OBJECTIVE

To investigate the effect of electroacupuncture (EA) on neurological function in rats with cerebral ischemia-reperfusion injury (CIRI) by regulating adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR)/Unc-51-like kinase 1 (ULK1) signaling pathway.

METHODS

Thirty-three male SD rats were randomly divided into a sham-operation group, a model group and an EA group, with 11 rats in each group. The right middle cerebral artery occlusion/reperfusion (MCAO/R) model was prepared by thread occlusion method in the model group and the EA group. In the sham-operation group, no thread was inserted after vascular separation. After the success of modeling, in the EA group, EA was applied to "Baihui" (GV 20) and "Zusanli" (ST 36) on the affected side, with disperse-dense wave, the frequency of 2 Hz/15 Hz, for 20 min, once a day. EA was delivered continuously for 3 days. On day 1 and day 3 of operation, the score of the modified neurological deficit scale (mNSS) was evaluated. After intervention completion, the cerebral infarction area was measured by the thiazolyl blue tetrazolium chloride (TTC) method. Nissl staining was used to observe the damage of cortical neurons on the ischemic side in each group. Using transmission electron microscopy, the ultrastructure of cortical neurons on the ischemic side was observed. With the immunofluorescence method adopted, the positive expression of the related protein 1 light chain 3 (LC3) and benzyl chloroform (Beclin-1) on the ischemic side was detected. The protein expression of p-AMPK, AMPK, p-mTOR, mTOR, pS757-ULK1, ULK1 and chelating ligand 1 (p62) in the ischemic cortex was detected using Western blot method.

RESULTS

① Compared with the sham-operation group, in the model group, the mNSS score increased (<0.01), the percentage of infarction area was increased (<0.01); the cortical neurons on the ischemic side were loosely distributed, with karyopyknosis and vacuolization, and the number of neurons was reduced (<0.01); the cells were swollen and ruptured, mitochondrial shrunk, electron density higher, and there were a large number of autophagic debris. The positive expression of LC3 and Beclin-1 was elevated (<0.01), and p-mTOR/mTOR, pS757-ULK1/ULK1 and the protein expression of p62 dropped (<0.01) in the ischemic cortex. ② Compared with the model group, in the EA group, the mNSS score was reduced (<0.05). The percentage of cerebral infarction area was descreased (<0.01); and the neurons were regularly distributed, the number of neurons increased (<0.01), the structure of mitochondria was clearer, the crest fracture alleviated, and the damage of neurons attenuated. The positive expression of LC3 and Beclin-1 was dropped (<0.01), and p-AMPK/AMPK reduced (<0.05), and p-mTOR/ mTOR, pS757-ULK1/ULK1 and the protein expression of p62 increased (<0.01) in the ischemic cortex.

CONCLUSION

EA at "Baihui" (GV 20) and "Zusanli" (ST 36) inhibits autophay, attenuates neurological deficit and cerebral pathological damage in CIRI rats to protect the nerves, which may be obtained by regulating AMPK/mTOR/ULK1 signaling pathway.