3.6 KM ameliorates chronic hyperalgesia and anxiety-like behavior caused by prolonged activation of BLA astrocytes
Since a single injection of CNO activated BLA astrocytes and instantly induced hyperalgesia, we next investigated whether prolonged activation of BLA astrocytes can induce persistent pain, mimicking the development of DNP. On this basis, we observed whether this pharmacogenetic-induced chronic hyperalgesia can also be alleviated by KM administration.
To this end, we induced repetitive pharmacogenetic activation of BLA astrocytes by daily i.p. injections of CNO for a week in naive rats with bilateral injection of gfaABC1D-M3-EGFP into the BLA and then treated them with KM (Fig. 8A). We found that daily CNO injections for one week induced a persistent decrease in the MWT in the rats expressing gfaABC1D-M3-EGFP, and KM reversed the mechanical allodynia induced by CNO injection in the rats transfected with gfaABC1D-M3-EGFP in the BLA within 1 hour, indicating that prolonged activation of BLA astrocytes was sufficient to induce the development of chronic pain-like hypersensitivity, which could be blocked by KM (7 mg/kg) (Fig. 8B).
We further determined whether this prolonged pharmacogenetic activation of BLA astrocytes was sufficient to induce anxiety-like behavior. We found that in the rats transfected with gfaABC1D-M3-EGFP, CNO injections made them less active in the central region in the OF and significantly reduced the number of entries and time spent in the open arms in the EPM; KM reversed these effects of CNO injections in the rats transfected with gfaABC1D-M3-EGFP (Fig. 8C-8H). The immunohistochemical results showed in the BLA of the rats that had BLA astrocytes continuously and specifically activated, the fluorescence intensity of GFAP was significantly enhanced, and the number of GFAP-positive cells was significantly increased. When combined with the KM intervention, BLA astrocytes were still, GFAP fluorescence intensity became weak, and the number of positive cells decreased (Fig. 8I-8K).
Collectively, these results indicated that prolonged activation of BLA astrocytes was sufficient to induce chronic hyperalgesia and anxiety-like behaviors that mimic STZ-induced DNP. After administration of KM, prolonged activation of BLA astrocytes induced a fully developed chronic hyperalgesia, and anxiety-like behaviors were significantly improved, which indicated that BLA astrocytes are one of the important targets of the anti-DNP effects of KM.