Repetitive transcranial magnetic stimulation ameliorates cognitive dysfunction in Alzheimer’s disease by regulating microglia-neuron crosstalk via the CX3CL1-CX3CR1 axis

Microglia-neuron crosstalk plays a pivotal role in the pathogenesis of Alzheimer’s disease (AD), yet whether repetitive transcranial magnetic stimulation (rTMS) modulates this interaction is unclear. This study aims to investigate rTMS effects on cognitive impairment in AD and on microglia-neuron communication, and to elucidate the underlying molecular mechanisms. In vivo, four-month-old male APP/PS1 mice were randomly allocated to three groups: AD group, rTMS group (10 Hz, 14 days), and rTMS + AZD8797 (a CX3CR1 antagonist) group. Age-matched C57BL/6J mice served as the normal control group. The Morris water maze and Y-maze were used to evaluate spatial learning and memory abilities. Golgi staining was performed to observe the morphology and density of neuronal dendritic spines in the hippocampus. Immunofluorescence, Western blotting, and RT-qPCR were employed to detect microglial activation, neuroinflammation, neuron synaptic-related proteins, and key signaling proteins. In vitro, a Transwell co-culture system of BV2 and HT22 cells was established. Cells were divided into four groups: Control, Aβ-treated, Aβ + rTMS (10 Hz, 2 days), and Aβ + rTMS + AZD8797, to explore the role of rTMS in neuron-microglia crosstalk via the CX3CL1-CX3CR1 signaling pathway. rTMS improved cognitive function in APP/PS1 mice, reduced the levels of pro-inflammatory cytokines IL-1β and TNF-α, and increased the expression of synaptic proteins SYN and PSD-95, both in vivo and in vitro. Sholl analysis revealed that rTMS increased the branching index and total intersections of microglia and neurons in vivo. In vitro migration assays revealed that Aβ induced excessive migration of BV2 cells, which was attenuated by rTMS treatment. Furthermore, rTMS activated the CX3CL1-CX3CR1 signaling axis and facilitated microglia-neuron crosstalk. Notably, administration of the CX3CR1-specific antagonist AZD8797 abolished the beneficial effects of rTMS, including cognitive improvement, suppression of microglial overactivation, and enhancement of synaptic plasticity, both in vivo and in vitro. Our data suggest that rTMS modulates microglia-neuron crosstalk via the CX3CL1-CX3CR1 axis, thereby enhancing synaptic plasticity, reducing microglial overactivation, and ameliorating cognitive dysfunction in AD. This study provides a novel intercellular communication perspective on the mechanisms underlying rTMS therapy, offering insights into the development of non‑invasive therapeutic strategies for AD.