Distinct roles of general anesthesia activated CeA neurons in acute versus late phase of neuropathic pain
A previous study discovered a distinct population of GABAergic neurons in the central amygdala (CeA) that can be activated by general anesthesia (CeA-GA) and exert analgesic functions (Hua et al., 2020). To independently reproduce these prior findings and to investigate the electrophysiological properties of CeA-GA neurons, we first used 1.2 percent isoflurane to induce c-Fos activation in the mouse brain and validated the Fos expression by RNAscope in situ hybridization. Indeed, isoflurane induced robust Fos expression in CeA and these Fos+ CeA-GA neurons are GABAergic neurons (Vgat+). We next used Fos-TRAP2 method (different from the CANE method used in the prior study) to label CeA-GA neurons (tdTomato+). Our ex vivo electrophysiological recordings in brain slices revealed that compared to Fos-negative CeA neurons, CeA-GA neurons had significantly higher excitability and exhibited distinct patterns of action potentials. Chemogenetic activation of Fos-TRAPed CeA-GA neurons was effective at increasing pain thresholds in naive mice and mice with early-phase neuropathic pain 2 weeks after spared nerve injury (SNI). However, the same chemogenetic activation of CeA-GA neurons only had modest analgesia in the late phase of SNI at 8 weeks, although it was highly effective in reducing chronic pain-associated anxiety behaviors at this stage. We found that Fos-negative CeA neurons, but not CeA-GA neurons, exhibited increased excitability in the late-phase of SNI, suggesting that chronic pain causes a shift in the relative activity of the CeA microcircuit. Interestingly, Fos-negative neurons exhibited much higher expression of K/Cl cotransporter-2 (KCC2), and KCC2 expression was downregulated in the CeA in the late-phase of neuropathic pain. These results support the idea that targeting CeA-GA neurons may provide therapeutic benefits for pain relief and chronic pain-associated anxiety. Our findings also suggest distinct roles of CeA-GA neurons in regulating physiological pain, acute pain, and chronic pain with a possible involvement of KCC2.