the CeA. Fiber photometry calcium imaging showed that opioid-naïve CeLCPKCδ neurons respond to salient noxious and startling stimuli. In fentanyl-dependent mice, naloxone-precipitated withdrawal increased spontaneous neural activity and enhanced responses to noxious stimuli. Chronic inhibition of CeLCPKCδ neurons throughout fentanyl exposure, via viral overexpression of the potassium channel Kir2.1, attenuated withdrawal symptoms in fentanyl-dependent mice. Lastly, we identified putative opioid-sensitive inputs to CeLCPKCδ neurons using rabies-mediated monosynaptic circuit tracing and color-switching tracers to map mu-opioid receptor-expressing inputs to the CeLC. Collectively, these findings suggest that the hyperactivity of CeLCPKCδ neurons underlies the somatic signs of fentanyl withdrawal, offering new insights into the amygdala cell-types and circuits involved in opioid dependence.