General anesthesia-activated neurons in the central amygdala mediate antinociception: Distinct roles in acute versus chronic phases of nerve injury.

BACKGROUND General anesthesia (GA), such as isoflurane, induces analgesia (loss of pain) and loss of consciousness through mechanisms that are not fully understood. A distinct population of GABAergic neurons has been recently identified in the central amygdala (CeA) that can be activated by general anesthesia (CeAGA) and exert antinociceptive functions. In this study, we aim to explore the underlying cellular mechanisms of CeAGA neurons across different phases of nerve injury-induced nociceptive sensitization in mice. METHODS This study used 107 mice, including 57 males and 50 females. We induced c-Fos activation in their brains using 1.2% isoflurane and validated Fos expression via RNAscope in situ hybridization. Unlike previous studies using the CANE method, we labeled CeAGA neurons (tdTomato+) with the Fos-TRAP2 method. We then performed ex vivo electrophysiological recordings to assess the properties of both Fos-positive/CeAGA neurons and Fos-negative CeA neurons. Using chemogenetic strategy to selectively activate the CeAGA neurons, we investigated pain-like behaviors and associated comorbidities in mice after spared nerve injury (SNI). RESULTS Isoflurane induced robust Fos expression in CeA GABAergic neurons. Electrophysiological recordings in brain slices revealed that compared to Fos-negative CeA neurons, CeAGA neurons had higher excitability and exhibited distinct patterns of action potentials. Chemogenetic activation of Fos-TRAPed CeAGA neurons increased nociceptive thresholds in naïve mice and in mice 2 weeks post-SNI, but demonstrated modest antinociception 8 weeks post-SNI. Finally, Fos-negative CeA neurons, but not CeAGA neurons, exhibited increased excitability in the chronic phase of SNI, which was correlated with a downregulation of K +-Cl - cotransporter-2 (KCC2) in the CeA (sham vs. SNI 8 weeks). CONCLUSIONS These results validate the antinociceptive power of CeAGA neurons using a different approach. Additionally, we highlight distinct roles of CeAGA neurons in governing physiological pain, acute pain, and the transition to chronic pain through KCC2 dysregulation.