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Background  Repetitive transcranial magnetic stimulation (rTMS) is a noninvasive treatment for ischemic stroke. Astrocytes regulation has been suggested as one mechanism for rTMS effectiveness. But how rTMS regulates astrocytes remains largely undetermined. There were neurotoxic and neuroprotective phenotypes of astrocytes (also denoted as classically and alternatively activated astrocytes or A1 and A2 astrocytes) pertaining to pro- or anti-inflammatory gene expression. Pro-inflammatory or neurotoxic polarized astrocytes were induced during cerebral ischemic stroke. The present study aimed to investigate the effects of rTMS on astrocytic polarization during cerebral ischemic/reperfusion injury.    Methods  Three rTMS protocols were applied to primary astrocytes under normal and oxygen-glucose deprivation/reoxygenation (OGD/R) conditions. Cell survival, proliferation, and phenotypic changes were assessed after 2-day treatment. Astrocytes culture medium (ACM) from control, OGD/R, and OGD/R + rTMS groups were mixed with neuronal medium to culture neurons for 48 h and 7 days, in order to explore the influence on neuronal survival and synaptic plasticity. In vivo, rats were subjected to middle cerebral artery occlusion (MCAO), and received posterior orbital intravenous injection of ACM collected from different groups at reperfusion, and at 3 days post reperfusion. The apoptosis in the ischemic penumbra, infarct volumes, and the modified Neurological Severity Score (mNSS) were evaluated at 1 week after reperfusion, and cognitive functions were evaluated using the Morris Water Maze (MWM) tests. Finally, the 10 Hz rTMS was directly applied to MCAO rats to verify the rTMS effects on astrocytic polarization.    Results  Among these three frequencies, the 10 Hz protocol exerted the greatest potential to modulate astrocytic polarization after OGD/R injury. Classically activated and A1 markers were significantly inhibited by rTMS treatment. In OGD/R model, the concentration of pro-inflammatory mediator TNF-α decreased from 57.7 to 23.0 рg/mL, while anti-inflammatory mediator IL-10 increased from 99.0 to 555.1 рg/mL in the ACM after rTMS treatment. The ACM collected from rTMS-treated astrocytes significantly alleviated neuronal apoptosis induced by OGD/R injury, and promoted neuronal plasticity. In MCAO rat model, the ACM collected from rTMS treatment decreased neuronal apoptosis and infarct volumes, and improved cognitive functions. The neurotoxic astrocytes were simultaneously inhibited after rTMS treatment.    Conclusion  Inhibition of neurotoxic astrocytic polarization is a potential mechanism for the effectiveness of high-frequency rTMS in cerebral ischemic stroke.

journal of neuroinflammation

High-frequency repetitive transcranial magnetic stimulation improves functional recovery by inhibiting neurotoxic polarization of astrocytes in ischemic rats